M1:09 Investigating Enzyme Action Flashcards
(24 cards)
What does an investigation into catalase entail? and what does the enzyme do?
it measures the rate at which H2o2 is used up, or the rate at which either water or oxygen gas is produced it catalyses the conversion of h202 into water and oxygen gas.
What is an independent and dependent variable?
the variable that is set by the investigator the variable that is measured by the investigator
What is a variable?
a variable is any factor that may change and therefore affect the reaction rate. Enzyme investigations require all variables to be controlled
What needs to be ensured before an investigation into enzymes can be carried out?
the enzyme and substrate must be brought to the required temperature separately (equilibration) to give the most accurate results
What is equilibration ?
the enzyme and substrate must be brought to the required temperature separately
What are the key variables when investigating enzymes?
temperature enzyme concentration substrate concentration pH value
How is temperature kept constant?
- carrying out enzyme-controlled reactions in a water bath with a thermostat- it a polystyrene sleeve is used as isulation can help keep temp constant
Why must temperature be controlled?
Room temp is too variable, and fluctuations in temp will affect the enzyme controlled reaction so readings taken will not reflect the action of the independent variable being tested.
Name 3 ways that enzyme concentration is controlled
- use accurately measured volumes of enzyme in solution - If you are using enzyme in a living tissue, accurate measurements of the mass of tissue are important - if you are using whole pieces of tissue, you must keep the surface area of the tissue constant as well as the mass.
Why must enzyme concentration be controller?
reaction rate depends on the concentration of enzyme molecules present. Using accurately measured volumes of enzyme solution gives a constant concentration of enzyme molecules
Why must you keep the surface area of tissue and mass constant
the number of enzyme molecules in contact with substrate molecules will affect the reaction rate if the surface areas of the pieces of tissue are different, the number of enzyme molecules exposed directly to the substrate will be different.
What is the method of keeping the substrate concentration constant?
use accurately measured volume or mass of substrate
Why must the substrate concentration be kept constant
as the reaction rate depends on the concentration of substrate molecules.
What method is used for keeping the pH value constant?
Use pH buffers- solutions that maintain pH at a set level by keeping the H+ concentration in solution constant
Why must the pH value be kept constant ?
as reaction rate depends on the pH because of its effect on the shape of the active site of the enzyme
What are the two main ways of measuring reaction rates?
- start the reaction then measure the concentration of product (or of the substrate used up) after a fixed period - monitor the reaction by taking readings of product formation (or of substrate used up) at a number of time intervals
What is the most suitable time scale usually for enzyme controlled reaction av
usually seconds or minutes
What is the calculation for rate of reaction ?
reaction rate = 1/time (or 100/time or 1000/time can be used to make the number friendlier )
which stage of the reaction should be compared if you are wanting to show the effect of any dependent variable?
initial reaction rates
How would you measure the initial reaction rates (off a graph)
plot then on a graph and take a tangent to the steepest portion of the graph and do x/y = rate in cm3 per second
Describe the steps of the investigation for effect of pH on amylase action
assay technique - cut wells into the starch-agar plate using a cork borer- into each well put the same volume of one of a range of pH buffer solutions - place the same amount of stock amylase solution into each well except 1 (add water to this one as a control) - incubate for 24hrs in a dry oven at 35deg - during this time the amylase will diffuse through the agar and catalyse the conversion of starch to maltose - flood the plate with iodine and rise with water (starch will turn blue/black) measure the diameter of the cleared zone, this shows how much substrate has been turned into product
Describe the way you can test the effect of temperature on catalase action
(catalase catalyses the break down of hydrogen peroxide into water and oxygen in potato tubers)- take samples of potato tissue using a cork borer and slice into discs.- place equal number of disks into test tubes bathing in water baths ranging from 20-80 degrees - place equal volume of pH buffer into each bath and allow it to equilibrate - add peroxide/buffer mixture to potato discs then fix stopper and side arm into the tube with the clip closed - time how long in seconds it takes for the bubble in the side arm to move 5cm
what is an anomalous result?
a result or reading that looks out of place with the others
Why should repeats always be done ?
in case of anomalous results and to add accuracy
