Download Photosynthesis 1 student notes File

Slide 1
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© SSER Ltd.
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Slide 2
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The ultimate source of all energy for life on earth is the sun
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Courtesy of ARTTODAY
The energy that powers our muscles, allows birds to fly and drives
the many chemical reactions that are characteristic of living cells
originates from the SUN
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Slide 3
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The light energy from the sun must first be TRANSFORMED
into chemical energy before it can be used by living organisms
ENERGY
FROM
THE SUN
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The process that transforms light energy into chemical energy and uses
this to manufacture organic food is called PHOTOSYNTHESIS
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Slide 4
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The living organisms that inhabit our planet can be classed as either
AUTOTROPHS or HETEROTROPHS
AUTOTROPHS are organisms that are capable of synthesising their
own complex organic food molecules from simpler inorganic ones
The term AUTOTROPH means “self feeding” and organisms in this group
make use of an external, non-living supply of energy to drive their
“self-feeding” way of life
The vast majority of AUTOTROPHS harness the energy of sunlight to
manufacture their own food during the process of
PHOTOSYNTHESIS
ENERGY
FROM
THE SUN
THESE ARE THE PHOTOAUTOTROPHS
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Slide 5
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PHOTOAUTOTROPHS capture the sun’s energy and use it to convert
simple inorganic molecules, such as carbon dioxide and water, into complex,
energy-rich organic food
The various species of green plants
in this woodland are photoautotrophs
and manufacture their own food
by the process of photosynthesis:
6CO2 + 6H2O
Light Energy
C6H12O6 + 6O2
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Light
Carbon dioxide + Water
Glucose + Oxygen
Energy
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Slide 6
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There is a huge diversity of PHOTOAUTOTROPHS in our ecosystem
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Giant Redwoods of California
Diatoms – microscopic algae
The various species of photoautotrophs
range in size from the Giant Redwoods
of California to the microscopic algae
that are the major producers of food for
animals living in marine waters
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Slide 7
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Light is not the only source of energy utilised by AUTOTROPHS
Various species of bacteria have evolved mechanisms for manufacturing
their own food through utilising the energy contained in certain inorganic
molecules
THESE ARE THE CHEMOAUTOTROPHS
ENERGY
FROM
inorganic
molecules
CHEMOAUTOTROPHIC BACTERIA obtain the energy they need for food
manufacture by OXIDISING inorganic molecules such as
ammonia and hydrogen sulphide
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Slide 8
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CHEMOAUTOTROPHS IN THE ECOSYSTEM
Species of CHEMOAUTOTROPHIC BACTERIA known as the
NITRIFYING BACTERIA play an essential part in the NITROGEN CYCLE
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NITRIFYING BACTERIA oxidise ammonium and nitrite ions to nitrates
NH4+
NO2
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NO3
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energy
energy
The energy released
from these oxidation
reactions is used by the
bacteria to manufacture
their own food
The nitrates are absorbed
by green plants and the
nitrogen is incorporated into
nitrogen-containing
organic compounds
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Slide 9
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CHEMOAUTOTROPHS IN THE ECOSYSTEM
Certain CHEMOAUTOTROPHIC BACTERIA obtain the energy
needed to manufacture their own food through the oxidation of
the inorganic compound hydrogen sulphide
In habitats devoid of light such as caves and deep ocean beds, these bacteria
are the PRIMARY PRODUCERS supplying the energy that supports an
entire community of organisms
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The deep
ocean bed
devoid of
light
The dark recesses of a cave where chemosynthetic
bacteria supply the energy to support a community
of organisms
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Slide 10
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The majority of living species are unable to carry out either photosynthesis or
chemosynthesis
THESE ARE THE HETEROTROPHS
Energy from
already
manufactured
organic
materials
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HETEROTROPHS must consume already manufactured organic materials
as a source of energy for their life activities
HETEROTROPHS are ultimately dependent upon the AUTOTROPHS for their
supply of organic food
