Human exploration of photosynthesis began in the 18th century. Before this, people generally accepted Aristotle's authoritative view that plants obtained all the nutrients they needed from soil. It seems absurd in current day because the air and light for plant growth wasn’t considered.
18th century: photosynthesis purify air
Before this, people had long questioned the idea that plants absorbed nutrients from soil, and proposed different views. But most ideas were based on observation and philosophical reasoning. Jan Baptist van Helmont was the first man to put these ideas into practice. He planted a willow sapling weighing about 5 pounds in a large tub containing 200 pounds dry soil. Other potential nutrients are eliminated by irrigation with rainwater or distilled water. An iron plate covered tub to prevent dust from entering the soil. After five years, the willow had grown. Van Helmont found that it weighed 169 pounds, while the dry soil had only lost a few ounces. If tree had absorbed soil substances to gain mass, the soil should have decreased significantly according to mass conservation. Thus, he concluded that plant growth requires water and not soil.
In 1727, Stephen Hales discovered that air might also be one of the plant's nutrients in the study of transpiration. So, what gases are involved in photosynthesis? British chemist Joseph Priestley conducted a famous experiment in 1771. A burning candle quickly extinguished in a sealed jar. However, if a mint plant and a candle were placed in jar together, the candle could burn for a long time. The similar experiments with mice were also conducted by him. He concluded that combustion and respiration deteriorate the air, but plants purify it.
It’s strange that his experiment failed sometimes. Eight years later, Dutch physician Jan Ingenhousz pointed out that sunlight was the key to success. He placed aquatic plants in a transparent container and submerged them in water. Only when plants were exposed to sunlight did small bubbles appeared around green leaves. However, these small bubbles disappeared when in darkness. Nicolas-Théodore de Saussure found that organic matter and oxygen produced by plants far exceeded the available carbon dioxide after calculating their mass. However, the experiment only included plants, water and air, so he inferred that water was also a raw material for photosynthesis. This demonstrated that quantitative analysis is important in scientific exploration.
By this period, it’s known that plants absorb water, carbon dioxide and sunlight to generate organic matter and oxygen.
19th century: chloroplasts, energy conservation, starch, chemical equation
In 1845, German physician Robert Mayer established the concept of energy conservation that was used to explain energy conversion in photosynthesis by botanists. Sunlight is converted into chemical energy stored in organic matter. Another significant achievement was proving that starch was in organic matter. German botanist Julius von Sachs placed leaves in darkness for some time to consume as much organic matter as possible. All the leaves were then exposed to sunlight. He found that uncovered leaves turned blue in iodine solution, while the parts covered by light-blocking paper didn't.
In 1880, Engelmann used spirogyra and an aerobic bacterium to study photosynthesis. When a narrow beam of light was directed at spirogyra, aerobic bacteria were attracted to the illuminated areas. If spirogyra was fully exposed to sunlight, bacteria evenly distributed around it. These areas with aerobic bacteria happened to be abundant in chloroplasts. He used a prism to separate different colors of light in another experiments. It’s surprising that bacteria tended to concentrate in blue and red areas. Engelmann's experiments proved that chloroplasts are the sites of photosynthesis in green plants that absorb red and blue-violet light.
During this period, the classical reaction equation had already been included in textbooks. 6CO₂+6H₂O+sunlight → C₆H₁₂O₆+6O₂
20th century: light and dark reactions, more details
British botanist Frederick Blackman studied how light intensity, carbon dioxide concentration and temperature affect photosynthesis in the early 20th century. He found that increasing weak light intensity accelerates photosynthesis. If light intensity exceeds a threshold (light saturation point), its rate remains constant. At this point, only increasing temperature or carbon dioxide concentration can speed up it. For every 10°C increase in temperature, the rate of a typical chemical reaction increases 1-2 times. However, the temperature independent photochemical reactions suggested that photosynthesis isn’t a simple light-induced reaction, and a light-independent process or dark reaction must be involved in.
German biochemist Warburg tried to separate light reaction from the dark reaction. He used flashes and continuous light of same energy to study Chlorella. The photosynthesis efficiency increased by 2-4 times in flashes. This indicated that light reaction was much faster than dark reaction, but the dark reaction could catch up under instantaneous flashes. It’s now generally believed that 8 photons reduce one CO₂.
Isotope labelling provided a deeper understanding of various microscopic processes. The oxygen released in photosynthesis comes from water molecules, not carbon dioxide. Calvin spent 9 years to explore carbon dioxide fixation in unicellular green algae with the help of carbon-14. Three-carbon compounds 3-PGA and G3P were shown to be intermediates in dark reaction, so it’s also called Calvin cycle or C3 pathway. C4 and CAM pathways were discovered shortly thereafter. However, both cycles ultimately involve the C3 pathway.
In the 1950s, American scientist Arnon found strongly reducing NADPH and high-energy compound ATP in isolated spinach chloroplasts. They serve as intermediaries to transfer solar energy to carbohydrates during carbon fixation. Subsequently, the breakthrough of X-ray diffraction technology and electron microscopy resulted in discovery of several proteins involved in photosynthesis: Photosystem I absorbing short wavelengths, Photosystem II for long wavelengths, and proteins located on electron transport chain.
The chemiosmotic hypothesis for ATP production wasn’t proposed until 1980s. Protons are stored in thylakoid to create a transmembrane proton gradient. Just like a dam stores gravitational energy of water, when they pass through ATP synthase, proton potential energy is converted into chemical energy in ATP.