Phenylalanine Deamination test procedure and result interpretation

Phenylalanine Deamination test MLTGEEKS

The ability of Proteus species to be able to deaminate phenylalanine to phenylpyruvic acid  using the enzyme
Phenylalanine ammonia-lyase (pal enzyme) is an important and useful reaction in the differentiation of members of the family of Enterobacteriaceae.This biochemical tests for identification of bacteria  is used for the differentiation of Proteus and Providencia group of organisms from other members of Enterobacteriaceae on the basis of their ability to form phenyl pyruvic acid from phenylalanine 

Principle of Phenylalanine Agar

  • The yeast extract in the medium supports the growth of the organisms.
  • Sodium chloride maintains osmotic equilibrium.
  • The phenylalanine serves as the substrate for enzymes, which are able to deaminate it in order to form phenylpyruvic acid.

Composition of Phenylalanine Agar slant

The following ingredients are used for the composition Phenylalanine agar slant

IngredientsGrams / Litre
Yeast extract3.000
Sodium chloride5.000
DL-Phenylalanine2.000
Disodium phosphate1.000
Agar15.000
Final pH at temperature 25°C7.3±0.2

How to Prepare Phenylalanine Agar slant

  1. Start by suspending 26 grams in 1000 ml distilled water.
  2. Heat to boiling in order to dissolve the medium completely.
  3. Dispense in tubes and sterilize by autoclaving at 15 lbs pressure (121°C) for 15 minutes.
  4. Allow the tubed medium to cool in a slanting position.

Appearance of Medium after preparation

It’s appear Light amber coloured slightly opalescent gel forms in tubes as slants

Phenylalanine Agar slant
Phenylalanine Agar slant

phenylalanine deaminase test reagent

  • 10% ferric chloride solution
  • 0.1N HCl

Phenylalanine Test Procedure

  1. inoculate the slant surface with plenty of inoculum then incubate it for 12-16 hours
  2. After incubation, now add 0.2 ml or 5 drops of 10% ferric chloride solution and 5 drops of 0.1N HCl so that the solution floods all over the growth.
  3. Gently shake the tube and observe for color change
  4. In a positive reaction, any phenylpyruvic acid present will react with the ferric salt in the reagent to give a green color
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phenylalanine deaminase test results and interpretation

Interpretation of  the results is done within 5 minutes upon addition of reagent as the green colour fades quickly

Positivegreen color develops within five minutes after addition of reagents
Negativeno color change (negative reaction)
1-Uninoculated-2-Negative-3-Positive
1-Uninoculated-2-Negative-3-Positive

Culture observation and Phenylalanine deamination

OrganismPhenylalanine Deaminase
Escherichia coli Negative : no change after addition of 10% Ferric chloride
Enterobacter aerogenesNegative : no change after addition of 10% Ferric chloride
Proteus vulgarisPositive : green colouration after addition of 10% Ferric chloride
Proteus mirabilis Positive : green colouration after addition of 10% Ferric chloride.

phenylalanine deaminase test positive bacteria

  • Providencia alcalifaciens
  • Proteus vulgaris
  • Proteus mirabilis

Reference

  • Singer J. and Volcani B. E., 1955, J. Bacteriol., 69:303.
  • Henrikson S. D., 1950, J. Bacteriol., 60:225.
  • Buttiaux R., Osteux R., Fresnoy R. and Moriamez J., 1954, Ann. Inst. Pasteur Lille., 87:375.
  • MacFaddin J. F., 1985, Media for Isolation Cultivation-Identification-Maintenance of Medical Bacteria, Vol. 1, Williams & Wilkins, Baltimore, Md.
  • Ewing W. H., Davis B. R. and Reavis R. W., 1957, Public Health Lab., 15:153.

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About the Author: Arthur Westmann

DEFFE ARTHUR (AMOEBAMANN) is the founder and author of MLTGEEKS and MLTEXPO.He’s from Cameroon and is currently a Final year State Medical Laboratory Technician (MLT MA). Beyond lab works, he’s a passionate internet user with a keen interest in web design and blogging. Furthermore He likes traveling, hanging around with friends and social networking to do in his spare time.

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