Metabolism Lab Exam

Front 1

What is the major storage form of glucose?

Back 1

Glycogen

Front 2

Where does glycogen exist?

Back 2

Liver and muscle

Front 3

What type of cells (specifically) require glucose?

Back 3

Cells with few to no mitochondria (red blood cells)

Front 4

What is the preferred energy source of the brain?

Back 4

Glucose

Front 5

What happens in animals if blood glucose levels fall?

Back 5

Glucagon is released if the a-cells sense the relative deficiency

Front 6

What does glucagon have to do with glucose?

Back 6

It affects its release from liver glycogen

Front 7

What happens in animals if glucose levels are high?

Back 7

Insulin is released from the beta cells in the pancreas

Front 8

What effect does insulin have on glucose?

Back 8

It affects glucose uptake by tissues (muscle and adipose) and it is needed for pancreatic a-cells to sense blood glucose levels

Front 9

What happens when pancreatic b-cells are destroyed?

Back 9

Insulin is no longer produced

Front 10

What happens when there is a high ratio of glycogen to insulin?

Back 10

Glycogen stores are broken down and depleted

Front 11

What is the structure of glycogen similar to?

Back 11

A-amylose

Front 12

How is glycogen broken down to glucose in the laboratory setting?

Back 12

Hydrolysis using a strong acid (ex. sulfuric acid)

Front 13

Purpose of experiment 1

Back 13

Determine physiological effects of fasting and feeding on liver glycogen content

Front 14

How was diabetes induced?

Back 14

Injection of streptozotocin (destroys beta cells)

Front 15

What properties of glycogen were important in lab 1?

Back 15

The stability of its glycosidic bonds under alkaline conditions, its precipitation by ethanol and its hydrolysis under acidic conditions

Front 16

How was glycogen precipitated?

Back 16

The addition of ethanol

Front 17

Why did the liver need to be removed from the rat quickly?

Back 17

Prevent anaerobic glycogenolysis

Front 18

How was the liver homogenized?

Back 18

With KOH.

Front 19

What does a reagent blank do?

Back 19

Correct for color due to the reagent itself

Front 20

What does the oxygen electrode observe?

Back 20

1. The oxidation of substrates permeable to mitochondria

2. the effect of respiratory inhibitors on mitochondrial respiration

Front 21

How does the oxygen electrode observe oxidation?

Back 21

It obseves substrate oxidation by measuring the depletion of oxygen at the platinum electrode.

Front 22

Where are substrates oxidized?

Back 22

In the mitochondria.

Front 23

How does biological oxidation occur?

Back 23

Addition of oxygen, removal of hydrogen, removal of oxygens

Front 24

What inhibitors were used in experiment 2?

Back 24

Sodium azide, antimycin A, rotenone.

Front 25

Why was NaOH used in experiment 2?

Back 25

Neutralize glucose solutions.

Front 26

What substrates were used in experiment 2?

Back 26

Succinate and B-hydroxybutyrate

Front 27

How did sodium azide affect the substrates in experiment 2?

Back 27

Partially inhibited them

Front 28

How did antimycin A affect the substrates in experiment 2?

Back 28

Inhibited them

Front 29

How did rotenone affect the substrates in experiment 2?

Back 29

Inhibited B-hydroxybutyrate but not succinate

Front 30

What does the change in osmolarity in experiment 1 indicate?

Back 30

One of the advantages of storing glucose as glycogen is that it permits mobilization of glucose without gross alterations in the solute content (and hence osmotic pressure) of the liver cell.

Front 31

What does the oxygen electrode's specificity depend on?

Back 31

The diffusion of oxygen through a membrane that prevents metal ions (or other reducible species) from diffusing to the electrode

Front 32

How is the removal of H2 from metabolites mediated in experiment 2?

Back 32

Cofactors/electron carriers--pyridine/flavonucleotides

Front 33

Where does rotenone enter the respiratory chain?

Back 33

Complex 1

Front 34

Where does Antimycin A enter the respiratory chain?