ENERGY is transferred from autotrophs to heterotrophs through
the food chain
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Slide 11
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This caterpillar is
obtaining organic
food material from
the products of
photosynthesis locked
up in this
photosynthesising
plant
Caterpillars are a
source of food for
small birds, which are
in turn eaten by
larger birds
Courtesy of ARTTODAY
The energy locked up in the organic material of this
autotrophic producer is passed along the food chain
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Slide 12
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AN OCEAN BED FOOD CHAIN DISCOVERED IN THE
PACIFIC OCEAN
H2S (Hydrogen Sulphide from the earth’s core)
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Chemosynthetic bacteria (PRIMARY PRODUCERS)
barnacles
clams
anemones
mussels
small worms
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Eaten by large crabs and fish
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Slide 13
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The overall process of photosynthesis in eukaryotes can be expressed by
the following equation:
6CO2 + 6H2O
Light Energy
Light
Carbon dioxide + Water
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C6H12O6 + 6O2
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Glucose + Oxygen
Energy
The equation shows that, when provided with light energy, photosynthesisers
utilise six molecules of carbon dioxide and six molecules of water in order
to manufacture one molecule of the carbohydrate glucose
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The equation gives no indication as to whether the released oxygen is obtained
from the water or the carbon dioxide
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The equation does, however, show that carbon dioxide is reduced during
photosynthesis and that this gas is the carbon source for the formation
of organic food (glucose)
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Slide 14
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The Source of Released Oxygen
In 1941, Ruben and
Kamen performed an
experiment that made
use of the heavy isotope
of oxygen – 18O
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OXYGEN GAS
One group of
plants was
supplied with
normal water
and labelled
carbon dioxide
C18O2
Another group of
plants was
supplied with
labelled water
H218O and normal
carbon dioxide
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The plants were allowed to photosynthesise for approximately 24 hours after
which the accumulated oxygen gas was analysed
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Slide 15
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The Source of Released Oxygen
As 18O is heavier than
normal oxygen 16O, then
the presence of the heavy
isotope could be detected
by analysing the
accumulated oxygen using
a mass spectrometer
One group of
plants was
supplied with
normal water
and labelled
C18O2
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OXYGEN GAS
Another group of
plants was
supplied with
labelled water
H218O and normal
carbon dioxide
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Regular oxygen 16O2
was detected from this group
of plants
Heavy oxygen 18O2
was detected from this group
of plants
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Slide 16
Ruben and Kamen concluded that water was being split during photosynthesis
and this molecule rather than the carbon dioxide was the source of the
evolved oxygen
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OXYGEN GAS
One group of
plants was
supplied with
normal water
and labelled
C18O2
Another group of
plants was
supplied with
labelled water
H218O and normal
carbon dioxide
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Regular oxygen 16O2
was detected from this group
of plants
Heavy oxygen 18O2
was detected from this group
of plants
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Slide 17
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Other experiments performed by various plant physiologists provided further
insight into the photosynthetic process
In 1905, Blackman provided vital evidence for the fact that photosynthesis is
A TWO STAGE PROCESS
Blackman worked on the interrelationships between the effects
of light intensity, temperature and carbon dioxide concentrations
on the rate of photosynthesis
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TEMPERATURE
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LIGHT INTENSITY
PHOTOSYNTHESIS
CARBON DIOXIDE
CONCENTRATION
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Slide 18
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Blackman used his knowledge of the concept of LIMITING FACTORS to explain
the results of his experiments
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THE LAW OF LIMITING FACTORS
When a process is affected by a number of factors, then the factor
that is in shortest supply at a given time determines the overall
rate of the process
Photosynthesis is affected by several factors that include temperature,
light intensity and carbon dioxide concentration
