Foods are made up of lots of different molecules - called Macromolecules and micromolecules. When we talk about a food's protein, carbohydrate and fat content (the information you see on food labels), we are referring to the macromolecules that make up the food. We can test for these different types of macromolecules in the laboratory to determine the other things that make up our favourite foods.
The Definition of a Biomolecule
A biomolecule has a few different definitions, being quite loosely defined. In general, a biomolecule is a molecule that has an importance of some sort in one or more Biological Processes.
The Biological Processes we're talking about range from Respiration and Photosynthesis to digestion and Cell Division. As you can imagine, the range of processes included in this definition can be vast; however, surprisingly, the majority of these processes involve the same small group of biomolecules, as we will see below!
The main biomolecules involved in any metabolic process are from the following list; Carbohydrates, Proteins, Nucleic Acids and Lipids. Aside from this list, there are some other, slightly less common biomolecules found in many different organisms that help carry out various metabolic processes; these are ATP, water and different minerals and ions. Each of these biomolecules plays a crucial role. Let's have a look at a few examples in the table below.
| Biomolecule | Example of a role |
| Carbohydrates | Carbohydrates like glucose are essential in the process of Respiration in many different organisms. |
| Proteins | Proteins are needed for the growth and repair of muscle tissue in Animals. |
| Lipids | Lipids play an important role in insulation, buoyancy, temperature regulation and energy provision in Animals. |
| Nucleic Acids | Nucleic acids are essential as a store of Genetic Information across many different organisms. |
| Minerals and ions | Magnesium is the core ion component of chlorophyll in Plants, allowing Photosynthesis. |
| Water | Water acts as a transport medium across many different organisms. |
| ATP | ATP is an energy store across many different organisms. |
Testing for Biomolecules in Food
We can test for lots of different biomolecules in foods. The main molecules we usually test for are proteins, fats (lipids), and carbohydrates (sugars and starch). The tests we use to look for the presence of these foods are called qualitative tests.
A qualitative test shows us whether something is present or not. It does not show us how much of a particular substance is present.
Let's think about flame tests in chemistry. A specific colour might tell us if a particular metal is present (e.g. a yellow flame often indicates sodium is the metal present), but it cannot tell us how many grams of the metal is present. Similarly, the tests that we use to identify the presence of biomolecules in food only show us whether the molecule is present or not, not how many grams of the molecule is present.
The opposite of a qualitative test is a quantitative test. A quantitative test shows us how much of a molecule is present, rather than just telling us whether the molecule is present or not.
Testing for Proteins
We can test for proteins by using a biuret solution. Biuret comes in two solutions, often named biuret A and biuret B. These two solutions are copper (II) sulphate and either potassium hydroxide or sodium hydroxide. When we add these two solutions together, we get biuret A; we then add potassium sodium tartrate and have biuret B.
Now that we understand the biuret solutions, let's have a look at the method behind testing for proteins;
Add 1 cm3 of biuret solution A to the food solution
Mix the two liquids
Add 1 cm3 of biuret solution B and shake OR add 1 cm3 of biuret solution B carefully down the side of the test tube to form two layers
The qualitative tests we are discussing can either have positive or negative results. A positive result shows us that the molecule we tested for is present in the solution. A negative result tells us that the molecule is not present in the substance. Have a look below at the positive and negative tests when using biuret solution to test for proteins:
- Positive test - colour change from blue to purple or a purple ring forming between the two layers.
- Negative test - colour remains blue, and no purple ring formed.
Remember when describing colour changes to always include the word from. The solution changes from x to y. We need to ensure we have both the colour before the change and after to get full marks in exams!
Testing for Fats/Lipids
The terms fats and lipids are often used interchangeably, so testing for lipids/fats will require the same test. We can use two tests to test for the presence of lipids/fats in a substance. The two tests that we can use are;
Sudan III Test
Emulsion Test
Let's look at the Sudan III test first. Sudan III is a dye that causes a colour change in fats.
Sudan III is a fat-soluble diazo dye. It is structurally related to azobenzene. You do not need to know about the structural formula of Sudan III (or the other solutions used as tests).
Here's how we carry out the test;
Add equal amounts of the food in question, and water to a test tube
Add a few drops of Sudan III solution to the test tube
Shake the test tube
A positive Sudan III will result in a red-stained layer on the water's surface.
A negative Sudan III will result in no red-stained layer forming.
Now let's have a look at the emulsion test:
Crush up the food in question and add it to a test tube
Add ethanol to the test tube
Take the liquid from the test tube, and pour it into a second test tube containing water
A positive emulsion test causes a white emulsion to appear in the second test tube.
A negative emulsion test leads to no emulsion being formed. Instead, a colourless solution remains.
An emulsion is a mix of two liquids that are typically immiscible. When fluids are immiscible, they do not mix.
Remember, when discussing the formation of an emulsion, we still have to include the word from. It is a colour change from a colourless solution to a white emulsion.
When discussing colour changes, we must avoid using the word 'clear'. Clear is not a colour but describes colours instead. Rather than using the word clear, we should use 'colourless'.
Biomolecule Test for Reducing Sugar
Sugars are carbohydrates. We can have two different types of sugars, either reducing or non-reducing. The tests that we use to test for the presence of reducing sugars and non-reducing sugars are slightly different, and we also have a completely different test for starch too. Let's look at each of these tests and work out when we need to use each.
Reducing Sugar
A reducing sugar acts as a reducing agent. This means that it reduces other compounds.
Remember that a reduction reaction is the gain of electrons, loss of oxygen or gain of hydrogen. As reducing agents, these sugars are oxidised.
