2.1 Enzymes and Digestion (1)
- Oesophagus - Carries food from the mouth to the stomach. Only adapted for transport (not absorption or digestion) so is made up of a thick muscular wall.
- Stomach - Muscular sac with an inner layer that produces enzymes. Role is to store and digest food, especially proteins, and so it has glands which produce enzymes to digest protein. Other glands in its stomach wall produce mucus to prevent the stomach from being digested by its own enzymes.
- Small Intestine - Long muscular tube. Food is further digested by enzymes produced by its walls and by secretary glands. Inner walls are folded into villi, giving them a large SA. The SA of these villi is further increased by microvilli (tiny projections on the epithelial cells of each villus). This adapts the small intestine for its role of absorbing the products of digestion into the bloodstream.
- Large Intestine - Absorbs water. Most of the water that is reabsorbed comes from the secretions of the digestive glands. The food within the large intestine therefore becomes drier and thicker, forming faeces.
- Rectum - Final section of the intestines. Faeces are stored here before being removed via the **** in a process called egestion.
- Salivary Glands - Secretion - Amylase. Passed via a duct into the mouth.
- Pancreas - Found below the stomach. Secretion - Pancreatic juice. Contains protease (digest proteins), lipase (digest lipids) and amylase (digest starch).
2.1 Enzymes and Digestion (2)
Digestion takes place in two stages: a) Physical Breakdown and b) Chemical Digestion.
a) Food is broken down by structures like the teeth, making it possible to ingest the food as well as provide a large surface area for chemical digestion. Food is churned by muscles in the stomach wall which also physically breaks it up.
- Carried out by enzymes named hydrolases because they function by hydrolysis.
- As enzymes are specific, more than 1 enzyme is needed to hydrolyse a large molecule.
- There are different types of digestive enzymes, three of which are particularly important:
- Carbohydrases - Break down carbohydrates into monosaccharides.
- Lipases - Break down lipids (fats and oils) into glycerol and fatty acids.
- Proteases - Break down proteins, ultimately to amino acids.
The products of digestion are absorbed from the small intestine into the blood where they are carried to different parts of the body and often built up again into large molecules (although not necessarily of the same type as the ones from which they derived). They are incorporated into body tissues and/or used in processes within the body. This is called assimilation.
2.2 Carbohydrates - Monosaccharides (1)
Carbon atoms readily form bonds with other carbon atoms, allowing a sequence of carbon atoms of various lengths to be built up. These form a backbone along which other atoms can join, hence permitting an immense number of different types and sizes of molecule, all based on carbon. These carbon-containing molecules are known as organic molecules.
- Monomer - An individual molecule making up a chain (polymer)
- Polymer - Longer chains of repeating monomer units, e.g. carbohydrates.
In carbohydrates, the basic monomer unit is a sugar, otherwise known as a saccharide.
- Single monomer - Monosaccharide
- Pair of monomers - Disaccharide
- Monosaccharides combined in larger numbers - Polysaccharides
- Dissolve in water to form sweet-tasting solutions and are white crystalline solids.
- Have the general formula (CH2O)n, where n can be any number from 3-7.
- e.g. Glucose (C6H1206). It is a hexose (6-carbon sugar). So n was 6.
2.2 Test for Reducing Sugars (Benedict's Test)
- All monosaccharides and some disaccharides (e.g. maltose), are reducing sugars.
- Reduction is a chemical reaction involving the gain of electrons (OILRIG), hence a reducing sugar is one that can donate electrons to (or reduce) another chemical, in this case Benedict's reagent.
- Benedict's reagent is an alkaline solution of blue copper (II) sulfate. When a reducing sugar is heated with Benedict's reagent, it forms an insoluble red precipitate of copper (I) oxide.
- The test is carried out as follows:
- 1. Add 2cm cubed of the food sample to be tested in a test tube. If the sample is not in liquid form, grind it up with water.
- 2. Add an equal volume of Benedict's reagent.
- 3. Heat the mixture in a gently boiling water bath for 5 minutes.
- No Reducing Sugar Present - Blue Precipitate/Solution
- Very Low Concentration of Reducing Sugar - Green Precipitate/Solution
- Low Concentration of Reducing Sugar - Yellow Precipitate/Solution
- Medium Concentration of Reducing Sugar - Brown Precipitate/Solution
- High Concentration of Reducing Sugar - Red Precipitate/Solution
2.3 Disaccharides and Polysaccharides
When combined in pairs, monosaccharides form a disaccharide. For example:
- Glucose linked to glucose forms maltose
- Glucose linked to fructose forms sucrose
- Glucose linked to galactose forms lactose
When the monosaccharides join, a molecule of water is removed and the reaction is therefore called a condensation reaction. The bond that is formed is called a glycosidic bond.