The rate at which photosynthesis proceeds is limited by whichever
of these factors is in short supply at a given time
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Slide 19
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Working with the aquatic
plant Elodea, Blackman reasoned
that if light energy drives
photosynthesis, then the
process should increase
in rate in more intense light
He measured the
rate of photosynthesis
in Elodea when subjected to
different light intensities
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Slide 20
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BLACKMAN’S RESULTS
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Rate of photosynthesis
Light is the limiting factor
in this region of the graph
Increasing the light intensity
to higher levels has no effect
on the overall rate of photosynthesis
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Increasing light intensity
Increasing the light intensity increases the rate of a photosynthesis but, at higher
intensities, increasing the light intensity has no effect on the rate
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Slide 21
Rate of photosynthesis
Blackman continued his experiments with limiting factors
High light
intensity
Low light
intensity
One experiment investigated
the effects of temperature on the
rate of photosynthesis under conditions
of (a) high light intensity and (b) low
light intensity
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Rate of photosynthesis at 25oC
Temperature (oC)
High light
intensity
Low light
intensity
Another experiment investigated
the effects of carbon dioxide
concentration on the rate of
photosynthesis under conditions of
(a) high light intensity and (b) low
light intensity – temperature kept
constant
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carbon dioxide concentration
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Slide 22
Rate of photosynthesis
Blackman continued his experiments with limiting factors
High light
intensity
Low light
intensity
Temperature (oC)
Rate of photosynthesis at 25oC
High light
intensity
Low light
intensity
carbon dioxide concentration
At low light intensities, the rate of
photosynthesis is unaffected by
the temperature
At high light intensities, the rate of
photosynthesis is temperature –
dependent
Thus when photosynthesis is limited
by light, it is temperature insensitive,
indicating that the overall process
is limited in rate by a reaction that is
probably NOT enzyme-controlled
When saturated by light, the rate is
determined by a stage that is
temperature- sensitive and, as such,
is likely to be an enzyme –
controlled chemical stage
At high light intensities the slowest
reaction is temperature –dependent
and does not involve the
absorption of light
Carbon dioxide is a reactant when
light intensities are high
Carbon dioxide is not involved when
light intensity is low and when
light absorption is the ratelimiting stage
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Slide 23
Rate of photosynthesis
CONCLUSIONS
High light
intensity
Low light
intensity
Rate of photosynthesis at 25oC
Temperature (oC)
High light
intensity
Low light
intensity
Blackman concluded that
photosynthesis is a
‘two-stage’ process
One stage is a light – requiring
non-chemical sequence of
reactions – this has come to be
known as the LIGHT DEPENDENT STAGE
The other stage is a chemical,
enzyme-controlled sequence
that utilises carbon dioxide –
this has come to be known as
the LIGHT – INDEPENDENT
STAGE
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carbon dioxide concentration
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Slide 24
FURTHER EVIDENCE
Further evidence has shown that the LIGHT DEPENDENT and LIGHT
INDEPENDENT reactions operate as separate stages
Experiments have shown that the amount of carbohydrate formed during
photosynthesis is GREATER IN FLASHING LIGHT than in
CONTINUOUS LIGHT
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CARBOHYDRATE
PER
PHOTON OF LIGHT
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Slide 25
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FURTHER EVIDENCE
It is envisaged that the LIGHT-DEPENDENT STAGE generates products
that are used in the LIGHT-INDEPENDENT STAGE
LIGHT-DEPENDENT STAGE yields products A and B
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A and B needed for the LIGHT-INDEPENDENT STAGE
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A and B accumulate in continuous light
In continuous light, the light-independent reactions cannot utilise the products of
the light-dependent stage as fast as they are produced
In flashing light, the short interval without light enables the light-independent
reactions to use the products of the light-dependent stage before more are produced
Thus we have a stage dependent upon light that is fairly rapid and a stage,
independent of light, that is slower and utilises the products of the faster process
PHOTOSYNTHETIC EFFICIENCY IS GREATER IN FLASHING LIGHT
PHOTOSYNTHESIS IS A TWO STEP PROCESS
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Slide 26
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Chloroplasts are the organelles within plant cells that contain all the
