I did an experiment in class; my hypothesis is that there will be less product when an inhibitor affects an enzyme. Tube 1: 10 drops of catechol + 5 drops of phenylthiourea + 10 drops of enzymes. Tubes 2 and 3, they have the same amount of catechol and enzymes but tube 2 has 10 drops of phenylthiourea and tube 3 has 20 drops. My hypothesis is supposed to be right. Why doesn't the experiment work?

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Catechol is an organic molecule that can be oxidized to benzoquinone using an enzyme called catechol oxidase commonly found in potato extract.  This reaction is the reason that fruits and vegetables turn brown over time.  Phenylthiourea (PTU) acts as a competitive inhibitor for catechol oxidase.  It will bind to the enzyme's active site instead of the catechol and halt the reaction.  It sounds in your question that you are keeping the amount of enzyme and substrate the same but increasing the amount of PTU inhibitor in each tube.  If the reaction is still occurring even with the inhibitor, you may be adding the reagents together in the wrong order.  The PTU must be added to the enzyme first and allowed to react before the catechol is added.  If the catechol is added first or too soon after the PTU, it will not give the inhibitor time to react with the enzyme.

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