To study the ability of microorganisms to hydrolyze gelatin with the proteolytic
The ability of microorganisms to hydrolyze gelatin is commonly taken as evidence
that the organism can hydrolyze protein in general. But there are exceptions.
Microorganisms vary from species to species with regard to their ability to
hydrolyze protein. This feature characterizes some species.
Gelatin is a protein obtained by the hydrolysis of the collagen compound
of connective tissues of animals. It is convenient as a substrate for proteolytic
enzymes in microorganisms.
Gelatin is used in the media from the experiment, which is liquid at room
temperature and solidifies at –4°C. If the gelatin has been hydrolyzed by the
action of the organism, the media will remain liquid.
- Nutrient gelatin media
- Test organism
- Test tubes
- Inoculation loop
- Preparation of Nutrient Gelatin Media
- Peptone - 5 g
- Gelatin - 20 g
- Beef Extract - 3 g
- Sodium Chloride - 5 g
- Distilled Water - 1000 mL
- pH - 7.2.
- Media is prepared according to the above composition.
- It is sterilized at 121°C for 15 minutes at 15 lb/inch square and poured
into presterilized tubes.
- Tubes were allowed to cool and then inoculated with test organisms. One
inoculated tube is used as a control.
- Tubes were incubated for 24 hrs and observed for liquefaction of gelatin
after keeping in ice for half an hour.
Gelatin is an incomplete protein, lacking many amino acids, such as tryptophan.
When collagen is heated and hydrolyzed, denatured protein gelatin is obtained.
Collagen accounts for 90%–95% of organic matter in the cell. It is the most
important protein, rich in amino acids. Microorganism-like bacteria can use
gelatin only if they are supplemented with other proteins. Bacteria produce the
gelatin-hydrolyzing enzyme, gelatinase. Since gelatin is a good solidifying agent
at low temperatures, its property of solidification can be used to distinguish
between gelatin-hydrolyzing and nonhydrolyzing agent. Most of the enterobacteriaceae
members are gelatin-hydrolysis-test-negative. Bacteria like Vibrio,
Bacillus, and Pseudomonas are gelatin-positive.