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Nitrate Reduction and Denitrification: A Laboratory Experiment, Study Guides, Projects, Research of Biology

A laboratory experiment on Nitrate Reduction and Denitrification, focusing on the role of bacteria in converting nitrate to nitrite and further to gaseous nitrogen compounds. Students are required to perform the experiment using three different bacterial cultures: Alcaligenes faecalis, Proteus vulgaris, and Pseudomonas aeruginosa. instructions on materials, reagents, steps, and possible results, as well as their interpretations.

Typology: Study Guides, Projects, Research

2021/2022

Uploaded on 09/27/2022

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Ex. 16: Nitrate Respiration
(Nitrate Reduction Test)
Objectives?
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Ex. 16: Nitrate Respiration

(Nitrate Reduction Test)

Objectives?

Vocabulary

• Oxidation – reduction reactions

• Reductase

• Nitrate vs. Nitrite

• Denitrification

Day 1:

Materials needed per table:  Nitrate broth tubes (4)  1 sterile Q-tip

Three pure cultures needed per table:Alcaligenes faecalis (negative)Proteus vulgaris (positive for nitrite)Pseudomonas aeruginosa (denitrifier) Each student takes responsibility to inoculate one of the nitrate broth tubes

Reagents and Steps

1. Test for accumulation of nitrite:

Add 5 drops of nitrate reagents A ( dimethyl- naphthylamine) and B (sulfanilic acid)  red color means nitrite present (+)

2. If step one is negative:

Add small pinch of Zinc powder  red means nitrate was present. Why? And what is the meaning?

3. No color development in step 1 or 2 

Why? And what is the meaning?

Nitrate Reduction Test

Test tubes after

24 hour

incubation and

before adding

reagents A and B

A. faecalis

Negative Control P. vulgaris^

P. aeru- ginosa

... after adding reagents A and B

What do you do next?

A. faecalis

Negative Control P. vulgaris

P. aeru- ginosa

Table 16-2: Nitrate Utilization Patterns Pattern Nitrate Nitrite Gas Interpretation NO 3 NOT used present^ absent^ absent^ No nitrate reduction NO 2 produced absent^ present^ absent^ Nitrate reduction^ to nitrite Gas produced absent^ absent^

Produced and escaped

Denitrification (Nitrate reduction all the way to gas)

Step 1 Step 2

Step 1

Step 2