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25 terms

Microbiology Lab- Oxidase, Catalase, Nitrate Reductase Tests

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Oxidase Purpose
to identify bacteria containing cytochrome C
Cytochrome C
an electron carrier in the electron transport chain
Phenylenediamine
artificial electron donor is used to reduce cytochrome C
colorless in reduced state
purple/blue when oxidized
Cytochrome C is present
phenylenediamine becomes oxidized
purple/blue color
Oxidase Procedure
collect bacteria from agar plate
place on blotter paper
apply reagent and observe color
Time for oxidase test
10 seconds
Purple/Blue
Oxidase Positive
Lack of color
oxidase negative
Catalase Test Purpose
to identify bacteria that produce the enzyme catalase
Catalase principle
most bacteria produce hydrogen peroxide, which is toxic to microbial cells. If the bacteria produces the enzyme catalase, which breaks down hydrogen peroxide into water and oxygen gas
Catalase procedure
place H202 on a colony
Gas bubbles
O2
bacterial colonies that produce catalase
catalase positive
No Gas Bubbles
catalast negative
Nitrase Reduction Test Purpose
to determine if an organism can reduce nitrate into nitrite using nitrate reductase
Nitrase Reductase outcome #1
NO3--->NO2
nitrate reduction positive
Nitrate Reductase outcome #2
NO3--->NO2--->other products (NH4, NO, N2, N2O)
nitrate reductase positive
Nitrase Reductase outcome #3
not metabolized
nitrate reduction -
genetics
presence of enzyme
Nitrase Reductase Procedure
10 drops of A and B
Red Color after Reagent A and B
positive result
nitrate reductase
prescense of nitrite
No color change after Reagent A and B
indicates no nitrate (NO2) present
add zinc powder
Nitrate C reagent
zinc powder
Red Color after reagent C
present of nitrate (NO3)
negative result
No color change after Reagent C
no nitrate (NO3) present
nitrates were reduced to nitrites which were then reduced to other products
positive result
API 20E
analytical profile index
20 biochemical tests
E- enterobacteriaceae