The Effect of Milk Fat on the Growth of Bacteria
Picture of the student researcher
 

Researched by Charley W.
2001-02




PURPOSE

The purpose of this experiment was to determine whether skim, 1%, 2%, or whole milk would grow the most bacteria after 1 hour, 2 hours, and 4 hours of incubation.

I became interested in this idea because I have tasted spoiled milk before and I wanted to know why I got sick from it.

The information gained from this experiment will help families, nutritionists, and schools because they would know whether to have skim, 1%, 2%, or whole milk for long-term storage.




HYPOTHESIS

My first hypothesis was that the longer the milk was incubated, the more bacteria it would grow. My second hypothesis was that as the percentage of milk fat increased, the more bacteria it would grow.

I base my first hypothesis on the book Bacteria that says "Most bacteria divide every two or three hours, but some divide every 15 minutes and others may wait as long as 16 hours." So the longer it is incubated, the more bacteria should grow.

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EXPERIMENT DESIGN

The constants in this study were:

  • The amount of milk in each sample 
  • The temperature it was stored at
  • The freshness of the milk (the same expiration date)
  • The storage temperature and method
  • The test method
  • The brand of milk
  • The way of counting the bacteria
  • The method of measuring the milk samples
  • The place the milk was stored before and during the experiment
  • The location the test took place
  • The type of agar plates used
  • The type of vials used


The manipulated variables were how much time the milk was left in the incubator and the percentage of fat the milk contained. 

The responding variable was the amount of bacteria that grew. 

To measure the responding variable, I counted all of the colonies of bacteria (big red dots.) 

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MATERIALS
 
QUANTITY ITEM DESCRIPTION
4 1 gallon cartons of milk
1 ml. of lactobacillus
1 incubator
29 3m Petrifilm Plates
28 vials
1 pipette
50 pipes for the pipette
1 plastic Spreader

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PROCEDURES

1. Gather all the materials.
2. Set out the milk, (skim, 1%, 2%, and whole) the vials, (28) and the plates (29.)
3. Take 12 of the vials and label then as follows: skim 1 hr. I, (Inoculate) 1% 1 hr. I, 2% 1 hr. I, and w.m. (whole milk) 1 hr. I. The others are labeled the same way, only 4 say 2 hrs., and the other 4 say 4 hrs., not 1 hr.
4. Repeat step three, only on the next 12 vials donít put an I because you arenít inoculating these, and on the last 4 vials, label them like this: skim c, (control) 1 % c, 2% c, and w.m. c.
5. Repeat steps 3 and 4 to label the plates. On the extra plate, put water control as the label.
6. Pour 30 ml. of each type of milk into the vials with a corresponding label on it.
7. Take the vials that arenít labeled with control to the incubator and stick them in at 33-35°C (85°F).
8. With the vials labeled control and the plates labeled control, line them up and plate them.
8a. To plate the milk, take the pippettor and a pipe, and suck up 1 ml. of milk into the pipe.
8b. Lift the agar flap and squirt the milk into the middle of the plate.
8c. Use the plastic spreader to spread the milk onto 20 squares. (Put the spreader in the middle of the milk sample and press down to make it spread.
9. Put the plates in the incubator with the vials of milk.
10. After an hour, take out the 1 hour vials of milk and plate them the same way as in 8a, 8b, and 8c.
11. Put these plates in the incubator with the other ones.
12. For the 2 hour vials of milk, get them out of the incubator and line them up with their matching plates. Now plate them.
12a. (To plate these you will need to get out your sterilized water.) Suck up 1 ml. of milk and squirt it into 100 ml. of sterilized water. Turn it upside down once or twice to mix it up.
12b. Draw up one ml. of control water and 1/10 of a ml. of the inoculated water (the water with milk in it.)
12c. Plate this, now, like you did in 8a, 8b, and 8c.
13. Put these plates in the incubator with the other plates.
14. On the 4 hour milk, you will plate it the same way as you did for the 2 hour milk.
15. Put these plates in the incubator with the others.
16. Plate the sterilized water (the water you used as a control on the 2 and 4 hour milk) like you did in 8a, 8b, and 8c. 
17. Put the plate in the incubator and leave them all in there over night.
18. Take the plates out the next day and count the big red dots, or the colonies, of bacteria. (Donít count the small dots because they have not yet grown into colonies of lacto bacillus.) (Before you take the plates out of the incubator the next day, you should make sure that there are colonies of bacteria that have grown. If not, leave them in for 1 more day.
19. Record the number of dots on your data sheet. (If there are so many colonies that you canít count them all, write down tntc, which stands for too numerous to count.

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RESULTS

The original purpose of this experiment was to determine whether skim, 1%, 2%, or whole milk would grow the most bacteria at 1 hour, 2 hours, or 4 hours.