When water is added to a disaccharide under suitable conditions, it breaks the glycosidic bond releasing the constituent monosaccharides. This is called hydrolysis.
- As polysaccharides are very large molecules, they are insoluble. This feature makes them suitable for storage. When hydrolysed polysaccharides break down into disaccharides or monosaccharides.
- Some polysaccharides, such as cellulose, are not used for storage but give structural support to plant cells.
2.3 Test for Non-Reducing Sugars
Other disaccharides e.g. sucrose are known as non-reducing sugars because they do not change the colour of Benedict's reagent when heated with it.
To detect a non-reducing sugar it must first be broken down into its monosaccharide components by hydrolysis. This is done by carrying out the Benedict's test.
After it is confirmed there is no reducing sugar present, i.e. if the reagent stays blue, the process continues as follows:
- 1. Add another 2cm cubed of the food sample to 2cm cubed of dilute hydrochloric acid in a test tube and place the test tube in a gently boiling water bath for 5 minutes. The dilute acid will hydrolyse any disaccharide present into its constituent monosaccharides.
- 2. Slowly add some sodium hydrogencarbonate solution to the test tube in order to neutralise the acid. (Benedict's reagent will not work in acidic conditions). Test with pH paper to check the solution is alkaline.
- 3. Re-test the resulting solution by heating it with 2cm cubed of Benedict's reagent in a gently boiling water bath for 5 minutes.
- 4. If a non-reducing sugar was present in the original sample, the Benedict's reagent will now turn orange-brown.
2.3 Test for Starch
Starch is a polysaccharide found in many parts of plants in the form of small granules or grains e.g. starch grains in chloroplasts.
It is formed by the linking of 200-100 000 alpha glucose molecules by glycosidic bonds in a series of condensation reactions.
Starch is easily detected by its ability to change the colour of the iodine in potassium iodide solution from yellow to blue-black.
The test is carried out at room temperature.
The test is carried out as follows:
- Place 2cm cubed of the sample being tested into a test tube (or add two drops of the sample into a depression on a spotting tile).
- Add two drops of iodine solution and shake or stir.
- The presence of starch is indicated by a blue-black colouration.
2.4 Disaccharide Digestion (1)
In addition to the digestion of maltose (starch digestion), two other dissacharides in the diet need to be broken down:
- Usually contained within cells which must be physically broken down by teeth to release it.
- The sucrose passes through the stomach and into the small intestine whose epithelial lining produces the enzyme sucrase. Sucrase hydrolyses the single glycosidic bond in sucrose to produce glucose and fructose.
- Is the sugar found in milk and milk products such as yoghurt and cheese.
- Digested in the small intestine whose epithelial lining produces the enzyme lactase.
- Lactase hydrolyses the glycosidic bond in lactose to produce glucose and galactose.
2.4 Lactose Intolerance
- Milk is the only food of babies, hence they produce large amounts of lactase to digest the lactose.
- As milk forms a smaller part of the diet in adults, the production of lactase naturally diminishes during childhood. In some people this reduction is so great that they end up producing little or no lactase.
- Modern storage and distribution methods mean adults now consume milk and milk products in greater quantities, therefore some people do not produce enough lactase to digest all the lactose they consume.
- When the undigested lactose reaches the large intestine, it is broken down by micro organisms leading to large volumes of gas being produced, resulting in bloating/diarrhoea/cramps/nausea.
- Avoiding lactose-containing foods will manage the symptoms but that presents the challenge of taking in sufficient calcium in the absence of milk. This can be resolved by taking in foods rich in calcium or by adding lactase to milk before drinking it.
- Babies with LI need to be fed special non-milk food rich in calcium and vitamin D.
N.B. Micro organisms produce this gas by respiration, therefore the gas is unlikely to be carbon dioxide as this is a product of aerobic respiration, whereas conditions in the large intestine are anaerobic.
2.5 Proteins - Structure of an Amino Acid (1)
- Amino acids are the basic monomer units which combine to make up a polymer called a polypeptide.
- Polypeptides can be combined to form proteins.
- Every amino acid has a central carbon atom to which are attached 4 different chemical groups:
- Amino group (NH2)
- Carboxyl group (COOH) - An acidic group
- Hydrogen atom (H)
- R Group - A variety of different chemical groups. Each amino acid has a different R Group.
- In the same way that monosaccharide monomers combine to form disaccharides, so amino acid monomers combine to form dipeptides. In the same way a glycosidic bond can be broken by hydrolysis, so can the peptide bond of a dipeptide.
- A water molecule is removed in a condensation reaction.
- The water is made by combining an OH from the carboxyl group of one amino acid with an H from the amino acid group of another amino acid. (creating H2O)
- The two acids them become linked by a new peptide bond between the carbon atom of one amino acid and the nitrogen atom of the other.
