Good afternoon, Anup. For Monday, you are to fix your DCPIP lab. For your discussion,
bring a few folder pages:
●
- DCPIP has a higher affinity for electrons than NADP
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- state clearly that CHLOROPHYLL loses an electron during the lightdependent stage and explain how it happened for the tubes
●
- EXPLAIN HOW the coldness of the buffer solution would have slowed down
enzyme activity and why that was beneficial prior to the start of the reaction
●
- EXPLAIN why the buffer solution had sucrose in it (check Aliyah)
●
- EXPLAIN the role of the sand during crushing and grinding of the leaf
You will fix the discussion on the drawing page after page 9
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Explain WHY a buffer solution had to be used to maintain pH at all
Anup Jagat U62
Date: 21/09/2022
Title: Photosynthesis
Exp#
Aim: To investigate the light-dependent stage in photosynthesis in a leaf extract, using the
dye DCPIP.
Introduction:
This experiment investigated the light-dependent stage in photosynthesis in a leaf extract,
using the dye DCPIP.
Photosynthesis can be described as the process by which plants use energy captured from
sunlight to convert carbon dioxide and water into glucose. The glucose can then be broken
down into smaller molecules such as sucrose for transport throughout the plant. The sugars
are carried to plant cells that need them by tubes called phloem.
Photosynthesis takes place in the chloroplasts of a plant and can be summarised by the
following equation:
6CO 2 + 12H 2 O (+ light energy) C 6 H 12 O 6 + 6O 2 + 6H 2 O
Photosynthesis takes place in two stages, the first of which is called the light-dependent
stage and needs light energy and consists of the light-dependent reactions. This stage takes
place in the thylakoids within the chloroplast. Chloroplasts are the main site of
photosynthesis. Their membranes form fluid-filled sacs called thylakoids. Thylakoids form
stacks called grana. Chlorophyll molecules are embedded in the membranes of the
chloroplast and are found in photosystems 1 and 2(PSI and PSII).
Photosystems are scattered across the thylakoid membrane which contain photosynthetic
pigments that absorb light energy across different wavelengths of light.
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Energy from sunlight is transferred into the production of ATP and the reduction of NADP to
NADPH. Both of the products are used in light-independent reactions. Absorption of light
done by photosystems 1 and 2 (PSI and PSII) which are attached to the thylakoid
membranes. The primary pigment chlorophyll molecule acts as a reaction centre for the PS.
Other pigments called accessory pigments capture photons of light and transfer their energy
to the central molecule. Energy that breaches the reaction centre excites an electron in the
primary chlorophyll molecule to a higher level.
The chlorophyll is then oxidised and taken up by an electron acceptor. The process by which
the water molecule is split into oxygen and hydrogen and the electrons from water are
carried to the reaction centre is called photolysis.In the photosystems, photons of light are
transferred from one molecule to another by resonance transfer. It is being tested that
isolated chloroplasts in the leaf extract obtained could still carry out the light-dependent
stages of photosynthesis if provided with the correct conditions.
When DCPIP is reduced, it changes from blue to colourless. DCPIP in the experiment takes
the place of NADP in this reaction. When NADP is exhausted, DCPIP continues to accept
the electrons from the electron transport chain allowing photolysis to continue. The DCPIP
changes colour from blue to colourless when it is reduced thus showing if light dependent
reactions are occurring.
Apparatus and Materials:
Apparatus
Materials
3 capillary tubes
Aqueous solution of 1% DCPIP
Petri dish
Leaves
White tile
Sand
Aluminium foil
Buffer solution
1 syringe
3 droppers
Glass rod
Scalpel
Specimen tube
Pestle and mortar
2
Spatula
Bench lamp
Paper towel
Method:
3
Control of experiment: Tube 1
VARIABLES:
Controlled variables:
Manipulated variables:
Responding variables:
Results:
Table showing the colour changes observed in the leaf extract of tubes 1,2 and
3 over a 10 minute time span
Time/min
Tube 1
Tube 2
Tube 3
No colour change
No colour change
No colour change
No colour change
No colour change
No colour change
No colour change
No colour change
No colour change
No colour change
No colour change
Turned light green
No colour change
No colour change
Remained light
green
No colour change
No colour change
Remained light
green
No colour change
No colour change
Remained light
green
No colour change
No colour change
Remained light
green
No colour change
No colour change
Remained light
green
No colour change
No colour change
Remained light
green
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Discussion:
In PS1 chlorophyll loses an electron in the reaction centre when light energy is absorbed by
chlorophyll molecules at the reaction centre. The energy is passed from along molecules
until chlorophyll emits a high-energy electron. To replace the electron in the chlorophyll, a
molecule of water is split by an enzyme. This splitting is called photolysis and it releases an
electron and results in the formation of oxygen (O2) and hydrogen ions (H+) in the thylakoid
space. The electron released goes into the chain of molecules that absorb light energy in the
chloroplast. DCPIP has a higher affinity for electrons than NADP. Therefore, the electrons
emitted in a photosynthesising PS1 in the light dependent stage would react with DCPIP.