The results of the experiment were that normal (uninoculated) skim milk and one percent milk grew almost no bacteria. Milk with two percent fat and whole milk grew so much bacteria within four hours that there was too much bacteria to count.

See the data and graphs.




CONCLUSION

My first hypothesis was that the longer the milk was incubated, the more bacteria it would grow. My second hypothesis was that as the percentage of milk fat increased, the more bacteria it would grow.

The results indicate that my first hypothesis should be rejected because the 2% milk grew more at 2 hours than at 4 hours. The results also indicate that my second hypothesis should be accepted because the whole milk grew the most bacteria.

Because of the results of this experiment, I wonder what would have happened if I had used different brands of milk. I also wonder if it would have changed the results if I had used a different method of measuring to find out how many bacteria grow.

My findings should be useful to families, nutritionists, and institutions like schools because they will know that if they have either skim or one percent milk, it should spoil less quickly. 

If I were to conduct this experiment again, I would do more trials of the experiment. I would also replicate my experiment, only I would do it with more brands of milk rather than just one. The other thing I would do differently is to count the bacteria on the plates with the microbiologist, not with my mom. 
 
 

Research Report

Bacteria

Bacteria are one-celled organisms that live almost everywhere. They are usually 0.3 to two microns in diameter, (a micron is 1/25,400 of an inch) and very microscopic. However, some are so small they arenít even visible under a microscope. They have a surrounding membrane that holds the cell together, and it also controls what substances go in and out. Bacteria also have a second wall around the membrane that determines its shape. This can either be stiff or flexible. 
Types of bacteria
There are three types of bacteria: cocci, which are round, bacilli, which are rods, and spirilla, which are spiral shaped. The cocci include staphylococcus, and they live on bodies. The bacilli with flagella include Escherichia coli, which live in the intestines. The spirillum spirochete includes leptospira, and they live in mammals and domestic animals. 
What they do
Bacteria may cause disease, but they help recycle the earth. That is because they are constantly making dead matter into useable materials for new life. Bacteria are very important to life as we know it because without them, we would have no new life, since there would be no raw nitrogen, carbon, and other things needed for living. Bacteria are also helpful in cheese production.
Bacteriaís intelligence
Bacteria are "smart" in some respects. Even without a nervous system or brain, they can tell that food is nearby and they move toward it. Bacteria can also sense that there are toxic chemicals and they move away from them. (They can sense 20 at a minimum.) Eschirechia coli (E. coli) have "nose spots" to help them detect any chemical that is around. Bacteria also have 60-second memories, and some have chains of "magnets" that line up toward the magnetic north and south poles.
Grouping Together
When bacteria line up or group together, they are called "diplo" if there is a pair of bacteria. Diplococcus causes pneumonia. If they form a chain of bacteria, it is called "strepto", and just one chain of streptococcus can cause strep throat. If bacteria form a clump or cluster, it is called "staphylo". Staphylococcus can cause boils and meningitis. The bacilli usually stay separate, but sometimes they form end-to-end chains. If they do, they look more like fungi than bacteria. Mycobacterium bacilli chains cause tuberculosis. 
Growth
Bacteria grow by making "diplo", "strepto", or "staphylo" groupings. These groups grow together and create colonies. The colonies of bacteria are what you can see under a microscope. A bacterium can reproduce by dividing and then grouping. The original bacterium splits in half and makes an exact replica of itself. It keeps splitting every two or three hours usually, but sometimes it can take 16 hours, and other times it can take only 15 minutes. The new bacteria grow together to make "diplo", "strepto", or "staphylo" groups, and then make more colonies. 
Spore Bacteria
Bacterial spores are bacteria that have a thickened cell wall. They then go into dormancy, boiling water and some can even live 100ís of years. Since they are protected from heat, spore bacteria can survive pasteurization and other food sterilizations.
Lacto Bacillus
Lacto bacillus is a non-spore bacterium that turns glucose into lactose. It is mostly found in things like dairy products.

Milk

Milk is the white liquid that comes from lactating mammals. Humans often consume milk from dairy cows. It has most of the essential vitamins and minerals needed to stay healthy. Many people have called milk "The nearly perfect food". However, milk doesnít have enough of some nutrients to be truly "perfect."

Milk has water, minerals, fat, protein, and carbohydrates. A lot of people drink milk because it has almost all the vitamins and minerals needed to stay healthy. There is 87.4 percent water, 0.7 percent minerals, 3.5 percent fat, 3.5 percent protein, and 4.9 percent carbohydrates. It also has lots of calcium to help your bones grow and remain strong. This is why doctors and nutritionists recommend that people drink milk. Children should drink three glasses of milk a day at the least. Adults should drink one glass of milk a day, but more is better. Pregnant women and women who are nursing are encouraged to drink three to four glasses of milk each day, to keep the baby and herself healthy. Anyone with high cholesterol should not drink whole milk because it has so much fat. If a person with high cholesterol does drink whole milk, they could be putting themselves at risk for arteriosclerosis, an artery disease, which you get from high cholesterol. This disease can also make you have a heart attack.