2.5 Test for Proteins (Biuret Test)
- The most reliable test
- Detects peptide links
- Is performed as follows:
- Place a sample of the solution to be tested in a test tube and add an equal volume of sodium hydroxide solution at room temperature.
- Add a few drops of very dilute (0.05%) copper (II) sulfate solution and mix gently.
- A purple colouration indicates the presence of peptide bonds and therefore a protein.
- If no protein is present, the solution remains blue.
2.5 Proteins - Primary Structure
- Through a series of condensation reactions, many amino acids can be joined together in a process called polymerisation resulting in a polypeptide.
- The sequence of amino acids in a polypeptide chain forms the primary structure in any protein.
- Since polypeptides have many (usually hundreds) of the 20 naturally occuring amino acids joined in any sequence, there is almost a limitless number of possible combinations and therefore types of primary protein structure.
- The primary structure of a protein determines its ultimate shape and therefore its function. A change in just a single amino acid in this primary sequence can lead to a change in the shape of the protein and may stop it carrying out its function. In other words, a proteins shape is very specific to its function. Change its shape, and it will function less well, if at all.
- A simple protein may consist of a single polypeptide chain. More commonly a protein is made up of a number of polypeptide chains.
2.5 Proteins - Secondary Structure
- The linked amino acids that make up a polypeptide possess both NH and C=O groups on either side of every peptide bond.
- The hydrogen of the NH group has an overall positive charge while the oxygen of the C=O group has an overall negative charge. Therefore these groups form weak bonds, called hydrogen bonds.
- The protein's secondary structure is formed when the long polypeptide is twisted into a 3D shape, such as the coil known as the alpha helix, or the beta-pleated sheet.
2.5 Tertiary & Quarternary Structure
- The alpha helices of the secondary structure can be twisted and folded even more to give the complex and unique 3D structure of each protein, known as the tertiary structure.
- This structure is maintained by a number of different bonds, including:
- Disulfide bonds - Fairly strong and therefore not easily broken down.
- Ionic bonds - Formed between any carboxyl and amino groups that are not involved in forming peptide bonds. They are weaker than disulfide bonds and are easily broken by changes in pH.
- Hydrogen bonds - Numerous but easily broken.
The 3D shape of each protein makes it distinctive and allows it to recognise and be recognised by other molecules.
This structure arises from the combination of a number of different polypeptide chains and associated non-protein (prosthetic) groups into a large protein molecule.
2.5 - Protein Shape & Function
- A protein's role depends on their molecular shape, which can be of two basic types:
- Fibrous Proteins - Have structural functions, e.g. collagen.
- Globular Proteins - Carry out metabolic functions, e.g. enzymes and haemoglobin.
- Form long parallel chains which are linked by cross-bridges and therefore form very stable molecules, e.g. collagen.
- Collagen is found in tendons. Tendons join muscle to bone. When a muscle contracts, the bone is pulled in the direction of the contraction.
- The individual collagen polypeptide chains in the fibres are held together by cross-linkages between amino acids of adjacent chains.
- The points where one collagen molecule ends and the next begins are spread throughout the fibre rather than all being in the same position along it.
2.6 Enzyme Action
- Enzymes are globular proteins that act as catalysts ( they alter the rate of a chemical reaction without undergoing permanent change themselves).
- They can be reused repeatedly and therefore are effective in small quantities
Enzymes work by lowering the activation energy level of a reaction, therefore enzymes allow reactions to take place at a lower temperature than normal (e.g. at the human body temperature of 37 degrees).
Activation energy - The minimum amount of energy needed to activate/kickstart a reaction
- Although an enzyme molecule is large, only a small region is functional (the active site), which forms a small hollow depression within the molecule.
- The molecule on which the enzyme acts is called the substrate which fits into the depression to form the enzyme substrate complex.
- The substrate molecule is held within the active site by bonds which temporarily form between certain amino acids of the active site and groups on the substrate molecule.
2.6 The Lock and Key Model
Proposes that enzymes work in the same way as a key fits a lock; each key has a specific shape that fits an operates only a single lock. In a similar way, a substrate will only fit the active site of one particular enzyme.
+ Supported by the observation that enzymes are specific in the reactions they catalyse.
- One limitation of this model is that the enzyme, like a lock, is considered to be a rigid structure. However, scientists have observed that other molecules could bind to enzymes at sites other than the active site, which suggested that the enzyme's shape was being altered by the binding molecule. In other words, its structure was flexibile, not rigid.
2.6 Induced Fit Model
- More refined model than the lock and key
- Proposes that in the presence of the substrate the enzyme may change shape slightly to fit the profile of the substrate. i.e. the enzyme is flexible.