Back 34

Complex III

Front 35

Where do sodium azide and cyanide enter the respiratory chain?

Back 35

Complex 4

Front 36

What does it mean if an inhibitor blocks one substrate but not another?

Back 36

It acts before the second complex or in one of the branches of the chain

Front 37

What does it mean if an inhibitor blocks both sustrates?

Back 37

It is acting in the common path of the chain

Front 38

What does EGTA chelate?

Back 38

Ca^2+ ions

Front 39

What is the major component of respiration medium?

Back 39

KCl

Front 40

What is in respiration medium?

Back 40

KCl, buffer, chelating agent (EDTA), bovine serum albumin

Front 41

What is the pH of respiration medium?

Back 41

7.2

Front 42

How is a polarograph interpreted?

Back 42

Front 43

How are P:O ratios interpreted?

Back 43

Front 44

What does a small pen deflection on the polarograph result from?

Back 44

The fact that the polarograph is not temperature equilibriated.

Front 45

How does the Clark electrode work?

Back 45

Platinum electrode and silver electrode are in contact with KCl but are separated by a thin Teflon membrane held in place by an O-ring.

The platinum electrode gets a negative charge relative to the silver electrode.

The sensor is placed in a water jacketed chamber so the sample and sensor can be
maintained at the desired temperature.

Front 46

What is the Teflon membrane permeable to?

Back 46

Molecular oxygen.

Front 47

How does the electrical current flow in the oxygen electrode?

Back 47

Oxygen goes through the Teflon membrane where it reaches the platinum electrode and is reduced to water.

The current flows from the silver electrode to the platinum electrode as electrodes are removed.

Silver ions combine with chloride to form silver chloride.

Front 48

What reaction coupling speeds up the oxidation of NADH and FAD?

Back 48

ADP/Pi

Front 49

Why aren't NADH and FADH2 oxidized when added to mitochondria?

Back 49

They cannot permeate the inner membrane and get to their oxidation sites.

Front 50

What permeates the inner mitochondrial membrane in experiment 2?

Back 50

B-hydroxybutyrate and succinate

Front 51

Once NADH and FADH2 permeate the inner membrane what are they oxidized by?

Back 51

NAD+ and FAD

Front 52

What determines permeation?

Back 52

Presence of transport systems (NOT molecular size)

Front 53

How is the physiological link between electron transport and ATP synthesis uncoupled?

Back 53

Disruption of the inner mitochondrial membrane or by the uptake of Ca2+.

Front 54

When is EGTA used?

Back 54

As a chelating agent to remove Ca2+ during isolation

Front 55

When is EDTA used?

Back 55

As a chelating agent to remove Ca2+ in respiration medium

Front 56

What substrates cannot permeate the inner membrane?

Back 56

NAD, NADH, FAD2, FADH2, FMN AND FMNH

Front 57

What is electron transport physiologically coupled to?

Back 57

Oxidative phosphorylation

Front 58

What is needed to rapid oxidation to occur?

Back 58

ADP and inorganic phosphate

Front 59

What inhibitor was used in experiment 3?

Back 59

Oligomycin

Front 60

What does oligomycin inhibit?

Back 60

ATP synthase/respiration

Front 61

What uncoupler is used in experiment 3?

Back 61

2,4-dinitrophenol

Front 62

What does 2,4-DNP uncouple?

Back 62

Oxidative phosphorylation

Front 63

What did the polarograph measure in experiment 3?

Back 63

Diffusion current--proportional to the oxygen concentration of the solution in the
polarograph chamber

Front 64

What could happen if the respiration medium is shaken too vigourously?

Back 64

The albumin could denature and cause froth

Front 65

What does the ADP/O ratio measure?

Back 65

the ratio of the number of moles of phosphorus atoms incorporated into "high energy bonds" in ATP to the number of moles of oxygen atoms utilized

or: number of ATPs produced per pair of electrons traveling through the electron transport system (ETS).

