BIO 102 Lab 03: Chemical Digestion and Nutrition

 

To submit, print this document, complete all lab activities, take a photo of your bile experiment, and answer the review questions. Scan your lab pages using the free phone app AdobeScan, and upload your PDF and your bile photo to Canvas.

· Please turn in only pages with completed work (ie., omit pages such as page 1 that only provide instructions)

· Please write your name on the 1st page you submit

 

If you have a disability that makes it difficult to complete this lab, please contact your instructor.

 

Copyright © 2016 by Mary Vander Maten and Jill Caporale.

OBJECTIVES

After completing this lab, you should be able to:

Provide an example of how an enzyme is named.

Explain the functions of enzymes during digestion.

Note the effect of bile salts on fats.

Complete a personal diet assessment

Explain Body Mass Index and its relationship to health.

INTRODUCTION

Enzymes are an important group of proteins that enable cells to carry out the chemical reactions needed to support life at temperatures that the cells can tolerate. Enzymes are organic, or biological, catalysts. They speed up chemical reactions that would otherwise occur at much slower rates and can be used over and over many times. The chemical that the enzyme affects is called the substrate. An enzyme is named by adding “–ase” to the name of the substrate it acts upon. For example, the enzyme that speeds up hydrolysis, or digestion, of fats is called lipase and the enzyme that breaks down cellulose is called cellulase.

 

 

Part 1: DIGESTION

•Mouth: The Digestion of Starch

Amylase is a protein that is an enzyme produced by the salivary glands and pancreas. Recall that starch is a complex carbohydrate composed of many glucose molecules bonded together. Amylase begins starch digestion in the mouth and hydrolyzes the starch into less complex carbohydrates, such as maltose, a disaccharide. Starch digestion continues in the small intestines.

Starch ⟶ Simple Sugars

via Amylase

 

•Stomach: The Digestion of Proteins

Protein digestion is initiated in the stomach by proteases such as pepsin and continues in the small intestine.

Protein ⟶ Amino acids

via Pepsin

Although there is some enzyme activity in the stomach, much of the work is mechanical. Searching online, find diagrams or photos of the human stomach’s interior. (Figure 33.8 in your textbook does a poor job clearly showing the stomach interior). Specifically pay attention to the illustrations of the stomach’s interior surface to observe the rugae lining the inside of the stomach.

•Small Intestine: The Primary Site of Digestion and Absorption

Don’t let the name “small intestine” deceive you. The name describes its relative circumference to the large intestine. In contrast, the length of the small intestine is about six meters whereas the length of the large intestine is about one and a half meters.

Most digestion and absorption takes place in the small intestine. Carbohydrate and protein digestion are completed in the small intestine. Lipid and nucleic acid digestion are initiated and completed. Although digestion occurs in the small intestine, some enzymes necessary for the digestion are produced by the pancreas.

The enzymes produced in the pancreas are transported to the duodenum by the pancreatic duct. Bile, produced in the liver and stored in the gall bladder, is delivered to the small intestine to aid in fat digestion. Bile is an emulsifier but is not an enzyme. It enables a more rapid enzymatic breakdown of lipids by increasing surface area of fats (by creating smaller droplets).

 

Microscopic Investigation of the small intestines

Look at a slide of columnar cells in the small intestine.

https://www.dartmouth.edu/

 

1. The brush border at the distal end of the columnar cells form the upper layer of the microvilli.

How do you think microvilli increase absorption?

 

 

2. You will notice some of the columnar cells contain small globules of mucus. These cells are called goblet cells. Why do you have cells in your intestines that produce mucus? (Isn’t mucus just for your nose?)

 

 

 

3. How many feet are in 6 meters? How many feet in 1.5 meters? Why are these numbers important in this lab? (For reference, the Chevy Suburban SUV is 5.7 m long)

 

 

 

Effect of Bile on Fats

Materials for at home demonstration:

2 clear drinking glasses

Dry mustard (see *Note below)

Spoon

Timer

Safety goggles recommended for all at-home experimentation/demonstration

Water

Into each glass, place both 3 Tablespoons of vegetable oil and 3 Tablespoons of distilled water (both glasses should have oil and water). You will be using dry mustard to substitute for bile. Add a pinch of dry mustard into glass B2. Stir the mixtures for one minute. Carefully observe both glasses for 3–4 minutes. Record your results in Table -2.

2-2 Effect of bile salts on fat dispersal.

Tube	Observation (size of fat droplets)
B1 (oil + water)
B2 (oil + water + dry mustard)

The mustard acts an emulsifier. Dispersing the fat globules of the oil into smaller droplets.

