Practicals Flashcards
(40 cards)
how can valid results be obtained with a potometer?
- use a healthy, fresh plant shoot
- cut plant shoot and assemble potometer underwater (avoid air bubbles)
- dry leaves (control humidity/initial Ψ around leaves)
how to prepare a photosynthetic pigment extract? (and why)
- soak leaves in acetone (propanone) to act as a solvent
- add a pinch of sand (help grind)
- use a mortar and pestle to grind up leaves with acetone
stationary phase in TLC
silica gel layer of TLC plate
what solvent is used for photosynthetic pigment TLC?
- cyclohexane
- propanone (acetone)
- petroleum ether
why must a lid be used on TLC?
- solvents are volatile
- evaporate quickly
- will not diffuse up the TLC plate and carry extract
how and why must you be careful when handling TLC plates?
- avoid fingerprints
- avoid damaging fragile silica gel layer
why must the TLC plate not touch the sides of the beaker/tube?
- solvent may evaporate off the plate, condense and drip down the side of the container
- can damage the silica gel
- can skew the movement of solvent (making it hard to measure Rf values)
how can photosynthetic pigments be identified from TLC?
- measure Rf values and record colours
- compare to known data
what can be used to add sample spots to TLC and why?
capillary tube for small spots
how many spots should be applied in TLC?
- multiple (3/4) spots of EACH sample in the SAME spot (concentrated pigment)
- if using different samples, spot them in different places
what is a TLC plate placed in?
developing chamber (tall beaker with lid)
how can solvent evaporation be reduced in a developing chamber?
- add a lid
- place a layer of filter paper around inner walls of developing chamber, wet the filter paper with solvent to adhere to the sides of the chamber and saturate it (prevent solvent from TLC evaporating and condensing on the side of the chamber)
how does the solvent travel through TLC?
capillary action
why must the TLC plate be taken out before the solvent reaches the top?
- once the solvent reaches the top, Rf values may be invalid
- pigments may start to run into each other (ones at the bottom catch up with the ones at the top etc)
why must you mark the solvent front immediately with a pencil?
- avoid solvent evaporating (will no longer be able to see the solvent front)
- pencil to avoid interference with the pigments and damage to the silica gel
how are molecules separated in TLC?
- relative adsorption with the stationary phase
(separated by polarity depending on the solvent: polar molecule in polar solvent will spend less time adsorbed to the stationary phase)
potential limitations of a respirometer
- measurements may only be taken at big intervals
- apparatus may not be air tight
- may be difficulty reading meniscus leading to inconsistent, invalid results
- may not be a scale on the capillary tube
what should you make sure to do whenever varying or controlling temperatures?
allow things to equilibrate (e.g. test tubes of water to equilibrate with the surrounding water bath temperature)
before placing beetroot cubes/cores in varying solutions what should you do? and why?
- wash them in distilled water to allow any excess pigment that has already leaked out of damaged cells to wash off
- improves validity and ensures results are comparable as each beetroot piece started at the same point
- blot the beetroot cells dry to remove excess water and pigment
how and when do you calibrate a colorimeter?
- use a cuvette of distilled water and “zero” the colorimeter
- between every sample
what filter do you use for colorimetry? and what does the % absorbance mean? use an example
- opposite colour to the thing you are testing
- higher % absorbance means there is more of the coloured pigment
- e.g. for beetroot solutions, use a blue filter because red absorbs blue light. higher % absorbancy means more red pigment
how should you label a petri dish?
- initials
- date
- microorganisms growing
- write on the base of the petri dish
four pre-inoculation aseptic techniques in microbiology PAG
- sterilise surfaces
- wash hands with soap
- don’t touch any sterile equipment
- sterilise equipment with ethanol and flaming
how do you make an agar plate using molten agar and a culture broth?
- flame the neck of the molten agar bottle and the culture broth bottle
- use a sterile pipette to transfer a certain volume of the culture into the molten agar and replace the lid
- roll the bottle of bacteria and agar to mix the solution without frothing it
- flame the neck of the bottle again
- lift the petri dish lid at an angle
- pour the bacteria agar solution into the petri dish
- replace the lid