Tube 2 and 3 showed different colour changes as seen in the table due to the different
conditions they were placed under in the experiment. Tube 2 had no colour change due to
the shielding of light experienced in the experiment. With no light source to provide the
photosystems in the leaf extract with photons, the accessory pigments or antenna system
had no photons to transfer to the primary pigments or reaction centre. Since no energy is
passed to the reaction centre,photolysis does not occur, it does not lose an electron and so
NADP is not reduced. Therefore, the DCPIP present never reacts and so there is no colour
change observed.
In contrast, Tube 3 has a light source directly above the leaf extract, thus providing lots of
energy for resonance transfer. Since DCPIP has a higher affinity for electrons than NADP,
all the DCPIP molecules react . This tube proves that light is a necessary condition for
photosynthesis to take place and that increased light speeds up the light-dependent
reactions until NADP is exhausted.
In tube 1 no change is observed as it is used as there are no conditions placed upon it. This
tube was not shielded from light so it can be assumed that, given enough time, the extract in
tube 1 would also decolourize. But not enough time for this to be observed was allotted for
the experiment. This tube was used as the control of the experiment. It serves this role well
because there is no condition placed on it and it shows what would happen to the leaf
extract in the tube if it was not placed under foil wrap or directly under a lamp. It shows what
conditions are necessary for transduction of light into chemical energy and which are not.
The purpose of the pestle and mortar was to decrease the particle size of the components of
the experiment by crushing and grinding. Adding sand to the mixture in the mortar helps
reduce the particle size by providing a rougher surface for the pestle to grind away at. It also
helps to extract substances from the plant material by breaking the plant cells.
Buffer Solutions resist change in pH since they contain high concentrations of both a weak
acid and its conjugate base. These components can neutralise added H+ and OH- ions and
as a result resist change in ph. A change in Ph can slow the rate of enzyme activity. This is
due to the fact that enzymes have an optimum temperature range and any pH outside this
range will affect charges in the amino acids that form the structure of the enzyme, ultimately
slowing the enzyme activity. The enzyme that splits water to release an electron into the
chlorophyll molecule chain and oxygen is necessary for the continuous process of
photosynthesis in the light dependent stage. Any negative effects on the enzyme activity will
therefore affect the rate of photosynthesis and therefore the results of the experiment.
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Using ice cold buffer solution and ice cold sucrose solution is necessary to slow enzyme
action and prevent damage to the chloroplasts while the extract is being crushed and ground
in the mortar. The coldness of the solution slows the speed at which molecules move and
hence the substrate collides with the enzyme less often. It also maintains osmotic potential
to prevent chloroplasts from swelling and bursting in the water. This occurs since sucrose
dissolved in water forms weak bonds with the water molecules, making it more difficult for
water to move out of the leaf extract by osmosis. The buffer solution is therefore beneficial to
the experiment by maximising the efficiency of the substances present in the leaf extract.
This allows for more accurate results as there is minimised interference.
Precautions:
Sources of error:
Limitations:
Improvements:
Conclusion:
It was determined from the experiment that light is necessary for photosynthesis to occur
since the DCPIP in tubes 1 and 2 did not decolourise while the solution in tube 1 was
decolourised in the presence of light.
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Lab 2
*Leaf cross section
Mag x40 and Magx10
*3 Palisade cells
Mag x 40
Title: Photosynthesis and Transportation in Plants - Leaf Structures
AIM:
1. To draw, label and annotate a transverse section through a dicot leaf under low
power (x10 obj lens)
2. To observe the different tissues in a dicot leaf under high power(x40 obj lens)
3. To observe 3 palisade cells under high power(x40 obj lens)
Lab 3
*Plan of Leaf vein
Mag x 40
*Plan of root
Mag x 40.
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