**Prelab 8 - Respiration**You have learned that in glycolysis and the Kreb's cycle, the compound NAD is reduced to form NADH. The hydrogen added on the end of the formula is an indication that NAD has been reduced. As you read in the textbook section on oxidative phosphorylation, a series of redox reactions oxidize NADH back to NAD. All redox reactions are paired oxidation and reduction reactions, so what was reduced? Ultimately O₂ is reduced to H₂O (note the addition of hydrogen due to reduction). Oxygen is said to be the final electron acceptor, as no further redox reactions take place.In lab we will again look at differences among bacteria in their metabolism as a way to distinguish different species. First we will look at anaerobic respiration. When oxygen levels are low, some microbes can use a different compound than oxygen as their final electron acceptor. Sulfate SO₄²⁻, carbonate CO₃²⁻, and nitrate NO₃⁻ are all common alternatives, rich in oxygen, which can accept those electrons. In lab this week we will use a nitrate media. Bacteria that reduce nitrate can form nitrite, NO₂⁻. Some bacteria go on to reduce nitrite to nitrogen gas, N₂, in which all the oxygen atoms have been removed by reduction. This test is more complicated because we must use reagents first to test if there is any nitrite produced (using reagents A and B). If not, then determine if any nitrate is still present or was everything reduced to nitrogen gas (using zinc dust).In the oxidase test we will determine which bacteria have cytochrome c, one version of the important enzyme in the electron transport chain. This enzyme plays a critical role in all those reactions with NADH, however some bacteria use alternative cytochromes. Thus, this becomes another way to distinguish between species.Lastly, we will conduct the catalase test. As mentioned in lecture, during respiration some of the hydrogen may combine with oxygen to form hydrogen peroxide, H₂O₂. Hydrogen peroxide is lethal to many bacteria. To protect themselves, they produce the enzyme catalase to break it down to water and oxygen. We will determine which bacteria produce this useful enzyme.1. When nitrate is used as the final electron acceptor, what two possible compounds might it be reduced to? ______________________ ____________________2. What reagent is added to culture

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Prelab 8 - Respiration You have learned that in glycolysis and the Kreb's cycle, the compound NAD is reduced to form NADH. The hydrogen added on the end of the formula is an indication that NAD has been reduced. As you read in the textbook section on oxidative phosphorylation, a series of redox reactions oxidize NADH back to NAD. All redox reactions are paired oxidation and reduction reactions, so what was reduced? Ultimately Oz is reduced to H;0 (note the addition of hydrogen due to reduction). Oxygen is said to be the final electron acceptor, as no further redox reactions take place. In lab we will again look at differences among bacteria in their metabolism as a way to distinguish different species. First we will look at anaerobic respiration. When oxygen levels are low, some microbe can use a different compound than oxygen as their final electron acceptor. Sulfate SO?², carbonate CO,, and nitrate NO, are all common alternatives, rich in oxygen, which can accept those electrons. In lab this week we will use a nitrate media. Bacteria that reduce nitrate can form nitrite, NO2. Some bacteria go on to reduce nitrite to nitrogen gas, N2, in which all the oxygen atoms have been removed by reduction. This test is more complicated because we must use reagents first to test if there is any nitrite produced (using reagents A and B). If not, then determine if any nitrate is still present or was everything reduced to nitrogen gas (using zinc dust). In the oxidase test we will determine which bacteria have cytochrome c, one version of the important enzyme in the electron transport chain,. This enzyme plays a critical role in all those reaction with NADH, however some bacteria use alternative cytochromes. Thus, this becomes another way to distinguish between species. Lastly, we will conduct the catalase test. As mentioned in lecture, during respiration some of the hydrogen may combine with oxygen to form hydrogen peroxide, H;O2. Hydrogen peroxide is lethal to many bacteria. To protect themselves, they produce the enzyme catalase to break it down to water and oxygen. We will determine which bacteria produce this useful enzyme. 1. When nitrate is used as the final electron acceptor, what two possible compounds might it be reduced to? 2. What reagent is added to culture in nitrate broth to determine if nitrite is present? 3. What does a positive test with zinc dust mean for nitrate media? 4. We will use liquid oxidase reagent for the oxidase test. What color will the colonies turn if they contain cytochrome с? 5. When liquid hydrogen peroxide is dropped on the bacterial colonies for the catalase test, what gas is produced that causes the bubbles?