Cap Lab 7: Chemical Digestion Experiment

Front 1

What are enzymes?

Back 1

Large proteins that are biologic catalysts and increase the rate of a chemical reaction without themselves becoming part of the product.

Front 2

What are all digestive enzymes classified as?

Back 2

Hydrolases
Break down organic food molecules by adding water to the molecular bonds, thus cleaving the bonds between the subunits or monomers

Front 3

Why can digestive enzymes be studied in a test tube?

Back 3

They function outside the body cells in the digestive tract, so their hydrolytic activity can be studied in vitro

Front 4

Where is salivary amylase made and secreted?

Back 4

Made in parotid salivary glands, sublingual salivary glands and submandibular salivary glands

Secreted into the mouth

Front 5

What does salivary amylase do?

Back 5

Hydrolyzes starch to maltose

Front 6

Why must controls be prepared?

Back 6

We need to identify the presence of starch, water and amylase at the beginning to determine to what extent the activity has occured; must provide a known standard against which comparisons can be made

Front 7

What is the equation of starch to maltose?

Back 7

Starch + water + amylase --> maltose (units) + amylase

As starch decreases, sugar increases as digestion occurs

Front 8

What does the IKI test determine?

Back 8

Whether starch is present in your final solution or not (don't want any present, want it all converted to maltose)

Blue/black color=starch present=positive starch test
Otherwise negative starch test, starch not present

Front 9

What does Benedict's solution test?

Back 9

Whether maltose is present or not
Put drops in mixture and boil
--green/orange precipitate forms, maltose present, positive sugar test
--otherwise negative sugar test, no color change

Front 10

What were the ideal conditions for the amylase digestion of starch to take place?

Back 10

Incubated at 37 degrees; would get negative IKI test and positive benedict's test (no starch present, maltose present)

When incubated at 0 degrees, get some sugar but not a whole lot, still have starch

When boil amylase, get starch but no sugar---denatured enzyme

Front 11

Where are monosaccharides absorbed once broken down?

Back 11

Into intestinal villi; transported via hepatic portal vein to the liver

Front 12

Label this chart:

Back 12

Front 13

How exactly does Bennedict's test work? (determines if sugar is present)

Back 13

It searches for the presence of "reducing sugars", or sugars with aldehydes. i.e. if a sugar has been broken up, it will probably have aldehydes on its end

Cu +2 is reduced to Cu +1 by reducing sugar which precipitates as a red copper oxide

Front 14

How do amino acids get absorbed?

Back 14

Absorbed into the intestinal villi and transported via the hepatic portal vein to the liver

Front 15

How does the IKI test work?

Back 15

Iodine ions in potassium iodide solution react with starch to change color of the solution

Front 16

What is trypsin and what does it do?

Back 16

An enzyme produced by the pancreas
Hydrolyzes proteins to small fragments (proteoses, peptones and peptides)

Front 17

What is BAPNA?

Back 17

A synthetic protein substrate consisting of a dye covalently bound to an amino acid

Front 18

What happens in trypsin hydrolysis of BAPNA?

Back 18

Cleaves the dye molecule from the amino acid, causing the solution to change from colorless to yellow

Front 19

Why are additional tests not needed in Activity B, with Trypsin and BAPNA?

Back 19

The covalent bond between the dye molecule and amino acid is the same as the peptide bonds that link amino acids together, so the appearance of yellow indicates the presence and activity of an enzyme capable of peptide bond hydrolysis.

Basically: The color change is direct evidence of hydrolysis. If the mixture remains clear, no detectable hydrolysis has happened.

Front 20

Draw the flowchart for protein digestion.

Back 20

Front 21

What were the best conditions for the hydrolysis of protein with trypsin and BAPNA?

Back 21

Occurred at both 37 degrees C and 0 degrees C; when we boiled the trypsin first it didn't work because our enzyme ended up becoming denatured

Front 22

How are lipids absorbed in the body?

Back 22

By lacteals in the villi and into the lymphatic vessels and eventually into the blood circulation.

Front 23

How are fats and oils digested and how is this different than proteins and carbs?

Back 23

Must go through more complicated treatment...two steps:
1. Fats/oils are EMULSIFIED in the presence of bile salts to make droplets
2. The droplets are digested by lipase to free fatty acids and glycerides

Front 24

What does pancreatin mean?

Back 24

The enzymatic product of the pancreas, which includes protein, carbohydrate, nucleic acid and fat-digesting enzymes.

Front 25

What is pancreatic lipase?

Back 25

Hydrolyzes fats and oils to monoglycerides and two fatty acids (and occasionally to glycerol and three fatty acids)

Front 26

What is an easy way to determine if fat digestion has taken place?

Back 26

Test pH using litmus blue, which changes from blue to pink as contents become acid.

Some of the end products of fat digestion are organic acids that decrease the pH, so we use pH to recognize the step of digestion of fats we are in.

Front 27

What is bile?

Back 27

Not an enzyme but a secretary product of the liver that is still important

Important to fat digestion because of its emulsifying action--the physical breakdown of larger particles into smaller ones, which provide a larger surface area.

Front 28

How did we demonstrate emusification of lipids?

Back 28

Added water, vegetable oil and a pinch of bile salts to test tubes, shook and let stand at room temperature

If emulsification did not occur: oil floats on surface of water

If it did occur: fat droplets will be suspended throughout water, forming an emulsion (7L)

Front 29

How did we test to see if lipid hydrolysis occurred?

