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Home > Biology > Options > Biochemistry > Biochemistry: 5. Using isotopes to prove that water is split in photosynthesis

Option 9.9 Biochemistry: 5. Using radioisotopes to prove that water is split in photosynthesis

Syllabus reference (October 2002 version)
5. The use of isotopes made the tracing of biochemical reactions much easier	Students learn to: explain why the tracking of biochemical reactions is difficult identify that isotopes of some elements may be unstable and emit radiation define what is meant by the half-life of an isotope and explain how this would affect its use in biochemistry outline the evidence provided by: Hill and Scarisbruck Ruben to confirm that the oxygen released by photosynthesis originated from water	Students: gather and process information from secondary sources to outline the range of isotopes that have been useful in studying photosynthesis and explain how radioactive tracers can be incorporated into plants to follow a biochemical pathway, such as photosynthesis

Extract from Biology Stage 6 Syllabus (Amended October 2002) © Board of Studies, NSW.
[Edit: 18 June 09]

Prior learning: Stage 4-5 Syllabus, 4.8.1(c) Structures and Systems 4.8.2 (c), 4.8.4(d) Interactions 4.10 (c); Models,Theories and Laws 5.6.5 (a) and(b), 5.7.1.

Preliminary module 8.2 (subsection 2) module 8.3 (subsection 4).

explain why the tracking of biochemical reactions is difficult

• The tracking of biochemical pathways is difficult because the substances taking part cannot be seen, even with an electron microscope. There are also many intermediate steps in reactions. The cell has therefore appeared like a “black box” with substances entering and leaving, having been transformed or changed. Cells do not function when dead and so methods of study have been limited to those that allow living cells to be studied.

gather and process information from secondary sources to outline the range of isotopes that have been useful in studying photosynthesis and explain how radioactive tracers can be incorporated into plants to follow a biochemical pathway, such as photosynthesis

• Gather information from various sources including Biochemistry textbooks. Practise efficient data collection techniques and extract the information to complete a table like the table shown below.

The range of isotopes useful in studying photosynthesis are:

Isotope	Radiation type	Half - life	Used for the movement of
Hydrogen as 3H	β	12.1 years	H+ across the thylakoid membrane
Carbon as 14C	β	5700 years	C from CO2(g)to glucose
Phosphorous as 32P	β	14.3 days	Pi and ADP to form ATP
Oxygen as 18O	None - measured by a mass spectrophotometer		O from H2O compared to CO2

• Process the information by looking for trends and patterns in the information. Use the information to write an outline of the range of isotopes that can be used to study photosynthesis and explain how isotopes can be incorporated into plants.

identify that isotopes of some elements may be unstable and emit radiation

• Isotopes are different forms of the same element. Most occur naturally and some are manufactured. Isotopes of an element have the same number of protons and electrons but a different number of neutrons in the nucleus. i.e. they have a different mass number.

Mass number = number of protons + number of neutrons.

• Isotopes of oxygen, for example, have all eight protons but ¹⁶O has eight neutrons while ¹⁸O has ten neutrons. The superscript before the symbol for the atom designates which isotope of the element is present. It might also be written as oxygen-16 or oxygen –18.
• The additional number of neutrons in the nucleus makes some isotopes unstable and the particles in the nucleus change until they reach a more stable state. If the isotope decays by α decay, two protons and two neutrons (a helium nucleus) are emitted from the nucleus. If the isotope decays by β decay then a neutron changes to a proton + an electron and the electron leaves the nucleus. Geiger Counters or Liquid Scintillation Counters can detect both types of particles.

define what is meant by the half-life of an isotope and explain how this would affect its use in biochemistry

• Half-life is defined as the time taken for 50% of the radioactive atoms to disintegrate. i.e. half of the radioactive mass decay. The length of a half- life varies between extremely long, 710 million years for uranium-235, to very short at 0.16 seconds for polonium-216.
• Substances with short or long half-lives can be used in Biochemistry. Radioactive substances, even those with long half-lives in small doses, will not effect the functioning of an organism.

outline the evidence provided by:

• Hill and Scarisbruck
• Ruben

to confirm that the oxygen released by photosynthesis originated from water

• Hill and Scarisbrick in 1939, shone light on chloroplast preparations in the absence of carbon dioxide. They supplied the chloroplasts with salts of ferric iron such as ferricyanide or ferrioxalate to accept the hydrogen instead of carbon dioxide. The equation for the reaction, known as the Hill reaction, is

2H₂O + 4[Fe(CN)₆]^3- → 4[Fe(CN)₆]^4- + 4H⁺ + O₂

• Despite the fact that Hill and Scarisbrick were not able to show that carbon dioxide received the hydrogen, they proved that oxygen was given off in the absence of CO₂ and came from water. This suggested that CO₂ fixation and O₂ evolution are separate processes.
• In 1940 Sam Ruben used an isotope of oxygen, ¹⁸O to find out where the oxygen atoms went in photosynthesis. Ruben fed plants water containing ¹⁸O but because ¹⁸O is not a radioactive isotope of the most common form of oxygen, ¹⁶O, Ruben used a mass spectrometer to determine the fate of the ¹⁸O. The ¹⁸O was found in the oxygen gas produced by the plant and none in the glucose formed.

Photosynthesis: Electron flow and the Hill reaction , Vanderbilt University, Nashville, Tennessee, USA.