Culture media are the nutrient mediums for the cultivation of microorganisms. Culture media provide different nutrients for the growth and survival of bacteria. These culture media vary in physical consistency, use, and chemical composition. Nutrient agar and nutrient broth are microbial culture media used for cultivating bacteria. Nutrient agar and nutrient broth are similar in composition except for agar. In nutrient broth, there is no solidifying agent, whereas nutrient agar media composition contains solidifying agent agar. Microorganisms can be grown either on solid or liquid media as per need.
Nutrient agar Introduction:
Nutrient agar is a common medium for cultivating and isolating a wide range of microorganisms. It is the most often used medium in all laboratories for the enumeration of microorganisms. Nutrient agar is very useful for studying the colony morphology of bacteria. Routine microbiological work like isolation and culturing, microbial enumeration, and preservation of pure cultures needs nutrient agar. The nutrient agar medium composition is made in such a way that it supports the growth of most non-fastidious bacteria. bacteria. In this article, we will see how to make nutrient agar media to grow bacteria in the laboratory.
The focus of this article will be on
1) Nutrient agar media composition and uses
2) How to prepare nutrient agar media for microbiological work.
Nutrient agar is a solid medium in microbiology to grow microbes, mainly bacteria. As per physical consistency nutrient agar is solid media.
If you categorize nutrient agar by use, it falls into the category of simple media. It is a general-purpose medium that supports the growth of most bacteria. Many non-fastidious microorganisms grow easily on nutrient agar.
Nutrient agar has ingredients such as peptone and beef extract, which provide individual chemical components like nitrogen, amino acids, organic carbon, and inorganic salt. Since we do not know the exact amount of each individual chemical component present in peptone and beef extract, it is also a complex media.
Watch following video on what is nutrient agar | Nutrient agar microbiology
Nutrient Agar media composition:
The main ingredients in nutrient broth are peptone, beef extract, and sodium chloride and agar as solidifying agent.
Peptone:
Peptone is a protein which is partly digested. It is a complex mixture of amino acids and polypeptides. Peptone is a rich source of organic nitrogen. Peptone is the product of protein hydrolysis. The proteins for making peptone can be from either plant or animal proteins, such as those found in meat, milk, soybeans, yeast, and gelatin. First, the proteins are hydrolyzed using various techniques such as acid hydrolysis, enzymatic hydrolysis, microbiological hydrolysis, and autolysis. As a result, a combination of amino acids and polypeptides of varying lengths are formed. Peptone has a high concentration of water-soluble minerals and micronutrients. Peptones provides nutrients that allow most microorganisms to flourish.
Beef extract:
It is a derivative of beef extract. It contains bovine tissue extract in dry form. Beef extract provides nutrition such as organic carbon, nitrogen, vitamins, and inorganic salts.
Agar:
Agar acts as the solidifying agent in nutrient media.
Sodium Chloride (NaCl):
Sodium chloride (NaCl) maintains osmotic equilibrium. Basically, it preserves osmotic balance thereby inhibiting bacterial cell lysis. Therefore, sodium chloride is an important ingredient in nutrient broth
If you are using a ready-made commercial nutrient broth bottle, then it is homogenous free-flowing cream to yellow powder. It is present in dehydrated form. You can directly suspend the required quantity of powder in distilled water as per the instructions given on the label.
Apparatus for nutrient agar media preparation:
1. Clean Erlenmeyer flask or conical flask
2. Spatula
3. Weighing Boat
4. Weighing Balance
5. Peptone
6. Beef extract
7. Agar powder
8. Sodium chloride
9. Measuring Cylinder
10. Distilled Water
11. 1N HCL
12. 1N NaOH
13. pH meter
How to prepare 1000 ml of nutrient broth media composition?
Watch the following video on how to make nutrient agar plates by the preparation of nutrient agar media.
The following table shows the quantity of reagents required for the preparation of nutrient agar in 100 ml and 1000 ml.
1. Firstly, take a clean conical flask. (Generally, if you are preparing 100 ml of nutrient agar, then you should use a 500 ml conical flask you are preparing a 1000 ml nutrient agar, then you should take a 2000 ml conical flask.)
2. Weigh the peptone, beef extract, NaCl, agar as per above table and add these weighed ingredients to the conical flask.
3. Pour 500 ml of distilled water into the flask.
4. Gently agitate and mix the contents well. Heat if it is necessary so as to dissolve the ingredients
5. Now add the remaining 500 ml distilled water and shake well to make homogenous mixture
6. Now check the pH of the broth. Adjust the pH to 7.2 ± 0.2. In case you observe the pH change, then you can adjust accordingly by adding 1N HCL or 1N NaOH.
7. Now you can either pour this broth into test tubes or keep as it is in the conical flask.
8. Cover the conical flask or test tube mouth with a cotton plug and wrap a paper on it.
9. Keep the flask with broth in autoclave and autoclave at 121 o C at 15 psi pressure for 15 minutes.
10. Lastly, after autoclave let the flask with nutrient broth to cool completely.
11. Store this sterile nutrient agar medium at cool temperature in clean dust free environment.
12. Before use, always open the flask for inoculation only inside the laminar in order to avoid contamination.
Uses nutrient agar:
1. Nutrient agar supports the growth of a wide variety of bacteria.
2. The bacteria streaking, spread plate method, and pour plate method need nutrient agar.
3. Purity checking and isolation require nutrient agar.
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