Front 66

In experiment 3, what do the moles of ADP measure?

Back 66

Phosphorylation

Front 67

In experiment 3, what does the decrease in concentration of O2 measure?

Back 67

Respiration rate

Front 68

What signifies a new rate of oxygen consumption on a polarograph chart?

Back 68

A plateau or a change of slope to that of the "background" apparent oxygen uptake.

Front 69

What are uncouplers?

Back 69

lipid soluble weak acids that uncouple respiratory chain activity from ATP synthesis

Front 70

How does oligomycin work?

Back 70

Oligomycin inhibits oxidative phosphorylation by inhibiting ATP synthase activity. Oligomycin blocks proton conduction through the F0 portion of the ATP synthase, preventing the synthesis of ATP

Front 71

What is required for oxidative phosphorylation?

Back 71

An intact inner mitochondrial membrane

Front 72

What is oxidative phosphorylation needed for?

Back 72

ATP synthesis

Front 73

What prevented ATP synthesis in experiment 3?

Back 73

Oligomycin

Front 74

Where does oligomycin bind in the ETC?

Back 74

Complex 5

Front 75

What is the advantage of using isolated cells over sliced cells?

Back 75

Adequate oxygenation and ease of pipetting

Front 76

What is K-H saline gassed with?

Back 76

95% O2, 5% CO2

Front 77

What controls the pH in experiment 4?

Back 77

CO2

Front 78

What substrates were used in experiment 4?

Back 78

Na pyruvate, alanine, leucine

Front 79

How does the OsMolar scale work?

Back 79

1 M of any species in solution exerts
an osmotic pressure of 1 OsM

Front 80

What is the osmolarity of human blood plasma?

Back 80

About 300mOsM

Front 81

Why didn't we need to correct for dilution on the standard curve in experiment 4 as in experiment 1?

Back 81

The same amount of sample was used in the experimental and the standards

Front 82

What did it mean in experiment 4 if the solution was light pink to orange?

Back 82

Too acidic

Front 83

What did it mean in experiment 4 if the solution was purple or blue?

Back 83

Too basic

Front 84

What was used for back titration in experiment 4?

Back 84

Perchloric acid

Front 85

How was the rate of gluconeogenesis calculated in experiment 4?

Back 85

u moles glucose produced/min/mg dry wt of cells

Front 86

Why were the rats fasted in experiment 4?

Back 86

Because gluconeogenesis occurs in times of starvation--in the liver glucose is made from amino acids or glycerol in times of starvation

Front 87

Why is K-H saline used?

Back 87

It is a physiological saline and thus prevents osmotic damage to the blood cells

Front 88

Why is leucine not gluconeogenic?

Back 88

Because it converts to acetyl-CoA which will be given to the Kreb's Cycle to make L-glucose

Front 89

Why did Na Pyruvate and Alanine work in experiment 4?

Back 89

Because they convert to oxaloacetate

Front 90

What is Friedwald's equation?

Back 90

LDLcholesterol=Total cholesterol - HDL cholesterol - (triglyceride/2.2)

All values must be in mmol/L.

Front 91

What is the reaction of cholesterol assay?

Back 91

cholesterol esters ---CEH-----> cholesterol + fatty acids

cholesterol + O2 -----------CO-------> cholest-4-en-3-one + H2O2

Front 92

What is the reaction of triglyceride assay?

Back 92

Triglycerides + H2O (Lipase)---> Glycerol + Fatty Acids

Front 93

How is mg/dl converted to mM?

Back 93

18 × mmol/l

Here, use the molar mass of cholesterol, 386.6 g/mol (therefore, 386.6 mg/mmol)

Front 94

What SI unit is used for cholesterol?

Back 94

mM

Front 95

What SI unit is used for triglycerides?

Back 95

mg/dL

Front 96

What levels of serum cholesterol is considered elevated?

Back 96

5.2mM

Front 97

What levels of serum triglyceride is considered elevated?

Back 97

150 mg/dL