PHOTOSYNTHESISIN HIGHER PLANTS Another experiment you may have carried out is the half-leaf experiment, where a part of a leaf is enclosed in a test tube containing some KOH soaked cotton (which absorbs CO2), while the other half is exposed to air. The setup is then placed in light for some time. On testing for starch later in the two halves of the leaf, you must have found that the exposed part of the leaf tested positive for starch while the portion that was in the tube, tested negative. This showed that CO2 was required for photosynthesis. Can you explain how this conclusion could be drawn? (a) (b) 13.2 EARLY EXPERIMENTS It is interesting to learn about those simple experiments that led to a gradual development in our understanding of photosynthesis. Joseph Priestley (1733-1804) in 1770 performed a series of experiments that revealed the essential role of air in the growth of green plants. Priestley, you may recall, discovered oxygen in 1774. Priestley observed that a candle burning in a closed space – a bell jar, soon gets extinguished (Figure 13.1 a, b, c, d). Similarly, a mouse would (c) (d)soon suffocate in a closed space. He concluded that Figure 13.1 Priestley’s experimenta burning candle or an animal that breathe the air, both somehow, damage the air. But when he placed a mint plant in the same bell jar, he found that the mouse stayed alive and the candle continued to burn. Priestley hypothesised as follows: Plants restore to the air whatever breathing animals and burning candles remove. Can you imagine how Priestley would have conducted the experiment using a candle and a plant? Remember, he would need to rekindle the candle to test whether it burns after a few days. How many different ways can you think of to light the candle without disturbing the set-up? Using a similar setup as the one used by Priestley, but by placing it once in the dark and once in the sunlight, Jan Ingenhousz (1730-1799) showed that sunlight is essential to the plant process that somehow purifies the air fouled by burning candles or breathing animals. Ingenhousz in an elegant experiment with an aquatic plant showed that in bright sunlight, small bubbles were formed around the green parts while in the dark they did not. Later he identified these bubbles to be of oxygen. Hence he showed that it is only the green part of the plants that could release oxygen. PHOTOSYNTHESIS IN HIGHER PLANTS of the F0 of the ATPase. The ATPase enzyme consists of two parts: one called the F0 is embedded in the membrane and forms a transmembrane channel that carries out facilitated diffusion of protons across the membrane. The other portion is called F1 and protrudes on the outer surface of the thylakoid membrane on the side that faces the stroma. The break down of the gradient provides enough energy to cause a conformational change in the F1 particle of the ATPase, which makes the enzyme synthesise several molecules of energy-packed ATP. Chemiosmosis requires a membrane, a proton pump, a proton gradient and ATPase. Energy is used to pump protons across a membrane, to create a gradient or a high concentration of protons within the thylakoid lumen. ATPase has a channel that allows diffusion of protons back across the membrane; this releases enough energy to activate ATPase enzyme that catalyses the formation of ATP. Along with the NADPH produced by the movement of electrons, the ATP will be used immediately in the biosynthetic reaction taking place in the stroma, responsible for fixing CO2, and synthesis of sugars. 13.7 WHERE ARE THE ATP AND NADPH USED? We learnt that the products of light reaction are ATP, NADPH and O. Of 2 these O2 diffuses out of the chloroplast while ATP and NADPH are used to drive the processes leading to the synthesis of food, more accurately, sugars. This is the biosynthetic phase of photosynthesis. This process does not directly depend on the presence of light but is dependent on the products of the light reaction, i.e., ATP and NADPH, besides CO2 and H O. You may wonder how this could be verified; it is simple: immediately2after light becomes unavailable, the biosynthetic process continues for some time, and then stops. If then, light is made available, the synthesis starts again. Can we, hence, say that calling the biosynthetic phase as the dark reaction is a misnomer? Discuss this amongst yourselves. Let us now see how the ATP and NADPH are used in the biosynthetic phase. We saw earlier that CO is combined with H O to produce (CHO)222n or sugars. It was of interest to scientists to find out how this reaction proceeded, or rather what was the first product formed when CO is taken 2into a reaction or fixed. Just after world war II, among the several efforts to put radioisotopes to beneficial use, the work of Melvin Calvin is exemplary. The use of radioactive 14C by him in algal photosynthesis studies led to the discovery that the first CO fixation product was a23-carbon organic acid. He also contributed to working out the complete biosynthetic pathway; hence it was called Calvin cycle after him. The first product identified was 3-phosphoglyceric acid or in short PGA. How many carbon atoms does it have? PHOTOSYNTHESIS IN HIGHER PLANTS incident light beyond a point causes the breakdown of chlorophyll and a decrease in photosynthesis. 13.10.2 Carbon dioxide Concentration Carbon dioxide is the major limiting factor for photosynthesis. The concentration of CO2 is very low in the atmosphere (between 0.03 and 0.04 per cent). Increase in concentration upto 0.05 per cent can cause an increase in CO2 fixation rates; beyond this the levels can become damaging over longer periods. The C3 and C4 plants respond differently to CO2 concentrations. At low light conditions neither group responds to high CO2 conditions. At high light intensities, both C3 and C4 plants show increase in the rates of photosynthesis. What is important to note is that the C4 plants show saturation at about 360 µlL-1 while C3 responds to increased CO2 concentration and saturation is seen only beyond 450 µlL-1. Thus, current availability of CO2 levels is limiting to the C3 plants. The fact that C3 plants respond to higher CO2 concentration by showing increased rates of photosynthesis leading to higher productivity has been used for some greenhouse crops such as tomatoes and bell pepper. They are allowed to grow in carbon dioxide enriched atmosphere that leads to higher yields. 13.10.3 Temperature The dark reactions being enzymatic are temperature controlled. Though the light reactions are also temperature sensitive they are affected to a much lesser extent. The C plants respond to higher temperatures and4show higher rate of photosynthesis while C plants have a much lower3temperature optimum. The temperature optimum for photosynthesis of different plants also depends on the habitat that they are adapted to. Tropical plants have a higher temperature optimum than the plants adapted to temperate climates. 13.10.4 Water Even though water is one of the reactants in the light reaction, the effect of water as a factor is more through its effect on the plant, rather than directly on photosynthesis. Water stress causes the stomata to close hence reducing the CO2 availability. Besides, water stress also makes leaves wilt, thus, reducing the surface area of the leaves and their metabolic activity as well.