Spices like dry mustard are used in the preparation of salad oil are used not only for taste, but to break the oil into smaller particles, thus keeping the salad dressing as a suspension. Bile, emulsifies fat in the small intestines, dispersing the fat into smaller droplets thereby creating more surface area for lipases to work, which speeds up the digestion of fats.

*Note: If you do not have dry mustard, you can use an eighth of a teaspoon of regular liquid yellow mustard

Part 3: Stomachache

We have all experienced the gastrointestinal discomfort and sometimes pain caused by a bout of diarrhea. Make an educated guess regarding the source of the pain. Observe a diagram of the human digestive system. (Refer to your copy of the textbook Campbell: Biology in Focus 2nd edition it is located on page 693). Although we commonly refer to such pain as a stomachache, where is the likely emanating? Explain your reasoning.

 

NUTRITION AND HEALTH – How healthy are you?

Obesity increases risk of diabetes, high blood pressure, heart disease, stroke, arthritis, and even some cancers. Chronic conditions such as diabetes and high blood pressure make COVID-19 outcomes worse, resulting in either a longer disease course or in ~3% of all COVID patients, death.

Activity 1: Personal Assessment

There are 2 ways to see if your weight is in a healthy range.

1) One method is to measure your waist circumference. It is best if women have a waist size under 35 inches and men under 40 inches. Find a tape measure and measure your waist. _________

2) The body/mas index (BMI) is another way to see if your weight is in a good range. The National Institutes of Health (NIH) has a quick BMI check here: https://www.nhlbi.nih.gov/health/educational/lose_wt/BMI/bmicalc.htm

Your BMI _____________

Activity 2: You are a health care worker

You are the nurse or physician for these patients. Determine the BMI and circle the correct weight range:

Patient 1 Male, 5’10” 200 pounds

 

BMI: ________

 

Underweight/Normal/Overweight/Obese

 

Patient 2 Female, 5’ 5” 110 pounds

 

BMI: _________

 

Underweight/Normal/Overweight/Obese

 

When can BMI be misleading? It does not allow for variations in muscle mass, gender, or body (ie., skeletal) frame.

Patient 3 Male college football player in top physical shape: 6’ 2” 245 pounds (average weight of NFL player)

 

BMI: _________

 

Underweight/Normal/Overweight/Obese

 

Is the B.M. Index representative in this case? __________________

 

 

 

 

Activity 3: Fat versus Muscle

 

Fat characteristics: less dense, less metabolically active

Muscle characteristics: more dense, more metabolically active

 

Will it be easier to maintain a target weight if your body contains more muscle or more fat in the body?

 

 

Why?

 

Fat tissue (like all tissue) is composed of living cells that require energy to function. Explain how this fact might impact an obese person’s efforts to lose weight?

 

Activity 4: Calculating Calories needed to survive

 

The calorie is a measure of the amount of energy contained in a substance. (To observe the chemical energy in a food being transformed into visible heat energy, look for videos online entitled “burning a sugar cube”). Hence, all foods have caloric value, even “junk” foods. To determine the number of calories in a given food, they are actually calories are measured by burning the food in a special container and measuring the heat released. (You may have used a device similar to this in chemistry class, a bomb calorimeter.)

 

How many calories do you need each day? It depends on your gender, age, and activities.

Here are approximate examples of Calories burned/hour for a 150-pound person:

Sleeping	40 Calories per hour
Sitting	80 Calories per hour
Standing	120 Calories per hour
Walking	280 Calories per hour
Swimming or dancing	450 Calories per hour
Running	600 Calories per hour

 

1. Calculate daily calories needed for a 150-pound person if s/he sleeps 8 hours, sits 10 hours, stands 3 hours, walks 2 hours, and dances 1 hour in each 24-hour period: _________________________

 

 

2. Calculate calories for your own typical day:

Activity	Time	Calories

 

3. What are obstacles to encouraging people to engage in more physical activity? What could be done to encourage more exercise in busy daily schedules?

 

 

 

Activity 5: What kind of Calories are best?

A healthy diet can make a big difference in overall fitness. According to the CDC (Centers for Disease Control), “poor nutrition contributes to many costly diseases, including obesity, heart disease, and some cancers.” (www.cdc.gov/nutrition)

Here are some recommended numbers, from U.S. Guidelines (https://health.gov/our-work/food-nutrition/2015-2020-dietary-guidelines/guidelines/appendix-7/ ), for a normal diet of 2200 calories per day:

10-30% protein, or 110 grams (20% of 2200 = 440 calories divided by 4 = 110 grams*)

 

25-35% fat, or 73 grams (30% of 2200 = 660 calories divided by 9 = 73 grams**)

 

45-65% carbohydrates, or 275 grams (50% of 2200 = 1100 calories divided by 4 = 275 grams*)

 

*There are 4 Calories per gram of protein or carbohydrate.

