In the nitrate reduction, Reagent A (sulfanilic acid) and Reagent B (a-naphthyla

Question

In the nitrate reduction, Reagent A (sulfanilic acid) and Reagent B (a-naphthylamine) are added to the tube after incubation. A postive nitrate reductase test will result in the media turning bright red.

a) Explain the functions of Reagents A and B?

b) If the media did not turn color after additionof the reagents, how would you interpret these results?

c) Explain why the development of a red color after the addition of zinc is considered a negative test.

d) if the media remains colorless after the addition of zinc, what final product of dentrification is being produced?

Explanation / Answer

Ans.

Ans. A. see step 1

Ans. B. See I and II of step 2.

Ans. C. See 2A.

Ans. D. ammonia, molecular nitrogen, nitrous oxide, nitric oxide, etc. See 2B.

Nitrate reduction test:

Aim: to test if Nitrate is reduced to nitrite by microbe-

            Nitrate ----------------> Nitrite

Step 1: Nitrite forms nitrous acid in aqueous system.

Nitrous acid + sulfanilic acid (reagent A) -------> diazotized sulfanilic acid

diazotized sulfanilic acid + a-naphthylamine (reagent B) --> RED colored compound

Step 2: To confirm false negative results

A negative test result (no development of red color with reagent A and B) indicates that- I. the bacteria does not reduce nitrate, or II. It converts nitrate and/or nitrite into ammonia or N2 gas. So, the test does not give positive result for nitrite because it has been denitrified.

To confirm this, a pinch of zinc is added to the test tube treated till step 1. Zn effects is summarized as follow-

2A. Zn reduces nitrate into nitrite (as bacteria do). The resultant nitrite gives positive red color as that of step 1. However, here it indicates that nitrate was present in the medium and NOT reduced into nitrite unless Zn was added. So, this color change indicates that nitrate was not reduced. Or, the bacteria was unable to reduce nitrate into nitrite.

2B. If addition of Zn does not result color change, it’s considered a positive result. No color change means that nitrate was reduced to byproducts like ammonia, molecular nitrogen, nitrous oxide, nitric oxide, etc. and no longer available in form of nitrite- that reacts with reagent A and B.     

            Tube (step 1 completed) + a pinch of Zn ------> Red/ Pink color (negative result)

            Tube (step 1 completed) + a pinch of Zn ------> No red color (positive result)

Related Questions

Navigate

• Browse (All)
• Browse I
• Subjects

Integrity-first tutoring: explanations and feedback only — we do not complete graded work. Learn more.