BIOCHEMICAL REACTION

FOR THE
IDENTIFICATION OF
BACTERIA
PRESENTED BY: NEETU SING
(NITRATE
MODERATED BY: Dr. Shinu P
REDUCTION )
Department of Microbiology

INTRODUCTION

Biochemical

tests are used for the identification

of the organisms
Bacteria

produces

metabolic products by utilizing

various substrates with the help of enzymes .

The detection of the either these enzymes or the end

products are utilized in the principle of biochemical

tests.

NITRATE REDUCTION
TEST

MEDIA AND RAEGENT

Potassium nitrate,KNO3(nitrite_free)
1g

Peptone
5g

Distilled water
1litre

REAGENT - A
Sulphanilic acid

8g
Acetic acid(5N),30%
1lit

PRINCIPLE
nitrate reductase

NO3 + 2e +2H

NO2 + Sulfanilic acid

NO2 + H2O

diazonium salt

diazonium salt + alpha-naphthylamine

Red color complex

IF NO COLOR

TRUE NEGATIVE

FALSE NEGATIVE
( NO3)/
(NH3,N2,NO,N2O and

hydroxylamine )
Zinc Dust
NO3 to NO2

Red colour

True Negative
reaction

PROCEDURE
Inoculate nitrate broth(5ml) with
organism and incubate at 37C , 18-24
hours

Add 1ml each of reagent A and B to

broth

Observe for atleast 3 minutes for red

color to develop

RESUL
TS

Positive :- Red colour

Negative:- No colour

NITRATE POSITIVE ORGANISMS WITH

EXCEPTIONS

Enterobactericae except- Yerisinia

enterocolitica biovar
Vibrionacae except- Vibrio melschnikovii
Pseudomonacae except- Pseudomonas
diminuta
Pseu. paucimobilis
Alkaliginacae except-Alkaligens fecalis
Pasteurella canis

NITRATE NEGATIVE ORGANISMS WITH

EXCEPTIONS
Actinomycetaceae, except- Actinomyces
Archnia propionica
Listeria monocytogenes
Bordetella avium
Bordetella pertussis