Bringing Back “Old” Biotechnology:
Little Miss Muffet*

            Enzymes have a central role in all living things.  People accidentally discovered the beneficial effects of enzymes long ago.  This discovery lead to leavened bread, brewed beer, and tenderized meat.  Ancient people, who used animal stomachs and bladders as canteens, must have filled it with milk one day.  Hours later, the milk clotted into creamy lumps that tasted pretty good!  These lumps, or curds, were the first cheese.  The nursery rhyme “Little Miss Muffet” mentions her consumption of curds and whey.  Curds are the coagulated milk protein that forms when milk clots; the remaining watery portion is the whey. 
            This transformation of milk was a result of an enzyme, then called rennin, that was released from the animal stomach canteen.  Today, this enzyme can be bioengineered to increase the yield dramatically.  In 1990, the U.S. Food and Drug Administration approved a recombinant rennin enzyme called chymosin.  Let’s use some “old” biotechnology and make our own cheese! 

Procedures:

1. Divide class into 3-4 test groups, depending on the number of variable enzymatic agents.
2. Using a 10 mL pipette, transfer exactly 7 mL of room temperature whole milk into a labeled, 15 mL conical tube.
3. Using a pre-set micropipette, add 0.25 mL (250 μL or 11 drops) of Rennin or Chymosin to one of the tubes of milk.

If using buttermilk or yogurt with live culture, add 500 microliters (22 drops) due to the lower concentration of enzymes. 

4. Leave one tube with whole milk and no added enzyme as the control.
5. Cap the tube and gently mix by inverting 3 times.  Record this “initial time”.
6. Place the milk-containing portion of the tube deep in your armpit, like a thermometer, and incubate it there for at least 15 minutes.  (A 37°C water bath could be substituted.)
7. Check for curdling every 5 minutes, recording the time to curdling in minutes.  Curds are large lumps of solidified milk. 
8. If curdling has not occurred by the end of the class period, keep it at room temperature and check the tube the following class period, keeping the tube upright.
9. On return to the lab, measure the amount of curds (solids) and whey (liquid) in the tube.  You may be able to read the volume of each directly off the tube although it may be difficult.
10. Pour the whey and curds through a filter paper funnel into a 10 mL graduated cylinder.  Determine the volume of whey collected in the graduated cylinder.  By subtraction, determine the volume of curds. 
11. Record the data for your sample plus one each of the other variable groups on the data table below. 

Data Table 1:  The Characteristics of Cheese made by Different Curdling Agents

*Adapted from Shoestring Biotechnology project by  DeAnn Campbell and Megan Waugh, Oklahoma City Community College, BBDiscovery Project.