Voges-Proskauer (VP) Test – Principle, Reagents, Procedure And Result

The Voges-Proskauer test is a type of test used in microbiology to detect the presence of acetoin in a bacterial broth culture.

First developed way back in 1898 by bacteriologists at the Institute for Infectious Diseases, this test is one of the oldest in microbiology that is used to find out if a bacterial organism can produce acetylmethyl carbinol during the glucose fermentation. 

So, this makes the Voges Proskauer test a pretty common one that most microbiologists try out during their studies or career.

Voges-Proskauer (VP) Test - Principle, Reagents, Procedure And Result

In fact, it is part of the IMViC series of tests which is used to classify enteric bacteria.

Here we have pulled together a guide for the entire test including its principle, what media is used in the test, a guide through the steps of the method and what the results mean.

So, check out the information below for everything you need to know about the Voges-Proskauer test! 

Principle Of The Voges-Proskauer Test

The whole point of the Voges Proskauer test is to find out which organisms (usually bacteria) can produce neutral end products during the fermentation process of glucose.

These neutral end products include acetoin as well as 2,3-butanediol. 

As all members of the enterobacteriaceae family of gram-negative bacteria can ferment glucose and convert it to pyruvic acid, this test can also be used to determine which pathway they take when it comes to metabolizing pyruvic acid.

To determine this, a pH indicator is used in the broth medium. If the organism used in the Voges-Proskauer test produced a lot of acids during the fermentation process of the glucose.

Then, the pH indicator will change color to indicate this.

However, if the organism metabolizes the other fermentation products and produces acetoin, this will keep the pH level of the broth at neutral and thus, the pH indicator will not change color. 

So, this test utilizes the pH indicator methyl red to indicate whether or not the organisms produce acetoin.

This further helps with the classification process of bacteria and helps microbiologists understand the full potential of each species. 

Reagens Of The Voges-Proskauer Test

The main media used for Voges-Proskauer test includes an MRVP broth that has a pH of 6.9. It is usually made up of 7.0 gm of buffered peptone, 5.0 gm of glucose, and 5.0 gm of dipotassium phosphate. 

The two reagents used for the Voges-Proskauer Reagent are A and B.

Reagent A contains 50 gm of Alpha-Naphthol, 5% and 1000 ml of Absolute Ethanol. Reagent B however contains 400 gm of Potassium Hydroxide and 1000 ml of deionized water. 

These two reagents are very important for the procedure of the Voges-Proskauer test, so let’s go through the steps one by one so you can see how it works. 

Procedure Of Voges-Proskauer Test

Before you start the procedure, you must have first allowed the medium to settle at room temperature.

You will also have isolated the organisms you are using, taking them from a pure culture that has had 18 to 24 hours to isolate. 

Once your organisms and medium are ready, you should inoculate the medium then leave it to incubate at 98.6 degrees Fahrenheit (or 37 degrees Celsius) for 24 hours.

The medium should also be left to incubate aerobically. 

Once the incubation period is over, you will need to aliquot (or draw out a portion) 2 ml of the broth into a clean tube. 

Now, you will need to incubate the rest of the broth again for another 24 hours in the same conditions. 

Once this is one, you need to add 6 drops of reagent A to the broth and mix well. Then, you need to add only 2 drops of reagent B and mix well.

It is important that you mix the broth well so it has a chance to properly introduce air into the broth for the reaction to take place. 

After this, you should be able to see some changes to the color of your broth.

Leave it for 30 minutes, shaking vigorously and then to continue introducing air to the broth to help activate the reaction (if there is one). 

After this 30 minute period, you should be able to record your observations as results. 

Interpreting Your Results

A positive result in the Voges-Proskauer test is when you see the color of the medium change from a beige-yellow color to a more pink-red shade.

This will more likely occur at the surface of the medium instead of affecting the whole tube, but this change in color indicates that acids are present.

Even if the change is only slight and the color changes to a rust color, this still counts as a positive result even though it is a sign of a weak reaction.

Examples of some Voges-Proskauer positive organisms include enterobacter, listeria, vibrio eltor, and all viridans group streptococci except for streptococcus vestibularis.  

Alternatively, a yellow-brown color or no change in color of your medium is a negative result, meaning that there are no acids present.

Some organisms you can expect to test negative in the Voges-Proskauer test include streptococcus mitis, shigella, salmonella, and edwardsiella.  


And that is everything you need to know about the Voges-Proskauer (VP) test!

If you are ever in doubt about the principle of this test and its use, as well as the steps of the method, the reagents you should use, and what your results mean, then you can just use this handy guide as a reference. 

Of course, the Voges-Proskauer test is not without its limitations as it has a long incubation process and failing to add reagents will result in weak or false negative results, but the Voges-Proskauer test is a vital one for all microbiologists to know.

It is one of the main four tests used to classify bacteria and other organisms, so check out the above guide again so you can understand this test in as much detail as possible!

Jennifer Dawkins