Development of the idea

The study of photosynthesis began in 1771 with observations made by the English clergyman and scientist Joseph Priestley. Priestley had burned a candle in a closed container until the air within the container could no longer support combustion. He then placed a sprig of mint plant in the container and discovered that after several days the mint had produced some substance (later recognized as oxygen) that enabled the confined air to again support combustion. In 1779 the Dutch physician Jan Ingenhousz expanded upon Priestley’s work, showing that the plant had to be exposed to light if the combustible substance (i.e., oxygen) was to be restored. He also demonstrated that this process required the presence of the green tissues of the plant.

(Read Britannica’s essay “What did Jan Ingenhousz discover about photosynthesis?”)

In 1782 it was demonstrated that the combustion-supporting gas (oxygen) was formed at the expense of another gas, or “fixed air,” which had been identified the year before as carbon dioxide. Gas-exchange experiments in 1804 showed that the gain in weight of a plant grown in a carefully weighed pot resulted from the uptake of carbon, which came entirely from absorbed carbon dioxide, and water taken up by plant roots; the balance is oxygen, released back to the atmosphere. Almost half a century passed before the concept of chemical energy had developed sufficiently to permit the discovery (in 1845) that light energy from the sun is stored as chemical energy in products formed during photosynthesis.

During the 20th century, comparisons between photosynthetic processes in green plants and in certain photosynthetic sulfur bacteria provided important information about the photosynthetic mechanism. Sulfur bacteria use hydrogen sulfide (H₂S) as a source of hydrogen atoms and produce sulfur instead of oxygen during photosynthesis. The overall reaction is

In the 1930s Dutch biologist Cornelis van Niel recognized that the utilization of carbon dioxide to form organic compounds was similar in the two types of photosynthetic organisms. Suggesting that differences existed in the light-dependent stage and in the nature of the compounds used as a source of hydrogen atoms, he proposed that hydrogen was transferred from hydrogen sulfide (in bacteria) or water (in green plants) to an unknown acceptor (called A), which was reduced to H₂A. During the dark reactions, which are similar in both bacteria and green plants, the reduced acceptor (H₂A) reacted with carbon dioxide (CO₂) to form carbohydrate (CH₂O) and to oxidize the unknown acceptor to A. This putative reaction can be represented as:

Van Niel’s proposal was important because the popular (but incorrect) theory had been that oxygen was removed from carbon dioxide (rather than hydrogen from water, releasing oxygen) and that carbon then combined with water to form carbohydrate (rather than the hydrogen from water combining with CO₂ to form CH₂O).

By 1940 chemists were using heavy isotopes to follow the reactions of photosynthesis. Water marked with an isotope of oxygen (¹⁸O) was used in early experiments. Plants that photosynthesized in the presence of water containing H₂¹⁸O produced oxygen gas containing ¹⁸O; those that photosynthesized in the presence of normal water produced normal oxygen gas. These results provided definitive support for van Niel’s theory that the oxygen gas produced during photosynthesis is derived from water.

Studies of chloroplasts

Detailed studies of the role of chloroplasts date from the work of British biochemist Robert Hill. About 1940 Hill discovered that green particles obtained from broken cells could produce oxygen from water in the presence of light and a chemical compound, such as ferric oxalate, able to serve as an electron acceptor. This process is known as the Hill reaction. During the 1950s Daniel Arnon and other American biochemists prepared plant cell fragments in which not only the Hill reaction but also the synthesis of the energy-storage compound ATP occurred. In addition, the coenzyme NADP was used as the final acceptor of electrons, replacing the nonphysiological electron acceptors used by Hill. His procedures were refined further so that small individual pieces of isolated chloroplast membranes, or lamellae, could perform the Hill reaction. These small pieces of lamellae were then fragmented into pieces so small that they performed only the light reactions of the photosynthetic process. It is now possible also to isolate the entire chloroplast so that it can carry out the complete process of photosynthesis, from light absorption, oxygen formation, and the reduction of carbon dioxide to the formation of glucose and other products.

Evidence of two light reactions

The general features of a widely accepted mechanism for photoelectron transfer, in which two light reactions (light reaction I and light reaction II) occur during the transfer of electrons from water to carbon dioxide, were proposed by Robert Hill and Fay Bendall in 1960. Many lines of evidence support the concept of electron flow via two light reactions. An early study by American biochemist Robert Emerson employed the algae Chlorella, which was illuminated with red light alone, with blue light alone, and with red and blue light at the same time. Oxygen evolution was measured in each case. It was substantial with blue light alone but not with red light alone. With both red and blue light together, the amount of oxygen evolved far exceeded the sum of that seen with blue and red light alone. These experimental data pointed to the existence of two types of light reactions that, when operating in tandem, would yield the highest rate of oxygen evolution. It is now known that light reaction I can use light of a slightly longer wavelength than red (λ = 680 nm), while light reaction II requires light with a wavelength of 680 nm or shorter.

Since those early studies, the two light reactions have been separated in many ways, including separation of the membrane particles in which each reaction occurs. As discussed previously, lamellae can be disrupted mechanically into fragments that absorb light energy and break the bonds of water molecules (i.e., oxidize water) to produce oxygen, hydrogen ions, and electrons. These electrons can be transferred to ferredoxin, the final electron acceptor of the light stage. No transfer of electrons from water to ferredoxin occurs if the herbicide DCMU is present. The subsequent addition of certain reduced dyes (i.e., electron donors) restores the light reduction of NADP⁺ but without oxygen production, suggesting that light reaction I but not light reaction II is functioning. It is now known that DCMU blocks the transfer of electrons between the first quinone and the plastoquinone pool in light reaction II.