- As it changes its shape, the enzyme puts a strain on the substrate molecule. This strain distorts a particular bond and therefore lowers the activation energy needed to break the bond.
+ Explains how other molecules can affect enzyme activity
+ Explains how the activation energy is lowered
2.7 Factors Affecting Enzyme Action
For an enzyme to work it must:
- Come into physical contact with its substrate
- Have an active site which fits the substrate
Almost all factors that influence the rate at which an enzyme works do so by affecting one or both of the above.
When measuring the progress of an enzyme-catalysed reaction, we measure its time-course. ("how long it takes for a particular event to run its course")
The two 'events' most frequently measured are:
- the formation of the products of the reaction e.g. the volume of oxygen produced when catalase acts on hydrogen peroxide
- the disappearance of the substrate e.g. the reduction in concentration of starch when it is acted upon by amylase.
2.7 Effect of Temperature on Enzyme Action
- A rise in temperature increases the kinetic energy of the molecules, causing the molecules to move around more rapidly and collide with each other more often.
- Therefore, in an enzyme-catalysed reaction, the enzyme and substrate molecules come together more often in a given time (i.e. more enzyme-substrate complexes formed), so that the rate of reaction increases.
- The temperature rise also begins to cause the hydrogen and other bonds in the enzyme to break, causing the enzyme and its active site to change shape. As a result, the substrate fits less easily, slowing the rate of reaction. (For many human enzymes this may begin at around 45 degrees).
- At some point (usually around 60 degrees), the enzyme is so disrupted that it stops working altogether. Therefore it becomes denatured - a permanent & irreversible change.
- Many human enzymes have an optimum temperature of 40 degrees despite our body temperatures being 37 degrees. They evolved this way because:
- Other proteins except enzymes may be denatured at higher temperatures.
- At higher temperatures any further rise e.g. from illness may denature the enzymes.
- Although higher body temperatures would increase the metabolic rate, the advantages are offset by the additional energy (food) that would be needed to maintain this.
2.7 Effect of pH on Enzyme Action
- The pH of a soultion is a measure of its hydrogen ion concentration.
- Each enzyme has an optimum pH.
- The pH effects how an enzyme works in the following ways:
- a change in pH alters the charges on the amino acids that make up the active site of the enzyme. Therefore the substrate can no longer become attached to the active site and hence the enzyme substrate complex cannot be formed.
- A change in pH can cause the bonds that maintain the enzyme's tertiary structure to break, so the enzyme changes shape. This may alter the shape of the active site and the substrate may no longer fit. The enzyme has been denatured.
- This is because even small changes in pH change the arrangement of the active site of an enzyme. The arrangement is party determined by the hydrogen and ionic bonds between NH2 and COOH groups of the polypeptides that make up the enzyme. The change in H+ ions affects this bonding, causing the active site to change shape.
N.B. As pH fluctuations inside organisms are usually small, they are far more likely to reduce an enzyme's activity than to denature it.
2.7. Effect of Substrate Concentration
- If the amount of enzyme is fixed at a constant level and substrate is slowly added, the rate of reaction increases in proportion to the amount of substrate added.
- This is because at low substrate concentrations, the enzyme molecules have a limited number of substrate molecules to colide with i.e. the enzymes are not working to full capacity.
- As more substrate is added, the active sites gradually become filled until the rate of reaction is at its maximum (Vmax). After that, the addition of more substrate will have no effect.
2.8 Enzyme Inhibition
Enzyme Inhibitors - "Substances that directly or indirectly interfere with the functioning of the active site of an enzyme and so reduce its activity"
Most inhibitors only make temporary attachments to the active site. These are called reversible inhibitors and are two types:
1. Competitive Inhibitors (which bind to the active site of the enzyme):
- Have a molecular shape similar to that of the substrate which allows them to occupy the active site on an enzyme, therefore they compete with the substrate for the available active sites.
- The difference between the concentration of the inhibitor and the concentration of the substrate determines the effect that this has on enzyme activity.
- If the substrate concentration is increased, the effect of the inhibitor is reduced. The inhibitor is not permanently bound to the active site so when it leaves, another molecule can take its place. This could be a substrate or inhibitor molecule.
- Sooner or later, all the substrate molecules will occupy an active site, but the greater the concentration of inhibitor, the longer this will take.
2.7 Enzyme Inhibition
2. Non-Competitive Inhibitors (which bind to the enzyme at a position other than the active site):
- Upon attaching to the enzyme, the inhibitor alters the shape of the enzyme's active site so that substrate molecules can no longer occupy it and hence the enzyme cannot function.
- As the substrate and inhibitor are not competing for the same site, an increase in substrate concentration does not reduce the effect of the inhibitor.