In biochemistry, chemosynthesis is the biological conversion of one or more carbon-containing molecules (usually carbon dioxide or methane) and nutrients into organic matter using the oxidation of inorganic compounds (e.g., hydrogen gas, hydrogen sulfide) or ferrous ions as a source of energy, rather than sunlight, as in photosynthesis. Chemoautotrophs, organisms that obtain carbon from carbon dioxide through chemosynthesis, are phylogenetically diverse. Groups that include conspicuous or biogeochemically important taxa include the sulfur-oxidizing Gammaproteobacteria, the Campylobacterota, the Aquificota, the methanogenic archaea, and the neutrophilic iron-oxidizing bacteria.

Many microorganisms in dark regions of the oceans use chemosynthesis to produce biomass from single-carbon molecules. Two categories can be distinguished. In the rare sites where hydrogen molecules (H₂) are available, the energy available from the reaction between CO₂ and H₂ (leading to production of methane, CH₄) can be large enough to drive the production of biomass. Alternatively, in most oceanic environments, energy for chemosynthesis derives from reactions in which substances such as hydrogen sulfide or ammonia are oxidized. This may occur with or without the presence of oxygen.

Many chemosynthetic microorganisms are consumed by other organisms in the ocean, and symbiotic associations between chemosynthesizers and respiring heterotrophs are quite common. Large populations of animals can be supported by chemosynthetic secondary production at hydrothermal vents, methane clathrates, cold seeps, whale falls, and isolated cave water.

It has been hypothesized that anaerobic chemosynthesis may support life below the surface of Mars, Jupiter's moon Europa, and other planets. Chemosynthesis may have also been the first type of metabolism that evolved on Earth, leading the way for cellular respiration and photosynthesis to develop later.

Giant tube worms use bacteria in their trophosome to fix carbon dioxide (using hydrogen sulfide as their energy source) and produce sugars and amino acids. Some reactions produce sulfur:

Instead of releasing oxygen gas while fixing carbon dioxide as in photosynthesis, hydrogen sulfide chemosynthesis produces solid globules of sulfur in the process. In bacteria capable of chemoautotrophy (chemosynthesis), such as purple sulfur bacteria, yellow globules of sulfur are present and visible in the cytoplasm.

In 1890, Sergei Winogradsky proposed a novel type of life process called "anorgoxydant". His discovery suggested that some microbes could live solely on inorganic matter and emerged during his physiological research in the 1880s in Strasbourg and Zürich on sulfur, iron, and nitrogen bacteria.

In 1897, Wilhelm Pfeffer coined the term "chemosynthesis" for the energy production by oxidation of inorganic substances, in association with autotrophic carbon dioxide assimilation—what would be named today as chemolithoautotrophy. Later, the term would be expanded to include also chemoorganoautotrophs, which are organisms that use organic energy substrates in order to assimilate carbon dioxide. Thus, chemosynthesis can be seen as a synonym of chemoautotrophy.