Litmus Milk Test – What Is It Used For And How It Works

The Litmus Test is used to help differentiate between microorganisms which are based on a number of metabolic reactions found within litmus milk.

Litmus Milk Test - What Is It Used For And How It Works

Some of these include the formation of a gas, as well as a clot, fermentation, digestion and reduction.

Milk is one of the best mediums when it comes to growth of certain microorganisms due to it containing the milk protein called casein, as well as vitamins, lactose (sugar), water and types of minerals.

The test uses Litmus milk because it can be used to distinguish the difference between bacteria species.

That is because the milk’s sugar (lactose), the milk’s protein (casein), and pH indicator (litmus) contained within the medium have the possibility of being metabolized by a number of species of bacteria.

With this in mind, let’s take a closer look at the Litmus Test.

What Is The Litmus Test Used For?

There are a couple of objectives when it comes to why the Litmus Test might be used. For example:

  • To seek out microorganisms that can enzymatically transform into different types of substrates of milk whose end products are varied metabolically.
  • To find out the ability of an organism to aid the metabolization of litmus milk.

The Principle Of The Litmus Milk

When it comes to what are the main substrates within the milk, the sugar and proteins (lactalbumin, lactoglobulin, and casein) are what makes it have the ability of transformation.

To help to distinguish the different changes metabolically within the milk, a pH indicator is needed. To do this, the oxidation reduction indicator litmus is included within the medium.

Litmus milk is great at this process because it can differentiate a plethora of microorganisms to see which ones metabolize within the substrates of milk. They do this by knowing which have a complementary enzymatic structure.

Depending on the biochemical result it can change the whole result.

To understand if any fermentation has occurred, the litmus milk turns a pink color due to the production of acid. If there has been a lot of acid produced within the litmus, then casein becomes thicker, and the milk becomes solid.

Other scenarios that might happen are:

  • Litmus can become reduced which causes the medium to be colorless at the bottom in a tube.
  • Hydrolyze casein (bacteria) can cause the milk to look like the color of straw.
  • Organisms can also make the curd of the milk begin to shrink. In doing so, whey is then formed on the surface.

Different Uses Of The Litmus Milk

Different Uses Of The Litmus Milk

So, what are the different uses when it comes to using the Litmus Test? Some of the examples of why the Litmus Test is used are:

  • The media used for it can be used to grow a type of bacteria which is formed for lactic acid. It can identify and also differentiate for the use of both lactic acid and enterococcus.
  • It is also used within clostridium to differentiate the members.
  • Also, the litmus milk can be used for enterobacteriaceae to again differentiate the different members. It differentiates it from ‘gram-negative’ bacilli.

Are There Limitations Of Using the Litmus Test?

There are some limitations to using the Litmus Test. Some of these include:

  • The results aren’t very specific and this means that you will need to do further testing to fully identify a particular microorganism.
  • The results can also be unreliable due to the fact the results are very varied.
  • When it comes to writing results, usually both a curd and clot that has been formed is recorded as a ‘clot’. This means the results are not specific enough.

Method Of Using Litmus Test

Here is a method of using the Litmus Test which includes the media:

  • 100g Skim milk which is powdered
  • 0.5g of litmus
  • 0.5g of Sodium sulphite with a pH of 6.8


  1. Inject up to 4 drops each within a cultured broth.
  2. Leave to sit at around 95 to 97 degrees Fahrenheit for up to 7 days.
  3. During this period an acid and alkaline reaction should be looked for and recorded:

Acid Reaction

The litmus will have turned pink. This might indicate peptonization or a rennet clot.

Alkaline Reaction

The litmus will have turned blue. This might indicate that there has been an indicator reduction, or that there is a clot within the acid.

Despite the ones listed above, there is a good chance that a number of different reactions might have happened. So, let’s take a look at them.

Results Of The Test

Here are the main things that a person might look for during the Litmus Test:

Positive Outcome

  • Production of gas – bubbles have formed within a solidified milk
  • Digestion has occurred – there has been a split within the milk which has caused a clot to appear that contains a gray, water like fluid, along with a pink colored clot.
  • Rennet curd – the milk has turned into a soft like curd which has been followed with peptonization.
  • Acid curd – this causes the curd to be hard and for a clear whey to form on the top.
  • Reduction – this causes it to turn white.
  • Acid pH – the result is a pink which turns into a red type color.
  • Alkaline pH – the result will be a purple to a blue type color.

Negative Outcome

Everything has stayed the exact same with both the consistency of the milk and the overall color.

Final Thoughts

The litmus test has its uses, and it can also produce various results. Whilst for something specific it can be useful, it can also prove to be unreliable due to the method concluding in answers that cannot be defined.

To put it simply, however, the Litmus Test is used to differentiate between a number of different microorganisms which have different metabolic reactions that can be found within litmus milk.

Using this method can help to grow bacteria such as lactic acid. Hopefully this article has helped you to understand the Litmus Test more clearly!

Jennifer Dawkins

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