When treated with certain detergents, lamellae can be broken down into smaller particles capable of carrying out single light reactions. One type of particle can absorb light energy, oxidize water, and produce oxygen (light reaction II), but a special dye molecule must be supplied to accept the electrons. In the presence of electron donors, such as a reduced dye, a second type of lamellar particle can absorb light and transfer electrons from the electron donor to ferredoxin (light reaction I).

The structural and photochemical properties of the minimum particles capable of performing light reactions I and II have received much study. Treatment of lamellar fragments with neutral detergents releases these particles, designated photosystem I and photosystem II, respectively. Subsequent harsher treatment (with charged detergents) and separation of the individual polypeptides with electrophoretic techniques have helped identify the components of the photosystems.

Electron flow and ATP

The theory relating the formation of ATP to electron flow in the membranes of both chloroplasts and mitochondria (the organelles responsible for ATP formation during cellular respiration) was first proposed by English biochemist Peter Dennis Mitchell, who received the 1978 Nobel Prize for Chemistry. This chemiosmotic theory has been somewhat modified to fit later experimental facts, and the general features are now widely accepted. A central feature is the formation of a hydrogen ion (proton) concentration gradient and an electrical charge across intact lamellae. The potential energy stored by the proton gradient and electrical charge is then used to drive the energetically unfavorable conversion of ADP and inorganic phosphate (P_i) to ATP and water.

Elucidation of the carbon pathway

Radioactive isotopes of carbon (¹⁴C) and phosphorus (³²P) have been valuable in identifying the intermediate compounds formed during carbon assimilation. A photosynthesizing plant does not strongly discriminate between the most abundant natural carbon isotope (¹²C) and ¹⁴C. During photosynthesis in the presence of ¹⁴CO₂, the compounds formed become labeled with the radioisotope. During very short exposures, only the first intermediates in the carbon-fixing pathway become labeled. Early investigations showed that some radioactive products were formed even when the light was turned off and the ¹⁴CO₂ was added just afterward in the dark, confirming the nature of the carbon fixation as a “dark” reaction.

American biochemist Melvin Calvin, a Nobel Prize recipient for his work on the carbon-reduction cycle, allowed green plants to photosynthesize in the presence of radioactive carbon dioxide for a few seconds under various experimental conditions. Products that became labeled with radioactive carbon during Calvin’s experiments included a three-carbon compound called 3-phosphoglycerate (PGA), sugar phosphates, amino acids, sucrose, and carboxylic acids. When photosynthesis was stopped after two seconds, the principal radioactive product was PGA, which therefore was identified as the first stable compound formed during carbon dioxide fixation in green plants. PGA is a three-carbon compound, and the mode of photosynthesis is thus referred to as C₃. In the two other known pathways, C₄ and CAM (crassulacean acid metabolism), the C₃ pathway follows the fixation of CO₂ into oxaloacetate, a four-carbon acid, and its reduction to malate. PGA is formed from 2-carboxy-3-keto-D-arabinitol 1,5-bisphosphate, which is a highly unstable six-carbon compound formed from the carboxylation of ribulose-1,5-bisphosphate, a five-carbon compound.

Further studies with ¹⁴C as well as with inorganic phosphate labeled with ³²P led to the mapping of the carbon fixation and reduction pathway called the reductive pentose phosphate (RPP) cycle, or the Calvin-Benson cycle. An additional pathway for carbon transport in certain plants was later discovered in other laboratories (see above Carbon fixation in C₄ plants). All the steps in these pathways can be carried out in the laboratory by isolated enzymes in the dark. Several steps require the ATP or NADPH generated by the light reactions. In addition, some of the enzymes are fully active only when conditions simulate those in green cells exposed to light. In living plants, these enzymes are active during photosynthesis but not in the dark.

Recent and contemporary research

Requirements for food, materials, and energy in a world where human population is rapidly growing have created a need to increase both the amount of photosynthesis and the efficiency of converting photosynthetic output into products useful to people. One response to those needs—the so-called Green Revolution, begun in the mid-20th century—achieved enormous improvements in agricultural yield through the use of chemical fertilizers, pest and plant-disease control, plant breeding, and mechanized tilling, harvesting, and crop processing. This effort limited severe famines to a few areas of the world despite rapid population growth, but it did not eliminate widespread malnutrition. Moreover, beginning in the early 1990s, the rate at which yields of major crops increased began to decline. This was especially true for rice in Asia. Rising costs associated with sustaining high rates of agricultural production, which required ever-increasing inputs of fertilizers and pesticides and constant development of new plant varieties, also became problematic for farmers in many countries.

A second agricultural revolution, based on plant genetic engineering, was forecast to lead to increases in plant productivity and thereby partially alleviate malnutrition. Since the 1970s, molecular biologists have possessed the means to alter a plant’s genetic material (deoxyribonucleic acid, or DNA) with the aim of achieving improvements in disease and drought resistance, product yield and quality, frost hardiness, and other desirable properties. However, such traits are inherently complex, and the process of making changes to crop plants through genetic engineering has turned out to be more complicated than anticipated. In the future such genetic engineering may result in improvements in the process of photosynthesis, but by the first decades of the 21st century, it had yet to demonstrate that it could dramatically increase crop yields.

Another intriguing area in the study of photosynthesis has been the discovery that certain animals are able to convert light energy into chemical energy. The eastern emerald sea slug (Elysia chlorotica), for example, acquires genes and chloroplasts from Vaucheria litorea, a yellow-green alga it consumes, giving it a limited ability to produce chlorophyll. When enough chloroplasts are assimilated, the slug may forgo the ingestion of food. The pea aphid (Acyrthosiphon pisum) can harness light to manufacture the energy-rich compound adenosine triphosphate (ATP); this ability has been linked to the aphid’s manufacture of carotenoid pigments.