Processing the Milk
The first step in processing milk is to get the milk from the farmers. Farmers often form a group called a dairy cooperative, which takes the milk sells it for the highest price possible. The government sets a minimum price that farmers can get for their milk. They sell it to the processing plant about every other day. They bring it in a large tank truck that is refrigerated. To make sure that the milk is good in composition, a laboratory technician checks it for any undesirable smell, taste, or look.

The second step is to separate and standardize the milk. This works by separating the cream and fat from the milk in a machine so that it can be made into butter and other things. The rest of the skim milk is made into other milk products or it is bottled. The other part of step two is standardization, which is when a machine called a Milko-Tester tells how much fat is in the milk. If the fat content is too low, then more cream is added, and if the content is too high, skim milk is added to it. That is how zero, one, two, and four percent milk are made. 

The third and most important step is pasteurization. Louis Pasteur was the person who discovered pasteurization. He knew that bacteria caused disease. He invented pasteurization to help stop the bacteria from growing and spreading by killing it. Pasteurization is where the milk is heated to a specific temperature for a certain amount of time to kill the bacteria. The most common method is the "high temperature short time", (htst) where the milk is heated to 72°C for 15 seconds and then cooled quickly. Another way to pasteurize the milk is by heating it to 63°C for half an hour, called the "batch" method. When milk is ultrapasteurized, it is heated to 138°C for about two seconds and then it is cooled very quickly. Milk pasteurized by any of these is able to spoil, so it must be refrigerated at all times. There is one type of milk, however, that doesnít spoil at all. It is when it is pasteurized at 149°C for six to nine seconds. This is called the "ultra high temperature" (uht) method. 

The fourth and last step, besides bottling, is homogenizing. This works by breaking up the fat globules in the milk so they donít rise to the top layer. The machine needed to homogenize milk is a homogenizer. Homogenizing milk works by forcing it through small holes under a lot of pressure. This makes more fat globules and cream in every drop.

Milk products
There are many different things made from milk, like cheese and yogurt. Some others are ice cream, butter, cultured buttermilk, cream, evaporated milk, and sour cream. The most common, of course, is skim, low-fat, and whole milk.

Skim milk has only about 0.5 percent milk fat. It is also called nonfat, even though it has this small amount of fat in it. One and two percent milk is milk with either one or two percents of fat in them. They are called low-fat milk. Whole milk has about 3.75 percent of fat in it. That is about how much fat is in it when it comes from a farm, so it is called whole milk.

Conclusion
In conclusion, food is important for everyoneís health and survival. It is very important to keep bacteria out of milk so that you donít get sick from food poisoning because of spoilage.

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BIBLIOGRAPHY

Facklam, Howard and Margery.  Bacteria.  New York: Twenty First Century Books, 1994. Pp. 7-10

Kalab, M. "Milk and Itís Constituents." December 11, 2001 http://anka.livstek.lth.se:2080/microscopy/f-milk.htm 

Marshall, Robert. "Milk." World Book Encyclopedia. 1991. Pp. 545-550

"Milk." World Book. 1998 edition CD-ROM

"Pasteurization." Microsoft Encarta, 2001. 

Puccio, Frank. Natureís Children Cows. Danbury CT: Grolier Educational, 1997. Pp. 27,28, 35 

University of Guelph. "Pasteurization" November 7, 2001 http://www.foodsci.uoguelph.ca/dairyedu/pasteurization.html

University of Texas Houston Medical School. "Lactobacillus" December 6, 2001 http://medic.med.uth.tmc.edu/path/00001446.htm

Worthington-Roberts, Bonnie. "Whole Milk, Low-Fat Milk, and Skim Milk." Microsoft Encarta, 2001. 
 


ACKNOWLEDGEMENTS

I would like to thank the following people. Without their help my project would not have been possible.

  • My mom for transporting me to Sunnyside, WA to do my experiment twice. Also, she encouraged me to keep going. 
  • The microbiologist Victor Martin for helping me do the experiment and getting me all the plates and vials and everything I needed.
  • Mrs. Helms for giving me tips on my board and my report.
  • My sister for giving me tips on doing a good report and board, and picking me up after school when I stayed late. 
  • Other 6th grade students for giving me tips to improve my board and journal.
  • Mr. Newkirk for going over my report, helping me with my graphs, and helping me with my data.

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