necessary structural and biochemical machinery for the process of photosynthesis
In 1882, a German biologist, Theodor Englemann provided strong evidence for
associating photosynthesis with the chloroplasts
Englemann worked with a species of freshwater alga, called Spirogyra
Spirogyra is made up of chains of regular-shaped cells in which the chloroplasts
are loosely coiled revealing areas of colourless cytoplasm
long spirally coiled
chloroplast
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colourless cytoplasm
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chain of cells
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Slide 27
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colourless cytoplasm
long spirally coiled
chloroplast
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Photomicrograph of Spirogyra
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Slide 28
Englemann placed his Spirogyra filaments on a microscope slide, together with
an even distribution of oxygen-sensitive bacteria
The microscope slide was enclosed in a chamber devoid of air, and red light
was directed onto a chloroplast and onto a portion of the colourless cytoplasm
After 30 minutes the slide was examined to determine the distribution of the
oxygen-sensitive bacteria
Englemann concluded that the bacteria had congregated at the site of
oxygen production and that this was the site of photosynthesis – the chloroplast
long spirally coiled
chloroplast
motile, oxygen-sensitive
bacteria
colourless cytoplasm
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chain of cells
Englemann’s findings support the view that photosynthesis takes place in the
chloroplasts
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Slide 29
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The chloroplast, the site of photosynthesis, is surrounded by an envelope of
two membranes and contains a jelly-like matrix called the stroma
Envelope
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Stroma
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Slide 30
Located within the stroma is an elaborate network of flattened membrane-bound
discs called thylakoids, in which light-capturing pigments, such as chlorophyll,
are contained
Envelope
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Thylakoids
Stroma
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Circular
DNA
molecule
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Lipid
droplets
Ribosomes
Many of the thylakoids
are stacked to form
The stroma also contains
A single granum
grana
a circular DNA molecule,
numerous ribosomes and lipid droplets
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Slide 31
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The photograph shown below details chloroplast structure
as viewed with a transmission electron microscope
Chloroplast envelope visible as two membranes
Stroma containing numerous
small ribosomes
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Lipid
droplets
Lamellae connecting
different grana
Courtesy of Dr. Julian
Thorpe – EM & FACS Lab,
Biological Sciences
University Of Sussex
A single
granum
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Slide 32
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The LIGHT-DEPENDENT REACTIONS take place within the thylakoid
membranes of the grana
thylakoid membranes
of granum
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The light dependent reactions begin when the energy in sunlight is captured
by the light absorbing pigments located within the grana
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Slide 33
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LIGHT AND PIGMENTS
Light energy travels in packets called photons
The energy content of a photon depends upon the
wavelength of the light
The shorter the wavelength, the higher the photon energy
The energy associated with a photon of light is called
a quantum of energy
Blue light has a relatively short wavelength whilst red light has
a relatively long wavelength
Plants are able to absorb a wide range of wavelengths because they contain
a variety of pigments with different structures and absorption properties
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Slide 34
THE ELECTROMAGNETIC SPECTRUM
Visible light represents only a small fraction of the full spectrum of electromagnetic
radiation produced by the sun
Plant pigments absorb light energy with wavelengths that fall within this
visible spectrum
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Visible light
Gamma rays
X- Rays
UV
Infrared
Radar
Short
wave
Radio waves
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400
500
600
700
750
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Wavelength (nm)
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Slide 35
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Pigments can be extracted from finely chopped leaves by grinding the tissues
in acetone
When the pigment extract is subjected to ascending paper chromatography, the
pigments can be analysed and identified
solvent
front
carotene
phaeophytin
xanthophyll
chlorophyll a
The principal plant pigments
