Chemists have been making important contributions in the exciting field of biosensors. A biosensor is a device that recognizes a target molecule in a sample and interprets its concentration as an electrical signal via a suitable combination of a recognition system and a transducer. Suppose, for example, you want to know whether there is oxygen in a coal mine. You attach a microphone (the transducer) to a canary (the sensing device) and lower the (caged) canary into the mine. The response to oxygen is the singing of the canary, and it is converted into an electrical signal by the microphone. As long as the canary’s song can be heard, there is a certain level of oxygen present. Chemical biosensors are analogs of the canary.

Why would one want to develop biosensors? Suppose, for example, we could monitor insulin and glucose concentrations in the blood with a suitable biosensor. The electrical signals thus produced could be used to drive (or retard) a small implanted insulin pump, thus providing just the right amount of insulin for the amount of glucose present. Such a system would in effect serve as an artificial pancreas! You can probably think of other situations in which chemical biosensors would be immensely useful. The caveat in developing such devices is that they must be highly specific for the molecules they are designed to detect; that is, the devices must not respond to other molecules.

There have been a huge number of papers in recent years in this area, but this concept can be illustrated with one recent example. In this work, chemist Tony James and his coworker Christopher Cooper at the University of Birmingham developed a sensor for the amino sugar glucosamine. (1)

Compare the structure of glucosamine with that of glucose; the two differ only in the change of a hydroxy group (–OH) in glucose for an amino group (–NH₂) group in glucosamine. A key to the development of the biosensor is that the amino group of glucosamine, as shown above, can be protonated, and that the pK_a of the protonated amino group is very close to physiological pH (7.4), so that glucosamine is protonated to an appreciable extent under the conditions where we would be interested in detecting it.

The sensor molecule developed in the study is compound 1. Two similar compounds ( the controls 2 and 3) were essential to prove the assertion that compound 1 is specific for glucosamine.

In the sensor molecule 1, we have a fluorescent anthracene “light bulb” (blue) and a three-way switch consisting of two nitrogen atoms in different parts of the molecule (red and green). In order for the light bulb to go on, both switches must be thrown. How do these switches get thrown, and why are they thrown only by glucosamine? To answer these questions, let’s start by looking at the anthracene ring system.

The anthracene ring system (blue) is fluorescent. This means that when anthracene absorbs near-ultraviolet light of certain wavelengths, it re-emits light at another (greater) wavelength. This property arises from the extensive system of conjugated double bonds (double bonds alternating with single bonds) in the anthracene ring. It is this fluorescence that serves as the “light bulb” for the biosensor. In molecules 1, 2, and 3, the fluorescence is quenched (by a mechanism we won’t consider) by interaction of the electrons in the anthracene ring (the electrons involved in fluorescence) with the unshared electron pairs of the nitrogens. Compounds 2 and 3 each have one nitrogen “off” switch, compound 1 has two. A key to the use of this biosensor is that when these unshared pair of electrons on nitrogen are involved in other chemical interactions, the anthracene fluorescence is no longer quenched–in other words, the switch is thrown and fluorescence is observed.

Glucosamine is able to throw the two nitrogen “off” switches in molecule 1 by specfic interactions with two reactive groups in the molecule, a boronic acid and a crown ether, and when the two nitrogen switches are thrown the anthracene group can fluoresce. The proximal diol groups of glucosamine interact with the boronic acid group, and the amino group of glucosamine interacts with the crown ether group. The closely related sugar, glucose, is able to interact with the boronic acid group but, because it lacks an amino group, it does not react with the crown ether group. First, let’s take a look at interactions with the boronic acid group.

Boronic acids (compounds with the general structure RB(OH)₂, where R is any group) are often used by chemists to interact with certain diols –compounds that have two proximal OH groups. The interaction occurs by the following equation.

You will note that both glucose and glucosamine contain such proximal OH groups. When compounds 1 and 3 are treated with glucose in aqueous solution, the sugar reacts with the boronic acid group in both 1 and 3. In both cases, it turns out that this reaction strengthens the Lewis acid-base interaction between the boron and the nitrogen in the connector arm (red) so that the nitrogen in the connector arm is not available to quench the anthracene fluorescence. The result is that when compound 3 is used as the sensor, the anthracene “lights up” when the glucose reacts with the boronic acid group. However, compound 1 has a second nitrogen (green) that also quenches the anthracene fluorescence, and this is unaffected by the reaction of glucose with the boronic acid. Hence, compound 3 fluoresces in the presence of glucose, but not compound 1.

The second nitrogen switch in compound 1 is part of an “azacrown ether” (green). Azacrown ethers are known to bind ammonium ions (compounds of the form RNH₃+, where R is any group), and the interaction that causes the binding is hydrogen bonding of the protons in the ammonium ion with the unshared electron pairs in the azacrown ether. Note that this binding involves the unshared electrons of the nitrogen; note also that, at physiological pH, a significant fraction of glucosamine exists as an ammonium ion. (Can you calculate how much from its pK_a?)

Thus, when compound 1 binds glucosamine, the electron pairs of both nitrogens are “tied up,” fluorescence quenching by the both nitrogen unshared pairs is removed, and the anthracene lights up. As noted above, however, when compound 1 binds glucose, the unshared electron pair of the nitrogen in the crown ether group remains unaffiliated. The result: compound 1 fluoresces (”lights up”) only when glucosamine is bound. The structure of the complex between protonated glucosamine and compound 1 can be depicted as follows:

This biosensor only works over a limited range of pH; can you see why?

Related posts:

1. Chemical Damage due to Cigarette Smoke Freebasing Nicotine A second issue associated with tobacco smoking is its addictive nature, which is due to the presence of nicotine. Recent research from James F. Pankow...
2. Observing Single Molecules, Watching Molecules Wink When scientists make observations of matter, they in most cases observe large numbers of molecules and base their conclusions about a “typical” molecule on the...
3. Chemical Alteration of Cell Surfaces Cellular Velcro The molecules at the surfaces of cells govern or modulate many important biological phenomena, such as cell growth, cell differentiation (the determination of cell type),...
4. Benzyne Incarcerated in a Molecular Container Chemical Bondage Some molecules of great theoretical interest are so unstable that they cannot be isolated under normal, everyday conditions. Two molecules [...]...
5. Al77 A Missing Link In Cluster Chemistry Metal-metal bonds between transition metals are not uncommon and have been known for many years. Recently there has been more research into metal-metal bonds occurring...