Amperometry in

chemistry Chemistry is the scientific study of the properties and behavior of matter. It is a physical science within the natural sciences that studies the chemical elements that make up matter and chemical compound, compounds made of atoms, molecules a ...

is the detection of ions in a solution based on electric current or changes in electric current. Amperometry is used in

electrophysiology Electrophysiology (from ee the Electron#Etymology, etymology of "electron" ; and ) is the branch of physiology that studies the electrical properties of biological cell (biology), cells and tissues. It involves measurements of voltage change ...

to study vesicle release events using a carbon fiber electrode. Unlike

patch clamp The patch clamp technique is a laboratory technique in electrophysiology used to study ionic currents in individual Cell isolation, isolated living cells, tissue sections, or patches of cell membrane. The technique is especially useful in the st ...

techniques, the

electrode An electrode is an electrical conductor used to make contact with a nonmetallic part of a circuit (e.g. a semiconductor, an electrolyte, a vacuum or a gas). In electrochemical cells, electrodes are essential parts that can consist of a varie ...

used for amperometry is not inserted into or attached to the cell but brought nearby of the cell. The measurements from the electrode originate from an oxidizing reaction of a vesicle cargo released into the medium. Another technique used to measure vesicle release is capacitive measurements. It is the electric current measured between a pair of electrodes. The measured current is directly proportional to the concentration of the analyte. Example: Blood glucose monitor Carbon is used as a working electrode which is coated with mediator and glucose oxidase. Ag/AgCl is used as the reference electrode. The enzyme oxidase catalyzes the reaction of glucose with oxygen. Hydrogen peroxide concentration is measured by oxidation which occurs at +0.6V. H₂O₂——>O₂+2H+ + 2e- Current is directly proportional to H₂O₂ concentration this in turn is directly proportional to glucose concentration. If O₂ is low, complete conversion is inhibited. Hence mediator ferrocene is introduced. Now, current is directly proportional to the concentration of ferrocene which in turn is directly proportional to glucose concentration.

History

Electrochemical Electrochemistry is the branch of physical chemistry concerned with the relationship between electrical potential difference and identifiable chemical change. These reactions involve electrons moving via an electronically conducting phase (typi ...

or amperometric detection as it was first used in ion chromatography was single-potential or DC amperometry, useful for certain electrochemically active ions such as cyanide, sulfite, and iodide. The development of pulsed amperometric detection (PAD) for analytes that fouled

surfaces when detected eventually helped create a new category of ion chromatography for the determination of

carbohydrates A carbohydrate () is a biomolecule composed of carbon (C), hydrogen (H), and oxygen (O) atoms. The typical hydrogen-to-oxygen atomic ratio is 2:1, analogous to that of water, and is represented by the empirical formula (where ''m'' and ''n'' ma ...

. Another advancement, known as integrated amperometry, has increased the sensitivity for other electrochemically active species, such as amines and many compounds that contain reduced

sulfur Sulfur ( American spelling and the preferred IUPAC name) or sulphur ( Commonwealth spelling) is a chemical element; it has symbol S and atomic number 16. It is abundant, multivalent and nonmetallic. Under normal conditions, sulfur atoms ...

groups, that are sometimes weakly detected by PAD. It was established that

neurotransmitter A neurotransmitter is a signaling molecule secreted by a neuron to affect another cell across a Chemical synapse, synapse. The cell receiving the signal, or target cell, may be another neuron, but could also be a gland or muscle cell. Neurotra ...

s could be electrochemically detected by placing a carbon electrode into tissue and recording the current from oxidizing neurotransmitters. One of the first measurements was made using an implanted carbon fiber electrode in the neostriatum of rats. Further work was done in chromaffin cells to investigate catecholamine release from large dense core vesicles.

Detection methods

Single-potential amperometry

Any analyte that can be oxidized or reduced is a candidate for amperometric detection. The simplest form of amperometric detection is single-potential, or

direct current Direct current (DC) is one-directional electric current, flow of electric charge. An electrochemical cell is a prime example of DC power. Direct current may flow through a conductor (material), conductor such as a wire, but can also flow throug ...

(DC), amperometry. A voltage (potential) is applied between two electrodes positioned in the column effluent. The measured current changes as an electroactive analyte is oxidized at the anode or reduced at the cathode. Single-potential amperometry has been used to detect weak acid anions, such as

cyanide In chemistry, cyanide () is an inorganic chemical compound that contains a functional group. This group, known as the cyano group, consists of a carbon atom triple-bonded to a nitrogen atom. Ionic cyanides contain the cyanide anion . This a ...

and

sulfide Sulfide (also sulphide in British English) is an inorganic anion of sulfur with the chemical formula S2− or a compound containing one or more S2− ions. Solutions of sulfide salts are corrosive. ''Sulfide'' also refers to large families o ...

, which are problematic by conductometric methods. Another, possibly more important advantage of amperometry over other detection methods for these and other ions, such as iodide, sulfite, and

hydrazine Hydrazine is an inorganic compound with the chemical formula . It is a simple pnictogen hydride, and is a colourless flammable liquid with an ammonia-like odour. Hydrazine is highly hazardous unless handled in solution as, for example, hydraz ...

, is specificity. The applied potential can be adjusted to maximize the response for the analyte of interest while minimizing the response for interfering analytes

Pulsed amperometry (pulsed amperometric detection, PAD)

An extension of single-potential amperometry is pulsed amperometry, most commonly used for analytes that tend to foul electrodes. Analytes that foul electrodes reduce the signal with each analysis and necessitate cleaning of the electrode. In pulsed amperometric detection (PAD), a working potential is applied for a short time (usually a few hundred milliseconds), followed by higher or lower potentials that are used for cleaning the electrode. The current is measured only while the working potential is applied, then sequential current measurements are processed by the detector to produce a smooth output. PAD is most often used for detection of carbohydrates after an anion exchange separation, but further development of related techniques show promise for amines, reduced sulfur species, and other electroactive compounds.

Principle

In order to record vesicle fusion, a carbon fiber electrode is brought close to the cell. The electrode is held at a positive potential, and when the cargo from a fused vesicle is near the electrode, oxidation of the cargo transfers electrons to the electrode. This causes a spike, the size of which can be used to estimate the number of vesicles, and the frequency gives information about the release probability.

References

{{reflist Electrophysiology