Harold Clayton Urey was born on April 29, 1893, in Walkerton, Indiana, the son of Samuel Clayton Urey, a school Teacher and a minister in the Church of the Brethren, and his wife Cora Rebecca née Reinoehl. He had a younger brother, Clarence, and a younger sister, Martha. The family moved to Glendora, California, but moved back to Indiana to live with Cora's widowed mother when Samuel became seriously ill with tuberculosis. He died when Urey was six years old.

Urey contributed a science article to The Scientific Monthly on Irving Langmuir, who invented atomic hydrogen welding in 1911 by utilizing 300 to 650 volts of electricity and tungsten filaments, and won the 1932 Nobel Prize in Chemistry for his work in surface chemistry.

Karl P. Cohen(1913–2012) was an American Chemist that worked as a research assistant under the direction of Harold Urey. Cohen’s greatest accomplishment was his work with Urey at Columbia, where he helped to perfect the enrichment techniques used in the Manhattan Project. He was a major player in the development of the centrifuge isotope separation, which is now the most commonly used way of uranium enrichment.He became an advisor for Standard Oil and worked towards the possibility of the nuclear Energy. Cohen later started his work with General Electric Company and created the Reactor Safety Committee there.

An academic career required a doctorate, so in 1921 Urey enrolled in a PhD program at the University of California, Berkeley, where he studied thermodynamics under Gilbert N. Lewis. His initial attempt at a thesis was on the ionization of cesium vapor. He ran into difficulties, and an Indian Physicist published a better paper on the same subject. Urey then wrote his thesis on the ionization states of an ideal gas, which was subsequently published in the Astrophysical Journal. After he received his PhD in 1923, Urey was awarded a fellowship by the American-Scandinavian Foundation to study at the Niels Bohr Institute in Copenhagen, where he met Werner Heisenberg, Hans Kramers, Wolfgang Pauli, Georg von Hevesy, and John Slater. At the conclusion of his stay, he traveled to Germany, where he met Albert Einstein and James Franck.

Born in Walkerton, Indiana, Urey studied thermodynamics under Gilbert N. Lewis at the University of California. After he received his PhD in 1923, he was awarded a fellowship by the American-Scandinavian Foundation to study at the Niels Bohr Institute in Copenhagen. He was a research associate at Johns Hopkins University before becoming an associate professor of Chemistry at Columbia University. In 1931, he began work with the separation of isotopes that resulted in the discovery of deuterium.

On returning to the United States, Urey received an offer of a National Research Council fellowship to Harvard University, and also received an offer to be a research associate at Johns Hopkins University. He chose the latter. Before taking up the job, he traveled to Seattle, Washington, to visit his mother. On the way, he stopped by Everett, Washington, where he knew a woman called Kate Daum. Kate introduced Urey to her sister, Frieda. Urey and Frieda soon became engaged. They were married at her father's house in Lawrence, Kansas, in 1926. The couple had four children: Gertrude Bessie (Elisabeth), born in 1927; Frieda Rebecca, born in 1929; Mary Alice, born in 1934; and John Clayton Urey, born in 1939.

At Johns Hopkins, Urey and Arthur Ruark wrote Atoms, Quanta and Molecules (1930), one of the first English texts on quantum mechanics and its applications to atomic and molecular systems. In 1929, Urey became an associate professor of Chemistry at Columbia University, where his colleagues included Rudolph Schoenheimer, David Rittenberg, and T. I. Taylor.

In 1931, Urey set out to find it. Urey and George Murphy calculated from the Balmer series that the heavy isotope should have lines redshifted by 1.1 to 1.8 ångströms (1.1×10 to 1.8×10 metres). Urey had access to a 21-foot (6.4 m) grating spectrograph, a sensitive device that had been recently installed at Columbia and was capable of resolving the Balmer series. With a resolution of 1 Å per millimetre, the machine should have produced a difference of about 1 millimetre. However, since only one atom in 4,500 was heavy, the line on the spectrograph was very faint. Urey therefore decided to delay publishing their results until he had more conclusive evidence that it was heavy hydrogen.

Around this time, william Giauque and Herrick L. Johnston at the University of California discovered the stable isotopes of oxygen. Isotopes were not well understood at the time; James Chadwick would not discover the neutron until 1932. Two systems were in use for classifying them, based on chemical and physical properties. The latter was determined using the mass spectrograph. Since it was known that the atomic weight of oxygen was almost exactly 16 times as heavy as hydrogen, Raymond Birge, and Donald Menzel hypothesized that hydrogen had more than one isotope as well. Based upon the difference between the results of the two methods, they predicted that only one hydrogen atom in 4,500 was of the heavy isotope.

Urey and Washburn attempted to use electrolysis to create pure heavy water. Their technique was sound, but they were beaten to it in 1933 by Lewis, who had the resources of the University of California at his disposal. Using the Born–Oppenheimer approximation, Urey and David Rittenberg calculated the properties of gases containing hydrogen and deuterium. They extended this to enriching compounds of carbon, nitrogen, and oxygen. These could be used as tracers in biochemistry, resulting in a whole new way of examining chemical reactions. He founded the Journal of Chemical Physics in 1932, and was its first Editor, serving in that capacity until 1940.

