PRACTICAL SKILLS 1

• Factors that affect enzyme activity

  • Enzyme concentration
  • Substrate concentration
  • Temperature
  • pH
• Setting up a control in a practical measuring enzyme activity

  Replace the enzyme solution with distilled water or boiled enzyme solution
• Using results to find the initial rate of reaction in practical measuring enzyme activity

  Plot results on a graph of ‘rate of reaction’ against ‘time’
• Practical procedure to measure the effect of temperature on enzyme activity using trypsin and milk


  Immerse equal volumes of trypsin and milk in different test tubes in a water-bath for 5 minutes, mix together, start timing, record time for milk to be completely hydrolysed, test at least 5 temperatures with 3 repeats each
• Calculating the rate of reaction from time


  Rate of reaction=1/time
• Effect of temperature on enzyme activity

  As temperature increases, kinetic energy increases leading to more ES complexes forming. Rate of reaction increases up to the optimum temperature. Beyond that, bonds in the enzyme tertiary structure break, changing the shape of the active site, leading to a decrease in the rate of reaction
• Risk associated with handling enzymes
• Where cells undergoing mitosis can be found in plants
• Mitotic index


  The ratio of cells undergoing mitosis to the total number of cells in a sample
• Procedure to prepare a root tip slide

  Warm 1M HCI, cut a root tip, add to HCI, wash with distilled water, add stain to make chromosomes visible
• Formula for the mitotic index
• Hazards and precautions for reagents used in preparing a root tip slide
• Purpose of calibration curves
• Using a calibration curve to find the concentration of plant tissue


  Plot a calibration curve of percentage change in mass against concentration, find the x-intercept where the plant tissue is isotonic to the sucrose solution
• What occurs when plant tissue is placed in a hypotonic solution

  Water moves into the plant tissue by osmosis, increasing its mass
• What occurs when plant tissue is placed in a hypertonic solution

  Water moves out of the plant tissue by osmosis, decreasing its mass
• Procedure of investigating osmosis using potato tissue

  Make a dilution of 1M sucrose, cut potato chips, place in test tubes with sucrose solution, leave for 20 minutes, weigh before and after, calculate percentage change in mass
• Reason for using percentage change in mass in investigating osmosis
• Indication of the x-intercept of the calibration curve
• Explanation of the change in mass in the potato chips
• The x-intercept of the calibration curve indicates the concentration that is isotonic to the solution used
• Change in mass in the potato chips

  1. Potato chips with concentration lower than the sucrose solution lose mass as there is a net movement of water out of the cells

  2. Potato chips with concentration higher than the sucrose solution gain mass as there is a net movement of water into the cells
• Potato chips are dabbed dry after removing from the sucrose solution to remove any excess water clinging to their surface
• Controlled variables of the practical

  • Volume of sucrose solution
  • Size of potato chips
  • Length of time left in solution
• Factors that affect the permeability of cell membranes

  1. Temperature

  2. Concentration of solvents (ethanol)
• How beetroot is used to measure the permeability of cell membranes

  The higher the permeability, the more red pigment leaks out into the surrounding solution within a given time. A colorimeter can be used to determine the absorbance, hence the concentration of pigment
• Procedure to investigate the effect of temperature on permeability of cell membrane

  1. Cut beetroot into 6 identical cubes with scalpel

  2. Place each cube in a different test tube with equal volumes of distilled water

  3. Place each test tube into water baths ranging from 30-80 degrees. Leave for 20 minutes

  4. Filter each solution out into a cuvette and measure the absorbance using a colorimeter
• Safety hazards involved in testing the effect of ethanol concentration on membrane permeability
• Effect of temperature on membrane permeability

  Increasing temperature results in increased membrane permeability
• Effect of ethanol concentration on membrane permeability

  Increasing ethanol concentration leads to increased membrane permeability
• How label lines in a diagram should be drawn

  With a ruler, no arrows, without crossing other label lines, in pencil
• How a diagram should be drawn

  1. Large diagram-at least half the space given

  2. No shading, single continuous lines (no sketching) in pencil
• Aseptic techniques


  • Wipe down surfaces with antibacterial cleaner, before and after experiment
  • Use a Bunsen burner in the work space so that convection currents draw microbes away from the culture
  • Flame the wire hoop before using it to transfer bacteria
  • Flame the neck of any bottles before using them to prevent any bacteria entering the vessel
  • Keep all vessels containing bacteria open for the minimum amount of time
  • Close windows and doors to limit air currents
• Bacteria is incubated at 25 degrees to prevent the growth of pathogens, which occurs at higher temperatures
• How to compare the effectiveness of different antibiotics applied to the same bacteria

  Measure the diameter and calculate the area of the zone of inhibition (clear zone) on the agar
• The zone of inhibition indicates the bacteria killed by the antibiotic. The larger the zone, the more effective the antibiotic. If an antibiotic has very little or no zone of inhibition, the bacteria is likely resistant to the antibiotic
• Hazards and precautions of the practical
• Why the lid should not be completely taped to the petri dish
• Describe the graph that can be plotted from the results of this practical