found in green plants are
the chlorophylls (a and b) and the
carotenoids (carotene and
xanthophyll)
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chlorophyll b
origin
The role of these pigments is
to capture light energy and
each of the pigments absorb
various wavelengths to different
extents
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Slide 36
ABSORPTION SPECTRA
When a beam of light is passed through a glass prism, the visible spectrum of white
light is obtained
ght
te li
whi
When a beam of light is first passed through a solution of leaf extract, then an
absorption spectrum is obtained
ght
te li
whi
Black bands appear in the spectrum where particular wavelengths of light
have been absorbed
Predominantly, wavelengths at the blue and red ends of the spectrum have
been absorbed
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Slide 37
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ABSORPTION SPECTRA
The spectrum obtained by passing white light through a leaf pigment
extract shows us the wavelengths that the pigments absorbed
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More detailed information about the absorption process can be
obtained using a spectrometer
The degree of absorption at each wavelength can be measured for
both the total plant extract and the individual pigments
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This information can be used to generate an
ABSORPTION SPECTRUM GRAPH
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In a similar way, the effectiveness of different wavelengths of
light in bringing about photosynthesis can also be determined
The rate of photosynthesis at different wavelengths can be experimentally
measured
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This information can be used to generate an
ACTION SPECTRUM GRAPH
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ABSORPTION AND ACTION SPECTRA GRAPHS
100
action
spectrum
80
80
60
60
40
40
absorption
spectrum
20
0
400
20
rate of photosynthesis
100
% absorption
Slide 38
0
500
600
wavelength / nm
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700
Although all wavelengths between 400 and 700 nm are absorbed by the plant extract,
maximum absorption occurs in the violet/blue and red regions of the spectrum
There is a close correlation between the action spectrum and the absorption spectrum
The rate of photosynthesis is largely determined by the amount of light of different
wavelengths being absorbed
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Slide 39
CHLOROPLASTS AND LIGHT ABSORPTION
Engelmann performed an elegant experiment in which he set out to determine
whether any particular wavelengths of light were preferentially absorbed
by chloroplasts
Englemann placed a chain of algal cells called Cladophora onto a microscope slide,
together with an even distribution of oxygen-sensitive bacteria
The microscope slide was enclosed in a chamber devoid of air and a
microspectrum of light was directed at the cells
After 30 minutes the slide was examined to determine the distribution of the
oxygen-sensitive bacteria
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400
500
600
700 nm
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Slide 40
CHLOROPLASTS AND LIGHT ABSORPTION
Englemann concluded that the distribution of bacteria coincided with
areas where oxygen production by the photosynthetic algal cells was greatest
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He proposed that the wavelengths of light mainly responsible for oxygen production
in the Cladophora cells were 650 nm and 450 nm (the red and blue wavelengths)
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The attraction of most of the aerobic bacteria to sites illuminated with the red
and blue wavelengths of light, led Englemann to conclude that these are
the wavelengths preferentially used by chloroplasts during photosynthesis
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500
400
600
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700 nm
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Slide 41
ORGANISATION OF THE LIGHT-HARVESTING PIGMENTS
LIGHT ENERGY
The photosynthetic pigments are
embedded in the thylakoid
membranes of the grana
in clusters called
PHOTOSYSTEMS
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Each photosystem
consists of a
specialised chlorophyll
molecule called the
REACTION CENTRE,
surrounded by a few
hundred pigment molecules
that make up an antenna
complex
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carotenoid
chlorophyll a
chlorophyll b
ANTENNA
COMPLEX
The antenna pigments gather
light energy of various wavelengths
and funnel this energy to the
chlorophyll molecule at the reaction
centre
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In this way, the chlorophyll molecule
at the reaction centre receives all the
absorbed energy
REACTION CENTRE
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Slide 42
THE MACHINERY OF THE LIGHT-DEPENDENT REACTIONS
The thylakoid membranes of the grana contain two types of photosystem:
Photosystem 1 and Photosystem 2
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The two photosystems differ by virtue of the wavelength of light that