Back 29

Put litmus cream (cream providing fat substance with litmus powder) and lipase in tubes, incubated for certain amounts of time...color control (.1 N HCl) added to tubes 1L and 2L only

When tubes turned pink, it meant hydrolysis had occurred, no additional assay reagents necessary

Front 30

What were the most favorable conditions of lipid hydrolysis?

Back 30

Incubated for 37 degrees C; that's where we got the color change

0 degrees too cold, boiled lipase denatured it

Front 31

What were the results of the lipid hydrolysis experiment with the control tubes, 1L and 2L?

Back 31

Added .1 N HCl to both
In 1L, since no lipid present, still did not get color change
In 2L, since lipid present but no lipase, wouldn't normally get color change but since we added the acid, we did

Front 32

What kinds of muscles are involved in movement of foods down digestive tract?

Back 32

Smooth and skeletal

Front 33

What is swallowing?

Back 33

i.e. deglutition
Skeletal muscle activity
2 steps = buccal and pharyngeal-esophageal

Front 34

Describe the two phases of deglutition

Back 34

Buccal: voluntary, controlled and initiated by the tongue

Pharyngeal-esophageal: involuntary, works through peristalsis, results in delivery of contents to stomach

Front 35

What two sounds can you hear in stomach when you take a swallow of water?

Back 35

Gastroesophageal sphincter: water splashes against it

Second when peristalic wave of esophagus arrives at sphincter and it opens, allowing water to enter stomach

Front 36

What is a catalyst?

Back 36

Increases the rate of a chemical reaction without becoming part of the product

Front 37

What is a control?

Back 37

Provides a standard of comparison for test results

Front 38

What is an enzyme?

Back 38

Biologic catalyst that is a protein in nature

Front 39

What is a substrate?

Back 39

Substance on which a catalyst works

Front 40

List three characteristics of enzymes

Back 40

1. Not used up in the reaction/does not change shape
2. Used to speed up a reaction
3. Will bind to a molecule if the enzyme-substrate complex can form (highly specific)

Front 41

The enzymes of the digestive system are classified as hydrolyses. What does this mean?

Back 41

They catalyze digestive hydrolysis reactions

Front 42

What produces salivary amylase, where is its site of action and what is the substrate?

Back 42

Salivary glands
Mouth
Starch

Front 43

What produces trypsin, what is the site of action and what is its substrate?

Back 43

Pancreas (exocrine)
Small intestine
Proteins

Front 44

What makes pancreatic lipase, where is its site of action and what is its substrate?

Back 44

Pancreas (exocrine)
Small intestine
Fats

Front 45

What is the end products of digestion for proteins?

Back 45

Amino acids

Front 46

What are the end products of digestion for carbohydrates?

Back 46

Simple sugars (monosaccharides/disaccharides)

Front 47

What are the end products of digestion for fats?

Back 47

Fatty acids and glycerol

Front 48

What conclusions can you draw when an experimental sample gives both a positive starch test and a positive maltose test after incubation?

Back 48

Not all starch has been broken down and converted to maltose/amylase

Front 49

Why was 37 degrees C the optimal incubation temperature?

Back 49

Because the body's enzymes work at their optimum temperature

Also, enzymes encounter substrate molecules more slowly at lower temperature

Front 50

Why did very little, if any, starch digestion occur in test tube 6A? (incubation at 0 degrees C)

Back 50

The temperature conditions were not ideal fo amylase to work

Front 51

In the exercise concerning trypsin function, why was an enzyme assay like Benedict's or Lugol's IKI (which test for the presence of a reaction product) NOT necessary?

Back 51

Because the color change from clear to yellow is direct evidence of hydrolysis--additional tests are not required

The other tests measure cleavage products, so are indirect

Front 52

Why did you do tube 1T? (no BAPNA)

Back 52

It was a negative control. BAPNA causes a color change when trypsin hydrolysis occurs. Since we didn't add BAPNA in the tube, if we get a color change we know our experiment is invalid and contaminated

Front 53

Why did you do tube 2T? (no trypsin)

Back 53

BAPNA only causes a color change in trypsin. Again, this is a control tube--if BAPNA causes a color change without trypsin present, our experiment is invalid/contaminated.

Front 54

Trypsin is a protease similiar to pepsin, the protein-digesting enzyme in the stomach. Would trypsin work well in the stomach?

Back 54

No because the stomach is highly acidic while trypsin works well in the neutral/basic environment of the small intestines

It would denature in the stomach

Front 55

In the procedure concerning pancreatic lipase digestion of fats, why did the litmus indicator change from blue to pink during fat hydrolysis?

Back 55

Because the change of blue to pink indicates the presence of acids as is the case when fats are broken down into glycerol and fatty acids

Front 56

What was used to test for protein hydrolysis, which was indicated by a yellow color?

Back 56

BAPNA

Front 57

What was used to test for the presence of starch, indicated by a blue-black color?

Back 57

IKI (Lugol's iodine)

Front 58

What was used to test for the presence of fatty acids, which was evidenced by a color change from blue to pink

Back 58

Litmus

Front 59

What was used to test for the presence of reducing sugars as indicated by a colored precipitate?

Back 59

Bennedict's solution