Blue bottle experiment

Reactions

History and general concept

The mechanism of the blue bottle experiment requires an understanding of rates and mechanisms of complex interacting chemical reactions. In complex chemical reactions, individual sub-reactions can occur simultaneously but at significantly different rates. These, in turn, can be affected by reagent concentration and temperature. In most cases, the overall reaction rate is determined by the fastest single component reaction. However, when some processes form intermediate molecules which then react in other processes to form the end product, the rate of the overall reaction is determined by the rate of the slowest reaction. In such circumstances the intermediate products are usually in a steady state at low concentrations because they are highly reactive. Equilibrium state requires that all reaction forward and backward mechanism happens at the same rate. Thus, the overall net reaction is determined by the sum of all the mechanism steps where the rate depends on the concentration and temperature. The blue bottle experiment illustrates this principle of interacting reactions with different rates.

The blue bottle experiment requires only three reagents: potassium hydroxide solution, dextrose solution, and dilute methylene blue solution. These reagents are added to a flask, mixed, and the flask is stoppered. The initial color of the solution is blue, but upon standing for a short interval it spontaneously fades to colorless, as the alkaline dextrose solution reduces the methylene blue to colorless leuco-methylene blue. Shaking the flask causes oxygen present in the head space air to dissolve in the solution and oxidize the leuco-methylene blue back to its colored form again. Another variation uses methylene blue in water, glucose, and caustic soda (NaOH). There are many versions of the experiment, however, unlike the classical version where dye is necessary to use as a catalyst for the reaction, the green and rapid versions undergo autoxidation even in the absence of the dye.

In the past, it was thought that the reaction occurred by the oxidation of an aldehyde group to a carboxylic acid under alkaline conditions. For instance, glucose would be oxidized to gluconate by oxygen. However, the experiment also works with compounds such as vitamin C and benzoin, which do not contain an aldehyde group. Thus, the reaction is actually the oxidation of an acyloin or related α-hydroxycarbonyl group, which is a structural feature of glucose, to a 1,2-diketone. The reduced redox dye (colorless state) is formed from oxidized redox dye (blue). The color-change that occurs in the blue bottle experiment has features of a clock reaction, in which a visible change in the concentration of one or more reagents suddenly occurs upon the exhaustion of a limiting reagent. For example, the limiting reactant, oxygen, is consumed by another reactant, benzoin, with the help of safranin as a catalyst. Once the limited amount of oxygen has been used up, the catalyst is unable to change forms, and as a result, the solution changes color.

Classical version

The aqueous solution in the classical reaction contains glucose, sodium hydroxide and methylene blue. In the first step an acyloin of glucose is formed. The next step is a redox reaction of the acyloin with methylene blue in which the glucose is oxidized to diketone in alkaline solution and methylene blue is reduced to colorless leucomethylene blue. If there is enough oxygen available (i.e., after shaking the bottle), leucomethylene blue is immediately re-oxidized to methylene blue and the blue color of the solution persists. However, when the solution is left to rest, the dissolved oxygen is gradually irreversibly consumed, and at the point where it has been completely exhausted, the glucose reduction of methylene blue proceeds unopposed and the color of the solution rapidly disappears. The reaction is first order in glucose, methylene blue and hydroxide ion and zero-order in oxygen. The process can be described as a pseudo first order reaction, and can be used to illustrate the changing concentrations of the reagents over the course of the reaction as the solution changes from blue back to colorless.

The final glucose oxidation products besides sodium gluconate have been identified as D-arabino-hexos-2-ulose (glucosone), the anion of D-arabinonate after splitting off of a formate anion and arabinonic acid.

Sustainable version

Wellman and Noble proposed a new formulation for the Blue Bottle experiment in which vitamin C serves as a reducing agent instead of glucose; the methylene blue and oxygen are still used. Copper is added as a catalyst for the reoxidation of leucomethylene blue to methylene blue. These modifications give an experiment that generates a smaller amount of waste that is less corrosive and easier to neutralize, along with changing the mixture color to a green-blue color, and therefore is an example of green chemistry modification.

Rapid version

The Chen autoxidation of benzoin had performed a similar experiment with respect to the classical and green versions. It was found that the traffic light and vanishing valentine experiments can become successful regardless of whether a sugar is added. One variation is more rapid, with the number of color change cycles do not last as long as the classical and green versions because the reactants are present in smaller amounts; also, the reducing agent for this experiment is benzoin, which is added to help increase the number of cycles in the solution. Moreover, the usable period in this experiment is quite short. Although the experiment is prepared overnight, the reducing agent can be added at any time to be able to observe the solution more.

Enzymatic version

Zhang, Tsitkov, and Hess from Columbia University proposed an enzymatic version of the "blue bottle experiment". They named it the "green bottle experiment", since the system is colored green and the reagents are safer than classical approaches. The experiment is performed in a clear glass vial containing two common enzymes (glucose oxidase and horseradish peroxidase), glucose, and 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (abbreviated as ABTS) in PBS buffer. A thin layer of oil is used to block the solution from the air. The solution initially turns green and then turns colorless with the depletion of dissolved oxygen. Shaking the solution introduces fresh oxygen and colors the solution green again until the oxygen is consumed.

This version relies on three enzymatic reactions. First, the glucose oxidase catalyzes the oxidation of glucose in the presence of oxygen and produces hydrogen peroxide. Second, the horseradish peroxidase utilizes the hydrogen peroxide to oxidize ABTS to its radical cationic form, ABTS^+•. As the dissolved oxygen is consumed in the solution, the third reaction occurs: glucose oxidase catalyzes the reduction of ABTS^+• back to ABTS in the presence of glucose. This system can also form beautiful patterns arising from reaction-driven Rayleigh–Bénard convection.