the reaction centre chlorophyll molecule absorbs
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The two photosystems are represented below on a kind of graph that
indicates the redox potential or energy content of the electrons
involved in the system
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Reaction
Centre
Reaction
Centre
Energy Level
of
Electrons
REDOX POTENTIAL
PS2
Light harvesting
pigments
PS1
Light harvesting
pigments
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Slide 43
THE MACHINERY OF THE LIGHT-DEPENDENT REACTIONS
Also located within the thylakoid membranes of the grana are:
• Prmary electron acceptor molecules • The electron acceptor, ferredoxin
• Carrier molecules that form an ELECTRON TRANSPORT CHAIN
Primary
Acceptor
Primary
Acceptor
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ferredoxin
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electr
on tr
a
chain nsport
Reaction
Centre
Light harvesting
pigments
PS1
Reaction
Centre
All of these components are required
for the LIGHT-DEPENDENT
STAGE and are ordered on the graph
in relation to their redox potential
REDOX POTENTIAL
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Light harvesting
pigments
PS2
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Slide 44
When light strikes the grana of the chloroplasts, the antennae pigment molecules
absorb light energy and become excited – this is PHOTOEXCITATION
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Primary
Acceptor
Primary
Acceptor
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2e_
The released electrons
are accepted by
Primary Acceptors located
within the thylakoid
membranes
2e_
PS1
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PS2
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2 photons
Energy is funnelled to each of the reaction centres
where the specialised chlorophyll molecule receives
sufficient energy to release an electron –
PHOTOIONISATION
A single electron is released for each photon absorbed
2 photons
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Slide 45
The release of electrons from the photosystems creates ‘electron holes’ at the
reaction centres that must be filled if PHOTOIONISATION is to occur again
Primary
Acceptor
Primary
Acceptor
½ O2 + 2H +
photolysis
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2e-
The ‘electron
holes’ of both
photosytems
have now been
filled
H2O
2e_
The ‘electron hole’ at
photosystem 1 (PS1) is
filled as electrons pass
along a chain of carriers
from the primary acceptor
for photosystem 2
2e2e_
+ 2e -
PS2
The ‘splitting of
water’ to form
hydrogen ions and
oxygen gas is
called
PHOTOLYSIS
2 photons
ADP
+Pi
The flow of electrons
along this chain of
carriers releases
sufficient energy for
the formation of ATP
ATP
PS1
The formation of ATP, as
the photoexcited electrons
pass along this chain,
is called
photophosphorylation
The ‘electron hole’ at
photosystem 2 (PS2) is
filled as water molecules
donate electrons to the
chlorophyll molecule
at the reaction centre
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2 photons
The donation of electrons to PS2 results in the
‘splitting of water molecules’ releasing hydrogen
ions and oxygen gas
Oxygen gas is released as a waste product of
photosynthesis
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Slide 46
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2H+
Primary
Acceptor
2e _
Primary
Acceptor
ferredoxin
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2e _
2e_
NADP
ADP
+Pi
2e_
H2O
½ O2 + 2H +
photolysis
+ 2e -
PS1
NADPH2
The electrons released
from photosystem 1
to the primary acceptor
are passed via
ferredoxin
to a molecule
2 photons of the hydrogen
carrier
NADP
PS2
ATP and NADPH2
are needed to
drive the
light-independent
stage of
photosynthesis
+ 2e+ 2H+
ATP
The light-dependent
stage of photosynthesis
has generated ATP
and reducing power
in the form of NADPH2
2 photons
These electrons, together with the hydrogen ions
formed during the photolysis of water, reduce
NADP to NADPH2
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Slide 47
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Summary of the Light-Dependent Stage – The Z-Scheme
2H+
Primary
Acceptor
2e _
2e_
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2e _
Primary
Acceptor
ferredoxin
NADP
2e2e2e
H2O
ADP
+Pi
_
+ 2e+ 2H+
ATP
Photophosphorylation
½ O2 + 2H +
photolysis
PS1
+ 2e -
NADPH2
PS2
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2 photons
REDOX POTENTIAL
2 photons
The purpose of the light-independent stage is
to generate ATP by photophosphorylation
and NADPH2 for use in the
light-independent stage
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Slide 48
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THE CALVIN CYCLE
Calvin and his co-workers determined the chemical steps involved in
the reduction of carbon dioxide to carbohydrate during the
LIGHT-INDEPENDENT STAGE of photosynthesis
Calvin was awarded the Nobel Prize for his work in 1961
The experimental procedure used by Calvin and his co-workers involved
the use of the radioactive form of carbon – 14C
By providing cultures of a single-celled algae with radioactive carbon dioxide
