LIPID HYDROLYSIS
The enzyme lipase is excreted out of the cells (an exoenzyme) into the surrounding media,
catalyzing the breakdown of tributyrin, a vegetable oil, into fatty acids which can then be
taken up by the organism. This reaction causes the agar, normally a light blue opaque color
to clear around the growth area.
OBJECTIVES:
Identify the presence of the enzyme lipase.
MATERIALS NEEDED:
1 tributyrin agar plate with methylene blue dye per unknown
THE PROCEDURE:
1.
2.
3.
4.
Run this test using your unknown bacterium.
Inoculate the organisms on the plate either a straight line or a zig-zag.
Incubate at 25 or 37 degrees C.
Record the results of your bacterial unknown in your journal.
INTERPRETATION:
Tributyrin oil forms an opaque
suspension in the agar. When an
organism produces lipase and breaks
down the tributyrin, a clear halo
surrounds the areas where the lipaseproducing organism has grown. Hold the
plate up to the light to see the zones
well.
Record positive reactions as weak + or
strong +.
There is already an indicator, methylene
blue, incorporated into the agar. A zone
of clearing around the growth area
identifies the presence of the enzyme
lipase.
QUESTIONS:
1. What is the substrate in this medium?
2. Name the indicator in this medium.
Fall 2011 - Jackie Reynolds, Richland College, BIOL 2421
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