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Photosynthesis Experiments - Coggle Diagram
Photosynthesis Experiments
Chromatography
used to separate mixtures of
plant pigments in leaf
principle
some pigments more soluble than
others in the same solution
more soluble pigments = travel further.
method
mixture dissolved in
suitable solvent
pencil line drawn across
bottom of paper - ORIGIN
(important line NOT TOUCH solvent)
using fine capillary tube
place drop of solvent containing mixture
on the origin
allow to dry
put another drop of solvent
on top of the first
Repeat = produce conc spot
suspend paper in glass jar
allow bottom of paper to dip into solvent
(make sure chromatogram NOT suspended @ angle)
place lid on jar
allow air become saturated with the solvent
leave to one side
allow solvent travel up paper
once solvent reached top of paper
paper removed
position of
Solvent Front
noted w/ pencil
when measuring distance moved by spot
BE consistent
measure to same position of each spot
explanation
solvent rises up paper
carries separate molecules @ different speeds depending on solubility
more soluble solutes travel further in solvent.
Rf value
= calculated (always less than 1)
compares distance moved by substance + distance moved by solvent.
each compound = unique Rf value.
allows identify which spot represents which substance
ensures valid
results
solvent placed in sealed
chromatography container beforehand
(to allow vapour to saturate jar)
avoid touching chromatography
paper with bare hands
all marks on chromatography
paper should be in pencil
solutions under investigation not be placed too close together or too close to the edge of chromatography paper
solutions be concentrated by repeated application in the same position
chromatography paper should not touch the sides of the jar/container
Melvin Calvin's Experiment
Light Independent
Reaction
referred to as Lollipop
experiment
method
radioactive carbon dioxide added to suspension of autotrophic protoctistan Chlorella
after short time interval...
tap opened
chlorella released into
hot methanol
(killing it immediately preventing
any further reactions)
chlorella homogenised +
compounds
present identified by
chromatography
results
any compound formed that
contains radioactive carbon
is made from light independent
stage of photoS
glycerate phosphate first
compound formed
questions
why was all chlorella run
through FAST?
to ensure no product missed
no. of products increase up to
point + level off - why?
more time allows all products be labelled
finite no. of products produced
compounds are recycled
time chlorella exposed to radioactive carbon dioxide increased by only few seconds - why?
if time interval longer
consecutive sequence of individual molecules
not worked out
why flattened flask?
maximise light reaching chlorella
Engelmann's Experiment
Action Spectrum for
Photosynthesis
reveals which wavelengths work
best for photosynthesis
green alga Spirogyra
filamentous alga that has
chloroplasts arranged in spiral pattern
just inside cell wall
method
light passed
through prism
split into spectral components
before reaching Spirogyra
aerobic bacteria accumulate
near parts of alga illuminated with
red + blue light
proves which wavelengths best
for photosynthesis
why alga released
more oxygen
in
red and blue light
(rate of photoS rate best here)
Hill Reaction
method
Divide suspension
into 3 equal volumes
in 3 clean boiling tubes
Label tubes A - C
Boil tube C for 5 mins
in water bath
Place tube B into
dark cupboard
Leave tubes A + C under
bench lamp
Leave for 30 minutes
observe the
colour of DCPIP
questions
Why was plant material ground up in ice
cold isotonic buffer solution?
buffer
= prevents any fluctuation in pH that disrupts enzyme activity
ice cold
= low temps = reduces enzyme activity
isotonic
= prevents osmotic changes to organelles
Why did the tube A colourless
(in intense light)?
light
absorbed by chlorophyll molecules + electrons were
excited
and
released
electrons
reduced DCPIP
so solution changed from dark blue to
colourless
(green colour is due to the chloroplasts)
Why no colour change in tube B
(left in the dark)?
no
light
no
electrons
excited
from chlorophyll molecules
DCPIP
remains oxidised
Why did boiling contents of tube C show no colour change after illumination?
boiling chloroplasts
destroys thylakoid membranes
pigment molecules not release electrons
Mitochondria (M) also release electrons
How modify experiment to find if M also present in extract. Explain?
electrons released during respiration (in M)
respiration not affected by light
if M were present = electrons still released in the dark (tube B)
investigates LIGHT DEPENDENT reactions
take place in thylakoid membranes of chloroplasts
involves
isolating chloroplasts from living cells
suspending them in coloured electron acceptor
blue dye DCPIP
dye is blue = oxidised
colourless = reduced
monitor the loss of blue colour
indication that DCPIP accepted electrons
DCPIP takes the place of NADP
allows photolysis to continue
even when supply of NADP exhausted
why?
DCPIP continues to accept electrons from
electron transport chain
chlorophyll suspension
derived from plant material
ground up in ice–cold buffer solution
filtered
& centrifuged.
use mix of:
intact chloroplasts
thylakoid membranes (without surrounding stroma)
& outer membranes