**There are 9 Calories per gram of fat.

 

You can check calories and grams on food packaging, or by looking online

 

Food examples

1. We likely consume large quantities of sugar in our diets. For example, a can of Coke contains 42 grams of sugar.

a. Which is true about this amount of sugar as related to the daily requirement for carbohydrates?

It is 5%

It is 15%

It is 25%

 

b. The American Heart Association recommends no more than about 30 grams of simple sugar a day.

(https://www.heart.org/en/healthy-living/healthy-eating/eat-smart/sugar/added-sugars)

 

c. Is this simple sugar the best carbohydrate to consume? Why or why not? __________________________________________________________

 

d. List food sources of complex carbohydrates: _________________________________

_________________________________

_________________________________

_________________________________

 

 

2. Many of us may also consume too much fat. For example, a small serving of French fries at a fast food restaurant has about 230 calories and contains 44% fat (11 grams).

 

Here are some other sources of fats. Which do you eat?

 

Nuts and seeds Fatty fish Avocado

Cheese Eggs Butter and oils

 

3. What about salt? The sodium in NaCl can cause problems such as hypertension in some individuals (this will be discussed when studying the circulatory system). Processed foods are typically high in salt.

 

If you want simple help with your diet, check out MyPlate at https://www.choosemyplate.gov

Review Questions: (to be completed and submitted)

1. Define the term “substrate”:

 

 

2. Explain the role of enzymes in chemical digestion:

 

 

3. a) Test your knowledge of naming enzymes. What is the enzyme that works on the following substrates?

Substrate: lipids. Enzyme: _______________________

Substrate: peptide. Enzyme: _______________________

Substrate: lactose. Enzyme: _______________________

b) You may have heard of a friend or family member being “lactose intolerant”. This is caused by the individual lacking an enzyme to digest the sugar most commonly found in milk. What is your guess of the name of this enzyme? Doing some independent online research, what happens when a person is incapable of digesting this milk sugar, lactose? Why do you think that evolutionary change has removed the ability of grown adults to digest milk sugars?

 

4. a) Bile is made in the liver, stored in the gallbladder and is secreted into the small intestines to assist with digestion of fats through its action as an emulsifier. What is an “emulsifier”?

 

 

b) What is the difference between and emulsifier and an enzyme?

 

 

c) Performing online research, what are everyday examples of emulsified items that you eat/drink?

 

 

 

5. Explain the function of bile:

 

 

 

6. How do you think microvilli work to increase absorption?

 

 

7. What cells in your intestines produce mucus?

 

8. For each of the following substrates, state the locations of its chemical digestion occurs in the body, which type of enzyme is involved, and what units these molecules are broken down into.

 

Organic Molecule	Location of digestion	Enzyme involved	Monomer units
Carbohydrate
Fats (triglyceride)
Proteins

 

 

9. The Body Mass Index uses __________ and _____________ to see if a person’s weight is healthy.

 

10. Fill in the chart below.

Type of Tissue	More or less dense	More or less metabolic activity
Fat
Muscle

 

11. Number these activities in order of activity, starting with the highest (you may rank activities as tied for equal activity, denoting them as, for example, “2-tied”):

___ walking ___ sitting ___ running ___ riding a bicycle

___ sleeping ___ swimming ___ standing ___ using a skateboard

Why did you rank a specific activity as highest, and why?

 

12. Most processed foods have too much (check all that apply)

___ fresh fruit ___ salt ___ fresh vegetables

___ water ___ whole grains ___ fat

___ simple sugar ___ protein ___ leafy vegetables

 

 

1

Bio 102 Lab 03: Chemical Digestion and Nutrition

 

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Module 6 Overview

In this week’s face-to-face lab session, you will measure the frequencies of two traits related to bitter  taste perception  within your lab section.  Then, you will use the bitter taste sensitivity data collected in your lab section to test hypotheses about the genetic and environmental basis of food preferences.

