Download Lab 5 - Photosynthesis Reading 09

Lab Five: Photosynthesis1
Introduction
Energy is the ability to do work. It comes in lots of forms: light, heat, radiation, movement, and chemical. Energy
itself cannot be created or destroyed, only transformed. Every time energy is transformed, changing from one type to
another, some of the original energy is changed to heat, which dissipates into the surroundings and is not usable. Because
of this, all living things must continually take in energy to survive. On Earth, photosynthetic organisms get their energy
from light. Using photosynthesis, the energy in light is transformed into potential chemical energy stored in the bonds of
glucose molecules. All living things depend on photosynthetic organisms to convert light energy into chemical energy.
Animals, many bacteria and protists, and fungi cannot use light directly. They use the chemical energy stored in their food,
all of which can be traced back to the photosynthetic organisms that first converted the light energy into chemical energy
(food). On land, the plants are the major photosynthetic organisms, while in the oceans it is the protists. There are also
some Archaea that can do photosynthesis, though it is of a type chemically different from that of plants and protists .
Part 1- Pigments
The first step of photosynthesis is capturing light energy using pigments. This step is called the light reaction, but is
more correctly called the light dependent reaction, because light is directly needed. There are several pigments that absorb
light, they include chlorophylls a & b, carotenoids, and xanthophylls. These pigments allow plants to absorb several
wavelengths of light.
Paper chromatography is a useful technique for separating and identifying pigments and other molecules from cell
extracts that contain a complex mixture of molecules. The solvent moves up the paper by capillary action, which occurs as
a result of the attraction of the solvent molecules to the paper and the attraction of the solvent molecules to one another. As
the solvent moves up the paper, it caries along any substances dissolved in it. The pigments are carried along at different
rates because they are not equally soluble in the solvent and because they are attracted, to different degrees, to the fibers in
the paper through the formation of intermolecular bonds, such as hydrogen bonds.
Beta carotene, the most abundant carotene in plants, is carried along near the solvent front because it is very soluble in
the solvent being used and because it forms no hydrogen bonds with the cellulose in the paper. Another pigment,
xanthophylls, differs from carotene in that it contains oxygen. Xanthophyll is found further from the solvent front because
it is less soluble in the solvent and has been slowed down by hydrogen bonding to the cellulose. Chlorophylls contain
oxygen and nitrogen and are bound more tightly to the paper than are other pigments.
Part 2- Photosynthesis
During the light reactions, when light energy is absorbed, the electrons within those pigments are boosted to higher
energy levels, and this energy is used to produce ATP and reduce NADP to NADPH. The energized electrons from the
pigments travel down a series of electron carriers known as an electron transport chain. As they are transferred, some of
their energy is used to pump hydrogen ions across the thylakoid membrane. This creates an electrochemical gradient.
There is more H+ on one side of the membrane. It is a chemical gradient, because of the molecule concentration difference,
and an electrical gradient, because of the excess positive charges on one side. Hydrogen ions are pumped at several points
along the gradient. The pigments and electron carriers together are known as photosystem II.
The electrochemical gradient made by the electrons moving down the electron transport chain stores a huge
amount of energy. That energy can be used to do the work of making ATP. A molecule called ATP synthase acts as a
channel, letting hydrogen ions diffuse down the gradient. The energy released as they move is used to phosphorylate ADP,
making it ADP. This is called photophosphorylation, because light energy (photo) is used to restore ATP
(phosphorylation).
When electrons reach the end of the electron carriers, the electron itself has two possible paths. In the first path,
called the non-cyclic photophosphorylation, the electron is added to photosystem I, another pigment and electron
transport chain. Here the electrons are excited again, and then added to NADP, making NADPH. This is a reduction
reaction (adding electrons). NADPH carries the last of the energy that was in the electron. The second path is known as
cyclic photophosphorylation. The electron is returned to the pigment molecule in photosystem I. There it can be
energized again by light, travel down the electron transport chains again, making more ATP. These two pathways exist
because of requirements for ATP and NADPH in the second part of photosynthesis; more ATP is needed than NADPH.
There is only one problem left for the light reactions. The original pigment has lost an electron and needs a new
one. The electron is taken from water making oxygen and hydrogen ion. Water is known as the electron donor; it donates
all the electrons that eventually wind up in the NADPH. NADPH is known as the final electron acceptor, because that is
where the electrons finish.
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Adapted from Lab 4 in AP Biology Lab Manual from the College Board, 2003.
In the second step of photosynthesis, the ATP and NADPH are then used to changing CO2 to organic molecules
like glucose, in a process called carbon fixation. This step is often called the dark reactions, because it can occur in the
dark. It is more correctly called the light independent reactions, because it occurs both during the day and night, but does
not require the light directly. The name for the reactions of this part is the Calvin Cycle.
We can study the light independent reactions using a dye-reduction technique. The dye-reduction experiment can
show us how much photosynthesis is occurring in chloroplasts. The dye is DPIP (2,6-dichlorophenol-indolphenol). It will
accept the electrons in place of NADP. When DPIP does not have the electron, it is blue. When it is reduced by an
electron, it turns colorless. We will measure this change in color using a spectrophotometer. If we try to pass blue light
through the DPIP before it is reduced, the light will not pass through (low transmittance). As it turns colorless, the
transmittance will increase.
Your set up will expose different combinations of solutions and chloroplasts to light. However, they should not be
exposed to heat. To accomplish this you will use a heat sink between the light bulb and the solutions. A heat sink is just a
clear flask or jar filled with water. The light will pass through but the heat will be absorbed by the water.