By the end of this chapter, you will be able to:
Mastering these skills will help you contribute confidently to important biochemical analyses in real laboratory settings.
Enzymes are biological catalysts, mostly globular proteins, that speed up chemical reactions in living cells without being used up. Because thousands of enzymes exist, they are classified and named in a standard way by the International Union of Biochemistry and Molecular Biology (IUBMB), according to the type of reaction they catalyse. Each enzyme receives a four-part Enzyme Commission (EC) number and a systematic name, alongside its common name. Enzymes can also be grouped by where they act, by their chemical make-up and by the substrate they act on. Example: the quality control laboratory of a sugar factory in Chemelil uses invertase, a hydrolase, to measure sucrose in cane juice samples.
| EC class | Name | Type of reaction | Examples |
|---|---|---|---|
| EC 1 | Oxidoreductases | Oxidation and reduction; transfer of hydrogen or electrons | Dehydrogenases, oxidases, catalase |
| EC 2 | Transferases | Transfer of a functional group (methyl, amino, phosphate) from one molecule to another | Transaminases (ALT, AST), kinases such as hexokinase |
| EC 3 | Hydrolases | Breaking bonds by adding water (hydrolysis) | Amylase, lipase, pepsin, sucrase, urease |
| EC 4 | Lyases | Adding or removing groups to form or break double bonds, without water or oxidation | Aldolase, carbonic anhydrase, decarboxylases |
| EC 5 | Isomerases | Rearranging atoms within a molecule to form an isomer | Phosphoglucose isomerase, triose phosphate isomerase |
| EC 6 | Ligases (synthetases) | Joining two molecules using energy from ATP | DNA ligase, pyruvate carboxylase |
| EC 7 | Translocases | Moving ions or molecules across membranes | Sodium-potassium pump (Na⁺/K⁺-ATPase), ATP synthase |
Class 7 was added in 2018; older books list six classes.
Isoenzymes (isozymes) are different forms of an enzyme that catalyse the same reaction but differ in structure and properties and are found in different tissues. Lactate dehydrogenase has five isoenzymes (LDH-1 to LDH-5); measuring which ones rise in the blood helps locate damage to the heart, liver or muscle.
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| Tools & Equipment | Materials |
|---|---|
| Mortar and pestle | Raw fresh pineapple fruit (source of bromelain enzyme) |
| Syringes (5 ml) | Raw fresh papaya fruit (source of papain enzyme) |
| Clean test tubes (15 ml) | Distilled water |
| Labeling tape and marker pen | Filter paper squares |
| Scalpel or sharp knife | |
| Beaker (100 ml) |
| S/N | Item | Quantity |
|---|---|---|
| 1 | Raw fresh pineapple fruit (source of bromelain enzyme) | 1 medium pineapple per 3 Candidates |
| 2 | Raw fresh papaya fruit (source of papain enzyme) | 1 medium papaya per 3 Candidates |
| 3 | Mortar and pestle | 1 set per Candidate |
| 4 | Distilled water | 200 ml per Candidate |
| 5 | Syringes (5 ml) | 1 per Candidate |
| 6 | Clean test tubes (15 ml) | 4 per Candidate |
| 7 | Labeling tape and marker pen | 1 set per Candidate |
| 8 | Scalpel or sharp knife | 1 per Candidate |
| 9 | Filter paper squares | 4 per Candidate |
| 10 | Beaker (100 ml) | 1 per Candidate |
| Items to be Evaluated | Marks Available | Marks Obtained | Comments |
|---|---|---|---|
| TASK 1: Preparation of Pineapple Enzyme Solution | |||
| Wore appropriate personal protective equipment (Lab coat, face mask, low-heeled closed shoes) (Award 1 mark for each correctly worn item) | 3 | ||
| Selected fresh pineapple sample and washed it with distilled water (Award 2 marks for correct washing procedure) | 2 | ||
| Used scalpel to cut small pieces (~5g) of pineapple flesh (Award 2 marks for correct cutting and quantity) | 2 | ||
| Placed pineapple pieces in mortar and added 10 ml distilled water (Award 2 marks for correct sample and volume) | 2 | ||
| Crushed pineapple thoroughly using pestle to obtain a fine pulp (Award 3 marks for thorough crushing) | 3 | ||
| Filtered the pulp through filter paper into clean test tube to obtain clear enzyme solution (Award 3 marks for proper filtration and clear solution) | 3 | ||
| Used syringe to transfer 10 ml of enzyme solution into labeled test tube (Award 3 marks for correct transfer and labeling) | 3 | ||
| Sub-Total | 18 | ||
| TASK 2: Preparation of Papaya Enzyme Solution | |||
| Selected fresh papaya sample and washed it with distilled water (Award 2 marks for correct washing procedure) | 2 | ||
| Used scalpel to cut small pieces (~5g) of papaya flesh (Award 2 marks for correct cutting and quantity) | 2 | ||
| Placed papaya pieces in mortar and added 10 ml distilled water (Award 2 marks for correct sample and volume) | 2 | ||
| Crushed papaya thoroughly using pestle to obtain a fine pulp (Award 3 marks for thorough crushing) | 3 | ||
| Filtered the pulp through filter paper into clean test tube to obtain clear enzyme solution (Award 3 marks for proper filtration and clear solution) | 3 | ||