Examples of reducing sugars are;
glucose
fructose
lactose
maltose
To test for reducing sugars, we use Benedict's solution. Benedict's solution is a complex mixture of sodium carbonate, sodium citrate, and copper (II) sulfate pentahydrate.
Again, you do not need to know this for your exams!
Let's have a look at how we can use Benedict's solution to test for reducing sugars;
Add Benedict's solution to the substance in question
Heat the solution in a water bath
A positive test for reducing sugars using Benedict's solution shows a colour change over time. This includes a change of green, yellow, orange, red or brown - depending on how much glucose is present. The solution changes from blue to brick red if we look at the overall colour change. If enough reducing sugar is present, a brick red precipitate (ppt) is formed.
A precipitate is a suspension of particles in a liquid formed when a dissolved substance reacts to form an insoluble substance. We can sometimes dissolve a precipitate by adding extra solvent.
In the Benedict's test, whether a precipitate is formed depends on how much reducing sugar is present. This makes the test semi-quantitative because the more reducing sugar there is, the greater the likelihood that a brick red (red/brown) predicate will be formed.
We need to use a water bath when testing for reducing sugars using the Benedict's solution. Remember to phrase this as 'heating' the solution in a water bath, rather than just adding the solution to a water bath, as this shows the examiner that you understand the necessity of heating the solution.
Water baths are the method of choice here because they allow us to maintain a specific temperature, especially with the use of a thermometer, better than a Bunsen burner would.
Non-Reducing Sugar
Non-reducing sugars do not reduce other compounds/substances. Examples of non-reducing sugars are;
Let's have a look at how we can test for non-reducing sugars;
Add dilute hydrochloric acid (HCl) to the substance in question
Heat in a water bath at boiling point
Neutralise the solution with sodium hydrogen carbonate
Using a suitable indicator (such as litmus paper), identify when the solution has been neutralised, then add more sodium hydrogen carbonate as the conditions need to be alkaline for the Benedict’s test to work.
Then carry out Benedict’s test as normal
You can see this test in full in our 'reducing sugars' section!
If a colour change occurs from blue to a brick red precipitate, a non-reducing sugar is present.
Neutralisation reactions produce salt and water. The production of water in these reactions is the reason behind the name of the reaction. Water has a pH of around 7 - which we describe as neutral. These reactions take place when we mix an acid and a base.
If we know that our substance is sugar, but we are not sure whether it is a reducing or non-reducing sugar, we can simply carry out a Benedict's test to determine which type of sugar is present. If the sugar causes a colour change with Benedict's without the addition of sodium hydrogen carbonate or dilute hydrochloric acid, then it is a reducing sugar. If it only has a colour change after the addition of these substances, then it is a non-reducing sugar.
Starch
Starch is another type of carbohydrate, like the reducing sugars and non-reducing sugars we have already discussed. However, we use a different solution to test for starch, as starch is too complex and large of a sugar.
Carbohydrates can either be Monosaccharides, disaccharides or Polysaccharides.
Polysaccharides are large, complex sugars, whilst Monosaccharides and disaccharides are small, simple sugars. Benedict's solution only works for small sugars, such as monosaccharides and some disaccharides. We have to use different tests for larger complex sugars such as polysaccharides and some disaccharides. For example, starch is a polysaccharide, so we have to use iodine.
As mentioned in the deep dive above, we have to use iodine to test for starch. Let's have a look at how to use iodine to test for starch;
Add the substance in question to a solution (for example, potatoes)
Add drops of iodine to the food in question
A positive result for starch will result in a colour change from yellow/brown to blue/black.
A negative result remains yellow/brown, with no colour change being observed.
When we talk about the colours observed in colour changes within different reactions, you will see a lot of hyphens (-) and forward slashes (/). So, what's the difference? A forward slash indicates that we can either or.
For yellow/brown, we might see yellow or brown and could write either in the answer in an exam. We could also write yellow/brown as the answer. On the other hand, the presence of a hyphen describes the colour. So, the flame test for copper is described as blue-green. We cannot pick blue or green here as an answer; we need to write green-blue or blue-green, as these describe the colour.
Testing for Biomolecules in the Lab
When we test for biomolecules in a laboratory, for example, when testing for these different molecules in school practicals, there are a few safety precautions that we have to follow. Let's look at some of the safeguards we must take and why we must take them.
Irritants can cause our skin to become itchy if they come into contact with our skin.
Corrosive chemicals can damage or destroy other substances if it comes into contact with them.
Flammable substances quickly light on fire.
Let's look at some of the substances discussed above, the potential risks of using them, and how we can reduce those risks as much as possible to carry out a safe biomolecule test in the lab.
Table showing the hazards and precautions associated with different food tests:
| Solution | Risk | Precautions |
Benedict's solution | Irritant | Avoid contact with skin and eyes; wear safety goggles |
Iodine solution | Irritant | Avoid contact with skin and eyes; wear safety goggles |
Biuret solution | Biuret solution A is corrosive, and biuret B is an irritant | Avoid contact with skin and eyes; wear safety goggles |
Sudan III solution | Flammable | Do not hold near a lit flame |
Ethanol solution (Emulsion) | Flammable | Do not hold near a lit flame |
When looking at the precautions and risks above, try to remember the biomolecules that each of the solutions are used to test for, and try to remember the results of a positive and negative test!
Testing for Biomolecules - Key takeaways
- We can test for proteins, fats (lipids) and carbohydrates (reducing and non-reducing sugars; starch)
- Each of these tests causes colour changes when positive.
- We describe these tests as being qualitative, as they only show us the presence of a specific molecule, not the quantity of the molecule.
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