Hill’s reaction is defined as the production of oxygen without accompanying CO2 fixation by isolated chloroplasts or intact cells illuminated in the presence of suitable electron acceptor such as ferricyanide. Ferricyanide is reduced to ferrocyanide by photolysis of water. The reaction can be given below:
A + H2O ——–> AH2 + ½ O2
R. Hill (1937) found that when isolated chloroplasts from ground up leaves of stellaria media placed in cell extracts, produced oxygen from H2O in illuminated stage. He also noticed that cell extracts can be replaced by salts of iron like ferric oxalate. The substances when added to illuminated isolated chloroplasts in the absence of CO2. acted as hydrogen acceptors i.e oxidants. These substances are called Hill oxidants or Hill reagents.
Demonstration of Hill Reaction
Hints: Vein-less part of mucilaginous leaf are taken and homogenized in 0.25 M sucrose solution. Homogenizer is a device used for cooling and tearing of leaf fragments to the level of suspension, though muslin cloth, it is filtered. The filtrate is then centrifuged. The supernatant is discarded and the pellet is re-dispersed to get the re-suspension of chloroplasts. For the demonstration of hill reaction a colored dye dichloro-phenol indo-phenol (DCPIP) which is blue in oxidized stage can be used. It becomes colorless when reduced. Due to photolysis of water reduced DCPIP turns colorless (by isolated chloroplasts). Thus this class room activity confirms that the photosynthesis is the light activated transfer of an electron from one substance to another involving oxidation (H2O —> O2) of one and simultaneous reduction of other.