| Used syringe to transfer 10 ml of enzyme solution into labeled test tube (Award 3 marks for correct transfer and labeling) | 3 | ||
| Sub-Total | 15 | ||
| TASK 3: Housekeeping | |||
| Cleaned workspace and equipment used (Award 1 mark for proper cleaning) | 1 | ||
| Disposed of waste materials according to laboratory procedures (Award 1 mark for proper disposal) | 1 | ||
| Organized and stored equipment and materials appropriately (Award 1 mark for good organization) | 1 | ||
| Sub-Total | 3 | ||
| PRODUCT CHECKLIST | |||
| Pineapple enzyme solution is clear, approximately 10 ml, correctly labeled and free of solid debris (Award 5 marks for clear, correct volume and labeling) | 5 | ||
| Papaya enzyme solution is clear, approximately 10 ml, correctly labeled and free of solid debris (Award 5 marks for clear, correct volume and labeling) | 5 | ||
| Sub-Total | 10 | ||
| GRAND TOTAL | 46 | ||
Type: Individual
| Tools & Equipment | Materials |
|---|---|
| Measuring cylinder 100 ml | Distilled water |
| Analytical balance | Sucrose powder |
| Beakers 250 ml | Glucose powder |
| Glass stirring rods | Fructose powder |
| Volumetric flasks 100 ml | Phosphate buffer pH 7.0 |
| Dropper pipettes | |
| Labels and marker pens | |
| pH meter |
| S/N | Item | Quantity |
|---|---|---|
| 1 | Distilled water | 500 ml per Candidate |
| 2 | Sucrose powder | 10 g per Candidate |
| 3 | Phosphate buffer pH 7.0 | 100 ml per Candidate |
| 4 | Glucose powder | 5 g per Candidate |
| 5 | Fructose powder | 5 g per Candidate |
| 6 | Measuring cylinder 100 ml | 1 Pc per Candidate |
| 7 | Analytical balance | 1 Pc per 5 Candidates |
| 8 | Beakers 250 ml | 2 Pcs per Candidate |
| 9 | Glass stirring rods | 1 Pc per Candidate |
| 10 | Volumetric flasks 100 ml | 3 Pcs per Candidate |
| 11 | Dropper pipettes | 2 Pcs per Candidate |
| 12 | Labels and marker pens | Sufficient per Candidate |
| 13 | pH meter | 1 Pc per 5 Candidates |
| 14 | Protective gloves | 1 Pair per Candidate |
| 15 | Lab coat | 1 Pc per Candidate |
| 16 | Face mask | 1 Pc per Candidate |
| 17 | Low-heeled closed shoes | 1 Pair per Candidate |
| Items to be Evaluated | Marks Available | Marks Obtained | Comments |
|---|---|---|---|
| TASK 1: Preparation of 0.1 M Sucrose Substrate Solution | |||
| Wore appropriate PPE: lab coat, face mask, gloves, low-heeled closed shoes (Award 1 mark for each correctly worn item) | 4 | ||
| Weighed 3.42 g of sucrose powder accurately using analytical balance (Award 3 marks for correct mass and use of balance) | 3 | ||
| Transferred sucrose into a 100 ml volumetric flask (Award 2 marks for correct transfer) | 2 | ||
| Added phosphate buffer pH 7.0 to the mark carefully (Award 3 marks for correct volume and use of buffer) | 3 | ||
| Mixed solution thoroughly using glass rod until fully dissolved (Award 2 marks for proper mixing) | 2 | ||
| Labelled the flask clearly as '0.1 M Sucrose substrate pH 7.0' (Award 2 marks for clear and correct labelling) | 2 | ||
| Sub-Total | 16 | ||
| TASK 2: Preparation of 0.1 M Glucose Substrate Solution | |||
| Weighed 1.80 g of glucose powder accurately (Award 3 marks for correct mass and use of balance) | 3 | ||
| Transferred glucose into a 100 ml volumetric flask (Award 2 marks for correct transfer) | 2 | ||
| Added phosphate buffer pH 7.0 to the mark (Award 3 marks for correct volume and use of buffer) | 3 | ||
| Mixed solution thoroughly until dissolved (Award 2 marks for proper mixing) | 2 | ||
| Labelled the flask clearly as '0.1 M Glucose substrate pH 7.0' (Award 2 marks for clear and correct labelling) | 2 | ||
| Sub-Total | 12 | ||
| TASK 3: Preparation of 0.1 M Fructose Substrate Solution | |||
| Weighed 1.80 g of fructose powder accurately (Award 3 marks for correct mass and use of balance) | 3 | ||
| Transferred fructose into a 100 ml volumetric flask (Award 2 marks for correct transfer) | 2 | ||
| Added phosphate buffer pH 7.0 to the mark (Award 3 marks for correct volume and use of buffer) | 3 | ||
| Mixed solution thoroughly until dissolved (Award 2 marks for proper mixing) | 2 | ||
| Labelled the flask clearly as '0.1 M Fructose substrate pH 7.0' (Award 2 marks for clear and correct labelling) | 2 | ||
| Sub-Total | 12 | ||
| PRODUCT CHECKLIST | |||
| Each substrate solution volume is exactly 100 ml (Award 3 marks if all three solutions meet this criterion) | 3 | ||
| Each substrate solution concentration is approximately 0.1 M based on correct mass (Award 3 marks if all three solutions have correct concentration) | 3 | ||
| Solutions are clear and free from undissolved particles (Award 3 marks if all three solutions are clear) | 3 | ||
| Each solution is correctly labelled with substrate name and pH (Award 3 marks if all three solutions are correctly labelled) | 3 | ||
| Sub-Total | 12 | ||
| GRAND TOTAL | 52 | ||
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