By the time World War II broke out in Europe in 1939, Urey was recognized as a world expert on isotope separation. Thus far, separation had involved only the light elements. In 1939 and 1940, Urey published two papers on the separation of heavier isotopes in which he proposed centrifugal separation. This assumed great importance due to speculation by Niels Bohr that uranium 235 was fissile. Because it was considered "very doubtful whether a chain reaction can be established without separating 235 from the rest of the uranium," Urey began intensive studies of how uranium enrichment might be achieved. Apart from centrifugal separation, George Kistiakowsky suggested that gaseous diffusion might be a possible method. A third possibility was thermal diffusion. Urey coordinated all isotope separation research efforts, including the effort to produce heavy water, which could be used as a neutron moderator in nuclear reactors.

Early reports on the centrifugal method indicated that it was not as efficient as predicted. Urey suggested that a more efficient but technically more complicated countercurrent system be used instead of the previous flow-through method. By November 1941, technical obstacles seemed formidable enough for the process to be abandoned. Countercurrent centrifuges were developed after the war, and today are the favored method in many countries.

Apart from his Nobel Prize, he also won the Franklin Medal in 1943, the J. Lawrence Smith Medal in 1962, the Gold Medal of the Royal Astronomical Society in 1966, and the Priestley Medal of the American Chemical Society in 1973. In 1964 he received the National Medal of Science. He became a Fellow of the Royal Society in 1947. Named after him are lunar impact crater Urey, asteroid 4716 Urey, and the H. C. Urey Prize, awarded for achievement in planetary sciences by the American Astronomical Society. The Harold C. Urey Middle School in Walkerton, Indiana, is also named for him, as is Urey Hall, the chemistry building at Revelle College, UCSD, in La Jolla. UCSD has also established a Harold C. Urey chair whose first holder is Jim Arnold.

Worn out by the effort, Urey left the project in February 1945, handing over his responsibilities to R. H. Crist. The K-25 plant commenced operation in March 1945, and as the bugs were worked out, the plant operated with remarkable efficiency and economy. For a time, uranium was fed into the S50 liquid thermal diffusion plant, then the K-25 gaseous, and finally the Y-12 electromagnetic separation plant; but soon after the war ended the thermal and electromagnetic separation plants were closed down, and separation was performed by K-25 alone. Along with its twin, K-27, constructed in 1946, it became the principal isotope separation plant in the early post-war period. For his work on the Manhattan Project, Urey was awarded the Medal for Merit by the Project Director, Major General Leslie R. Groves, Jr.

Urey actively campaigned against the 1946 May-Johnson bill because he feared that it would lead to military control of nuclear Energy, but supported and fought for the McMahon bill that replaced it, and ultimately created the Atomic Energy Commission. Urey's commitment to the ideal of world government dated from before the war, but the possibility of nuclear war made it only more urgent in his mind. He went on lecture tours against war, and became involved in Congressional debates regarding nuclear issues. He argued publicly on behalf of Ethel and Julius Rosenberg, and was even called before the House Un-American Activities Committee.

In the late 1950s and early 1960s, space science became a topical field of research in the wake of the launch of Sputnik I. Urey helped persuade NASA to make unmanned probes to a moon a priority. When Apollo 11 returned moon rock samples from the moon, Urey examined them at the Lunar Receiving Laboratory. The samples supported Urey's contention that the moon and the Earth shared a Common origin. While at UCSD, Urey published 105 scientific papers, 47 of them about lunar topics. When asked why he continued to work so hard, he joked, "Well, you know I’m not on tenure anymore."

In later life, Urey helped develop the field of cosmochemistry and is credited with coining the term. His work on oxygen-18 led him to develop theories about the abundance of the chemical elements on earth, and of their abundance and evolution in the stars. Urey summarized his work in The Planets: Their Origin and Development (1952). Urey speculated that the early terrestrial atmosphere was probably composed of ammonia, methane, and hydrogen. One of his Chicago graduate students, Stanley L. Miller, showed in the Miller–Urey experiment that, if such a mixture be exposed to electric sparks and to water, it can interact to produce amino acids, commonly considered the building blocks of life.

Urey spent a year as a visiting professor at Oxford University in England in 1956 and 1957. In 1958, he reached the University of Chicago's retirement age of 65, but he accepted a post as a professor at large at the new University of California, San Diego (UCSD), and moved to La Jolla, California. He was subsequently made a professor emeritus there from 1970 to 1981. Urey helped build up the science faculty there. He was one of the founding members of UCSD's school of chemistry, which was created in 1960, along with Stanley Miller, Hans Suess, and Jim Arnold.

Urey speculated that the early terrestrial atmosphere was probably composed of ammonia, methane, and hydrogen. One of his Chicago graduate students was Stanley L. Miller, who showed in the Miller–Urey experiment that, if such a mixture were exposed to electric sparks and water, it can interact to produce amino acids, commonly considered the building blocks of life. Work with isotopes of oxygen led to pioneering the new field of paleoclimatic research. In 1958, he accepted a post as a professor at large at the new University of California, San Diego (UCSD), where he helped create the science faculty. He was one of the founding members of UCSD's school of chemistry, which was created in 1960. He became increasingly interested in space science, and when Apollo 11 returned moon rock samples from the moon, Urey examined them at the Lunar Receiving Laboratory. Lunar Astronaut Harrison Schmitt said that Urey approached him as a volunteer for a one-way mission to the Moon, stating "I will go, and I don't care if I don't come back."