(14CO2), Calvin was able to follow the fate of radioactively
labelled carbon compounds
14CO
2
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Slide 49
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CALVIN’S EXPERIMENT
Calvin grew cultures of the green alga
Chlorella in his famous ‘lollipop’
apparatus
The lollipop-shaped flask was
designed to ensure that there
was even illumination to
the culture of cells
14CO
LIGHT
ENERGY
2
The illuminated culture was
exposed to radioactive carbon
dioxide (14CO2) for various
intervals of time
suspension of the single-celled
alga, Chlorella
At specific intervals of time
after exposure to 14CO2, samples
of the algae were rapidly
killed by opening the sampling
valve to release a sample into
boiling alcohol
lollipop-shaped
flask
sampling valve
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The killed algal cells were then
homogenised and their contents
extracted
The chemical content of the
extract was then analysed using
TWO-DIMENSIONAL
CHROMATOGRAPHY
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boiling alcohol
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Slide 50
CALVIN’S EXPERIMENT
TWO-DIMENSIONAL PAPER CHROMATOGRAPHY
A drop of the concentrated algal extract was placed at one edge of a square of
chromatography paper
The chromatography paper was dipped in the first solvent to separate the chemical
compounds in the spot
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dry and
turn
through
90o
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concentrated
Develop with second
Develop with first extract
solvent of butanol:proprionic acid
solvent of phenol:water
FINAL
CHROMATOGRAM
The chromatogram was then dried and turned through 90o and run again using a
different solvent
The identity of each of the separated compounds was then made and the
determination of which of these compounds was radioactive was achieved
through the technique of AUTORADIOGRAPHY
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Slide 51
CALVIN’S EXPERIMENT
X-ray sensitive
film
AUTORADIOGRAPHY
In order to determine which of the chemical compounds
on the final chromatogram were radioactive, Calvin placed
a sheet of X-ray sensitive film in contact with the chromatogram
The chromatogram and X-ray sensitive film were left in
contact in the dark for about two weeks
During this period, any radioactive carbon compounds (14C)
blacken the X-ray sensitive film and become visible as dark spots
The resulting AUTORADIOGRAMS enabled Calvin to determine
the order in which compounds were formed during the lightindependent stage
malic acid
GP
triose phosphate
diphosphates
after 5 seconds
of photosynthesis
after 15 seconds
of photosynthesis
EXAMPLES OF AUTORADIOGRAMS FROM CALVIN'S EXPERIMENT
These autoradiograms show
that, after a short time
period, the spot most heavily
labelled is GP (glycerate 3 –
phosphate)
Calvin determined that the
first stable compound into
which carbon dioxide had
become incorporated was
glycerate 3-phosphate (GP)
- a three carbon compound
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Slide 52
THE CALVIN CYCLE – LIGHT INDEPENDENT STAGE
The first stage of the
Calvin Cycle is
Carbon Dioxide
Fixation
The Light-Independent Stage
of photosynthesis takes place in
the stroma of the chloroplast
CO2
GP – glycerate 3-phosphate
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RuBP – ribulose bisphosphate
Located within the stroma of the chloroplast is the five-carbon, phosphorylated
sugar known as RIBULOSE BISPHOSPHATE (RuBP)
The enzyme ribulose bisphosphate carboxylase catalyses the reaction between
carbon dioxide and RuBP – CARBON DIOXIDE FIXATION
This reaction forms an UNSTABLE SIX CARBON COMPOUND that immediately
splits into TWO MOLECULES of the compound glycerate 3-phosphate (GP)
Glycerate 3-phosphate (GP) is the first STABLE compound to be formed during
the Calvin Cycle
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Slide 53
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CO2
2 ATP
GP – glycerate 3-phosphate
2 ADP + Pi
2 NADPH2
RuBP – ribulose bisphosphate
2 NADP
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Triose Phosphate
Each molecule of GP is now reduced to the phosphorylated, three-carbon sugar
TRIOSE PHOSPHATE
This reduction step requires the hydrogens from NADPH2 and is also an endergonic
(energy demanding) reaction requiring the utilisation of ATP molecules
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The conversion of GP to triose phosphate is the step that utilises the products
of the LIGHT-DEPENDENT STAGE – ATP and NADPH2
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Slide 54
As triose phosphate molecules accumulate, some are used to form glucose which
can be converted into starch for storage or used to manufacture other food
molecules such as amino acids and fatty acids
Some of the triose
phosphate molecules
are used to regenerate
the compound RuBP
to enable the cycle
to continue operating
CO2
2 ATP
GP – glycerate 3-phosphate
2 ADP + Pi
2 NADPH2
RuBP – ribulose bisphosphate
ADP
+ Pi
ATP
This stage
utilises ATP for
the phosphorylation
of RuP
2 NADP