 

Objectives

By the end of this week students will be able to:

1. Differentiate between environmental and genetic influence on phenotype

2. Formulate a testable hypothesis

3. Use phenotype frequencies to test for alternative patterns of inheritance

4. Analyze data to evaluate alternative hypotheses

 

Activities In Lab This Week

1. Complete the food preference survey on page one of the lab activity handout and discuss the genetic and environmental basis of food preferences as a class  [satisfies Learning Objectives #1-2]

2. Work with a partner to record taste perceptions, and share data as a class [satisfies Learning Objective #3]

3. Use the class food survey and  taste perception  data to test hypotheses about the relative importance of genetics versus the environment in shaping food preferences [satisfies Learning Objective #4]

 

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Write

Objective, Background, method, result and conclusion (ALL IN PAST TENS )

ANSWER THE Questions.

SEE THE RESULTS IN EXCEL SHEET

 

• a table of the data from the growth experiment
  • Time in mins
  • count date from each group.
  • average data
  • actual dilution (remember the 0.1 ml added to the plate is another 10 fold dilution)
  • Cfu/ml (represent with one digit to the left of the decimal point and two to the right then the exponent)
  • Absorbance at 600nm
  • Calculated CFU using the Aligent website
• GraphTitleY axisX axisE coli Growth CurveAbsorbance (600nm)Time (mins)E. coli Growth CurveLog bacterial numbers (CFU MM/ml) Aligent)TimeE. coli Growth CurveLog bacterial numbers (CFU MM/ml)TimeE coli Standard CurveAbsorbance (600nm)Cell numbers (MM/ml) for both Aligent and your CFU data
  • 

Questions

• Look at the growth curves using Aligent CFU and your CFU: are they similar or different.  If they are different why is that..
• Look at the aligent  plate count vs absorbance graph  – are they similar are they different.  If they are different why do you think that
• Look at the aligent and your cfu data vs time  – calculate the maximum time of doubling
• 

Lab 2 Bacterial Growth Curve

 

 

 

 

4

Objective

· Follow the growth of a Escherichia coli using cell counts and absorbance · Determine the maximum growth rate · Develop a relationship between cell counts and absorbance

 

Background

· Bacterial growth follows a standard pattern of lag, log, stationary, and death phase.   · Bacteria grow at different rates depending on the strain and the temperatures.

 

 

 

Materials	· 100 ml LB broth in 500 ml flasks – 2 · 50 ml LB in 250 ml flasks – 2 · Sterile distilled water for dilutions – 3, 100ml bottles · Sterile microfuge tubes · Sterile serological pipettes – 5ml · Sterile Falcon tubes · LB Plates – made in previous lab · Alcohol · Glass spreaders · Sterile 1000 ul tips · Sterile 200 ul tips · Sterile test tubes · Distilled water · Spectrophotometer · Cuvettes Notes on preparation – · Two days before the lab inoculate E.coli into 10 ml nutrient broth tube and incubate at 37oC · The night before inoculate a 50ml/250 ml flask with 1 ml of the broth from the culture. Shake overnight at 37oC _______________________________________________
Procedure	· Inoculate the 100 ml flask with 5 ml from an overnight culture. Mix by swirling · Aseptically take a 2 ml sample after inoculation = T0 · Place in shaker set at 37oC and shake at 300 rpm · Take 2 ml samples using a 5 ml pipette and place it in a sterile Falcon tube at: · 20 min · 40 min · 60 min · 80 min · 100 min · 120 min · 150 min · 180 min · Note you may need to gather some samples outside of normal class times · For each sample you will need to · dilute it and plate on LB plates · measure absorbance at 600 nm · Absorbance · Take a 1 ml sterile sample and place it in a cuvette. Measure the absorbance at 600nm. If needed, dilute the sample with DI water so it is in the 0.05 to 0.7 range. Record the absorbance and the dilution. Multiply the absorbance value by the dilution to get the actual value. · Use the absorbance at 600nm to estimate the number of cells using the website http://www.genomics.agilent.com/biocalculators/calcODBacterial.jsp calculate the amount of number of cells present in your sample. · Dilution and plating · Use aseptic practices · For each samples, set up sterile Eppendorf tubes containing 1000ul of sterile DI water . · Remove 100 ul from each tube using a sterile pipette tip. · Take 100 ul from the sample you took from the flask and add to the first dilution tube: this is the 10 -1 dilution or 1/10th ). Mix the 10-1 dilution in the vortex mixer for 20 seconds, then take a 100ul sample and place it in the next tube. Continue until you have a 10-7 dilution · Sample · 10-1 · 10-2 · 10-3 · 10-4 · 10-5 · 10-6 · 10-7 · Using an estimation of the cell counts from absorbance select the dilution that will give you between 30 to 300 colony-forming units (cfu). Remember you will be plating 0.1 and not 1ml so take this into account with your dilutions. Also plate the next highest and next lowest dilutions. · Identify the appropriate dilution of the sample in sterile water in sterile tubes so one of the plates has between 30 – 300 cfu. · Plate 100ul ml of suspension on the plate in duplicate · Dip a glass spreader in alcohol and flame (Note: do not heat in the flame) . Use the spreader the inoculum on the plate. · Incubate plates upside down at 37oC for 24 hours. Store plates at 4oC until needed
Data Tabulation	Absorbance Data · | Time | absorbance (600nm)) | dilution | Final Absorbance |Aligent CFU| Cell counts · |Time | Dilution | CFU plate 1 | plate 2| plate 3 | Average cell count (cfu/ml) |
Data Analysis	Make the following graphs Title Y axis X axis E coli Growth Curve Absorbance (600nm) Time (mins) E. coli Growth Curve Log bacterial numbers (CFU MM/ml) Aligent) Time E. coli Growth Curve Log bacterial numbers (CFU MM/ml) Time E coli Standard Curve Absorbance (600nm) Cell numbers (MM/ml) for both Aligent and your CFU data   Calculate · Estimate the maximum doubling time using the growth curves from Aligent and also your CFU data . · See video · https://www.youtube.com/watch?v=W6OkbTAuQVc · For more information, see details under assignments.
Report	· The information from this lab will be written as a formal report.

