An Investigation On The Effects Of Calcium Ions On The Coagulation Of Milk

Introduction

In this investigation, we are to observe the effect of calcium ions on the coagulation of milk. Here we can bring about the effect of coagulation by introducing the enzyme rennin into the milk where as the addition of calcium chloride solution will increase the concentration of calcium ions in our sample of milk. Initially I believe that in the presence of more calcium ions, the results would be a higher rate of coagulation. We can observe this by dipping a microscope slide in a sample of milk with the rennin and to observe the “flecks of curd” on the slide.

Theory and Background

Rennin is an enzyme which is synthesized in the stomach, its role is to curdle milk; this is especially important in organisms as coagulation enables the digestion of the milk protein; in this case casein. Otherwise the milk would just pass through the digestive system as a liquid, hence unable to be absorbed. It should be noted that there are 4 types of casein, which we can divide into 3 groups; alpha, beta and kappa. Kappa casein plays an important part in milk coagulation via rennin as the enzyme targets a specific bond in this and transforms it into para-kappa-casein. This is not able to stabilize the other casein types which are calcium insoluble. Thus this causes spontaneous coagulation. Also note that rennin is important in the cheese making industry due to its ability to encourage coagulation.

Equipment and Apparatus

Rennin solution Pasteur pipettes

Milk Distilled water

Sodium citrate solution ...

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Equipment and Apparatus

Rennin solution Pasteur pipettes

Milk Distilled water

Sodium citrate solution Stopwatch

mol dm-3 calcium chloride solution

Test tubes

Microscope slides

Graduated pipettes

Method

The purpose of this investigation is to observe the effect of calcium ions on the coagulation of milk via the addition of rennin solution. Therefore in order to do so we must construct a series of coagulation in which only one factor is changed, that is the concentration of calcium ions. Other environmental factors must be kept constant in order to ensure the investigation is fair as possible, also to ensure it is reliable and to reduce results being called into question by the effect of changing external factors. These are the precautions which I took to ensure the above criteria are met:

All experimental work will be performed in the same area on the same day. This will ensure that the temperature in which the reactions are taking place are all room temperature and factors such as humidity are constant.

The sample of milk I used is used in all experimental work, causing the initial composition of the milk to be exactly the same for each test.
During the production of each milk mixture, I will ensure that the volume will be exactly the same; in this case 9 cm3 in any given solution. Distilled water is to be added in order to make a mixture to the given amount, note that water is not reactive hence it will not cause any deviations or sudden change in the reaction.

Altogether there will be 6 different sample of milk to test, each will have a different concentration of calcium ions. As a control, there will be a sample of milk in which all the calcium ions are removed via the addition of sodium citrate, this will prove that calcium ions are required for the coagulation process. The following composition in the test samples are shown in a diagram below. Test tubes were to be cleaned out first in order to prevent contamination, also for each solution that was added, a different graduated pipette was used; this was to prevent any effects of cross- contamination. Due to the rather complex nature of obtaining the ‘flecks or curds’ via dipping the microscope slides, I have devised an improvised method in which coagulation can be directly observed. It will still take place on a microscope slide, however on the slide a drop of rennin from a pasteur pipette was added, and onto this, 2 drops of the milk samples from another pipette. The stopwatch is started straight after this and then this concoction was evenly spread across the slide using the specified milk sample pipette. Via this method one can see the obvious appearance of the curd forming on the surface of the sample, in which the stopwatch is stopped and the time is recorded.

As a safety precaution due to the fact that calcium chloride is an irritant substance, safety goggles were worn for eye protection and gloves were also worn when handling the solution.

Preliminary Results

As shown on the graph, I have coded the time using the function R.O.R = (1/time) X 100, this gives me a set of data which can be interpreted as the rate of reaction. Also the x- axis shows the additional amount of calcium chloride solution I have added, with 0 being the unaltered sample of milk with distilled water

Evaluation

As shown on the graph we can see that the rate of reaction is proportional to the increase of CaCl2 solution (i.e. more calcium ions), which I predicted earlier on in this investigation. However we can improve certain aspects of this experiment and investigate further into it. Firstly we can create a larger range of concentrations which we could test; this would give us a larger range of data which we could work with. Also in order to keep a constant environment we can employ the use of a water bath, perhaps set to 25oC – room temperature and also use a buffer to keep pH constant at neutral. Due to the unsuitable nature of the microscope slide, watch glasses should be used instead as the concaved shape enables the solution to stay in the container unlike the slides. For reliability it is necessary to obtain repeats, perhaps up to 3 sets of data, this will reinforce

the fact that the results are accurate and that there were no anomalous results in the data which I obtained. However it is difficult to calculate the amount of calcium ions due to the fact that some are present in the milk beforehand, one variation may be to remove it beforehand and from there add excess artificial ions via the solution. Further investigates can be attempted in the form of the reaction’s environment such as temperature and pH conditions.

References

Chymosin (Rennin) and the Coagulation of Milk

R.Bowen