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Triose Phosphate
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RuP – ribulose phosphate
Glucose
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STARCH
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Slide 55
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SUMMARY
CARBON DIOXIDE
FIXATION
STAGE
REDUCTION STAGE
CO2
2 ATP
GP – glycerate 3-phosphate
2 ADP + Pi
2 NADPH2
RuBP – ribulose bisphosphate
ADP
+ Pi
2 NADP
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Triose Phosphate
ATP
REGENERATION
STAGE
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FOOD
SYNTHESIS
STAGE
RuP – ribulose phosphate
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Glucose
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STARCH
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Slide 56
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BALANCING THE BOOKS
The overall process of photosynthesis in eukaryotes is expressed by
the following equation:
6CO2 + 6H2O
Light Energy
C6H12O6 + 6O2
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The equation shows that SIX MOLECULES of CARBON DIOXIDE are
required for the synthesis of ONE MOLECULE OF GLUCOSE
The reactions in which these six molecules of carbon dioxide are
utilised can be shown by following the Calvin Cycle
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Slide 57
Six molecules of carbon dioxide combine with six molecules of RuBP to form six
molecules of an unstable 6-carbon compound that immediately splits into
TWELVE molecules of the 3-carbon compound GP
CO 2FIXATION
STAGE
6CO 2(1C)
12 GP (3C)
Glyceraldehyde
phosphate
12 ATP
REDUCTION
STAGE
12 ADP + 12 P i
6RuBP (5C)
ribulose bisphosphate
12 ATP and 12 NADPH2 from
the light-dependent stage
are required to drive this step
The remaining 10 molecules of
triose phosphate are utilised
to regenerate 6 molecules of RuBP
6ADP
+ 6Pi
6ATP
Each RuP
molecule is
phosphorylated
to RuBP at the
expense of ATP
6RuP (5C)
ribulose phosphate
REGENERATION
STAGE
12 NADPH2
12 NADP
The 12 GP
molecules are
reduced to
12 molecules
of triose
phosphate
12 TRIOSE PHOSPHATE (3C)
2 molecules
of triose
phosphate
are used to
form 1
molecule of
glucose
PRODUCT
SYNTHESIS
STAGE
GLUCOSE
STARCH
(glucose polymer)
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Slide 58
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12 GP (3C)
Glyceraldehyde
phosphate
CO2 FIXATION
STAGE
6CO 2(1C)
12 ATP
REDUCTION
STAGE
12 ADP + 12 Pi
12 NADPH 2
6RuBP (5C)
ribulose bisphosphate
12 NADP
2 out of 12 or 1/6th of the triose
phosphate molecules are used to form
glucose (6C)
10 out of 12 or 5/6ths of the triose
phosphate molecules are used to
regenerate 6 molecules of RuBP
(30 carbon atoms in total)
6ADP
+ 6Pi
6ATP
12 TRIOSE PHOSPHATE (3C)
6RuP (5C)
ribulose phosphate
PRODUCT
SYNTHESIS
STAGE
GLUCOSE
REGENERATION
STAGE
STARCH
(glucose polymer)
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THE IMPORTANCE OF THE LIGHT-DEPENDENT STAGE FOR
THE OPERATION OF THE CALVIN CYCLE
Throughout the course of his experiments, Calvin measured the relative amounts
of GP and RuBP present first in continuous light and then when the light was
switched off
In the light, both GP and RuBP
remain at a steady level
In the light, RuBP is converted into
GP which, in turn is converted
back to RuBP during the
regeneration stage
GP
In darkness, GP accumulates and
levels of RuBP fall
This provides evidence for the
RuBP
fact that the Calvin Cycle is
disrupted in darkness
This effect is observed because
the reduction of GP to triose
phosphate requires ATP and
NADPH 2 from the
light
dark
light-dependent stage
The Calvin Cycle is dependent upon
The formation of triose phosphate
the light-dependent stage
is therefore prevented and regeneration
for its operation
of RuBP cannot take place –
GP thus accumulates and RuBP levels fall
relative amount of substances
Slide 59
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Slide 60
manufacture their own organic food
SUMMARY Photoautotrophs
during the process of photosynthesis
Photosynthesis is a two-stage process that generates ATP and NADPH2 during
the light-dependent stage and utilises these products to reduce carbon dioxide to
carbohydrate in the light-independent stage
water
LIGHT
ENERGY
CO2
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CHLOROPLAST
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NADP and ADP + Pi
LIGHT
INDEPENDENT
STAGE
LIGHT-DEPENDENT
STAGE
GRANUM
NADPH2 and ATP
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STROMA
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oxygen
sugar
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Slide 61
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Acknowledgements
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Photographs on slides 2, 6 and 9 – Courtesy of ArtToday ©
FOR VIEWING PURPOSES ONLY
Visit: www.arttoday.com
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Chloroplast Electron Micrograph – Courtesy of Julian Thorpe
EM and FACS lab
Biological Sciences
University of Sussex
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END SHOW
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