 

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How to Proceed

• Read through the introductory materials below.
• Open the Unit 5 Experiment Answer Sheet and complete the following Experiment exercises this unit:
  • Experiment 5 Exercise 1 – Transcription and Translation (~15 min)
  • Experiment 5 Exercise 2 – Translation and Mutations (~1 hr)
  • Experiment 5 Exercise 3 – Mutation Rates (~30 min)
• Save your completed Unit 5 Experiment Answer Sheet and submit it no later than Sunday midnight (CT).

Transcription and Translation – Introduction

Be sure that you have read over our online lecture this unit on DNA and read pp 177 to 181 in your book before starting. DNA can be a complex concept to grasp, and there is a lot of terminology to keep straight. These first two exercises will focus on transcription and translation, the two processes responsible for taking the information embedded in our DNA and using it to create a protein.

There are segments in our DNA called genes that code for the proteins needed to carry out cellular functions. These genes are a sequence of nucleotides; adenine (A), thymine (T), cytosine (C) and guanine (G) and the specific sequence of these nucleotides is what conveys the information needed to produce a given protein. In humans, the smallest gene is 252 nucleotides long, whereas the largest is more than 2 million nucleotides long! The genetic code is used to decipher the sequence of nucleotides into a sequence of amino acids. The code uses a series of three-nucleotide sequences called codons. Each different codon codes for an amino acid and it is this specific sequence of amino acids that determines what protein is formed.

DNA is found in our nucleus, yet our proteins are synthesized in the cytoplasm. A gene must first be transcribed into a form that can leave the nucleus. Transcription is the process in which a sequence of DNA used to synthesize a complementary strand of messenger RNA (mRNA). This mRNA acts a template and is used to translate the original DNA sequence into a protein, based on the information in its codons and the Genetic Code.

For example, the DNA sequence ATG-CGT-TAG-CGT-ATTC would be transcribed into the mRNA sequence UAC-GCA-AUC-GCA-UAA. Then, using Fig 10.11 on p 180 in your book, you can determine that this mRNA would be translated into the amino acid sequence Tyrosine-Alanine-Isoleucine-Alanine-Stop.

In Exercise 1, you will have the opportunity to demonstrate your understanding of transcription and translation. You will be using the following website; be sure that you are able to access and use the site:

University of Utah. No date. Transcription and Translation
https://learn.genetics.utah.edu/content/basics/transcribe/ (Links to an external site.)

In Exercise 2, you will apply what you learned in Exercise 1 and evaluate the effect that different types of mutations have on the outcome of transcription and translation. You’ll want to review these mutations on pp 186-187 of your book and in our online lecture on DNA before starting. You will be using the following website; be sure that you are able to access and use the site:

McGraw Hill. No date. Virtual Lab: DNA and Genes
http://glencoe.mheducation.com/sites/dl/free/0078802849/383936/BL_26.html (Links to an external site.)

Finally, in Exercise 3, you will complete a series of calculations to determine the probability of a mutation occurring within a gene that results in a change in protein structure. These are straight-forward math calculations; do not let them overwhelm you.

 

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