Activity No.

3 Laboratory Sheet

MEDIA DISPENSING

Name: Buenaflor, Andrea Mae P. Date Submitted: 03-10-23

Enorio, Isabelle D.
Reyes, Shylene Kaye A.
Rivera, NJ Aleah F.
Yap, Fhardina D.

I. Introduction

Microorganisms nearly always exist in nature as mixed populations of

numerous, significantly diverse varieties. But still, isolation in pure culture is
required before the preponderance of an organism's qualities and traits may be
discovered. One strain of the microorganism makes up a pure culture.
Microorganisms need nourishment and a good environment to thrive and reproduce,
just like all other living things. Typically, a culture medium that contains necessary
nutrients and creates an appropriate environment (e.g., a proper pH) is used to grow
microbes containing necessary nutrients and creates an appropriate environment
(e.g., a proper pH) is used to grow microbes. Overall, microbes are cultured on agar
or in broth, a liquid medium.
A number of the cells, the inoculum, are transferred, or inoculated, into or
onto the medium in order to grow a microbial culture in a sterilized medium while
taking extreme precautions to preserve the purity of the culture. The term "sterile
technique" refers to the procedures employed in the microbiology lab to prevent
contamination of pure cultures.

II. Objectives
• to introduce the two different forms of culture media, nutritional broth and
nutritional agar.
• to acquire the ability to isolate a pure culture.

III. Requirements:

A. Materials: Erlenmeyer Flask, Petri Dish, Hot Plate, Test Tubes, Beaker,
10mL syringe, alcohol lamp
B. Chemicals: Nutrient Broth
IV. Procedure

Preparation of Solid Media

Generally, a solidifying agent is added to a broth medium to create a solid
medium. Agar, a material derived from sea algae and available in dried, pure form,
is the most commonly used solidifying agent. Although the physical characteristics
of various agars differ drastically, the typical melting point ranges from 97 to 100°C.
Thus, during heat sterilization, solid agar can be introduced to liquid culture media
and melted. This liquid medium can be put into test tubes, bottles, or petri dishes
after sterilization. This agar-containing mixture cools to about 42 °C before
solidifying.
1. Place five petri plates side up in front.
2. Lit the alcohol lamp
3. In order to complete pouring the plates before the agar solidifies, it must be done
quickly.
4. Bottle cap should be loosened but not taken off. Once the cap has been completely
loosened, work near the alcohol lamp to maintain the sterility of the procedure.
5. Lightly pass the lip of the bottle in front of the alcohol lamp to burn off any
adhering dust and to also to set up a negative air current. The lip of the bottle would
be sterile from the autoclaving procedure.
6. Using your left or right hand, gently lift the cover of a petri dish so that you can
pour some of the agar medium into the bottom of the dish.
7. Add just enough medium onto the bottom of the petri dish to cover it. (With 100
mL of medium, you should have no trouble pouring four plates, possibly five.)
8. Slide the poured plates slowly and carefully to the back of the benchtop, where
they may cool and harden without being disturbed. For at least 20 minutes, don't
relocate these plates for the agar to solidify. Don't forget to label your petri plates.

V. Conclusion
We concluded that there are essential things that we should take note of during
the dispensing of media. In dispensing the agar, it must be done close to the alcohol
lamp to maintain the sterility of the procedure. Then always remember to pass lightly
the materials in the alcohol lamp before and after opening. Also, when pouring tha
agar to the petri dish, time should be observed as the agar is solidifying. This
procedure was repeated due to the agar being contaminated. On the other hand, in
dispensing the broth, accuracy of measurements and sterility must be noted. It has
been realized that the procedures must be properly done in order to produce the
expected results. Each measurement of the chemical and material are significant in
 obtaining the objectives of this laboratory activity, which are mainly to introduce
both nutritional agar and broth and to isolate a pure culture.

VI. Documentation
a. Agar Dispensing

b. Broth Dispensing
VII. . References
• Mettler-Toledo International Inc. all rights reserved. (2022, November 16).
Culture Media Preparation. Mettler-Toledo International Inc. All Rights
Reserved.
https://www.mt.com/ph/en/home/applications/Laboratory_weighing/Culture-
Media-Preparation.html
• Aryal, S. (2022, August 10). Nutrient Agar: Composition, Preparation and Uses.
Microbiology Info.com. https://microbiologyinfo.com/nutrient-agar-
composition-preparation-and-
uses/#:~:text=Water%20is%20essential%20for%20the,various%20nutrients%20
can%20be%20transported.