Yellapragada Subbarow Co-Created a Phosphorus Test Cited More Than 21,000 Times, Was Among the First to Report ATP in 1929 and Directed the Lederle Laboratories Research Behind Folic Acid, Aminopterin and Aureomycin

Yellapragada Subbarow and Cyrus Fiske published “The colorimetric determination of phosphorus”, a test that reads the amount of phosphate in a sample from the strength of a colour, in the Journal of Biological Chemistry in 1925. Ankit Bharti and Yogesh Marfatia, writing in the Indian Journal of Dermatology, Venereology and Leprology, put its citations at more than 21,000, and Hektoen International counted 22,547 in October 2013. Subbarow was born on 12 January 1895 in Bhimavaram, in the West Godavari district of today’s Andhra Pradesh, and went on to head research at Lederle Laboratories in New York State, where his teams worked on folic acid, the leukaemia drug aminopterin and the antibiotic aureomycin.

His surname appears as Subbarow, SubbaRow and Subba Row in the literature. This article uses the first.

From Bhimavaram to Harvard in 1923

Subbarow came from a Telugu Brahmin family, Bharti and Marfatia write. He entered Madras Medical College and, answering Mahatma Gandhi’s call during the non-cooperation movement, began wearing a khadi gown there in August 1920. His surgery professor, M C Bradfield, was displeased, and Subbarow was given the lesser Licentiate of Medicine and Surgery certificate instead of an MBBS. That kept him out of the Madras Medical Services, so he took a post as lecturer in anatomy and physiology at the Madras Ayurvedic College, where he rose to vice principal.

A visiting American doctor, John Fox Kendrick, encouraged him to apply to the Harvard School of Tropical Medicine. A loan from his father-in-law and a scholarship of Rs 1,500 a year for three years from the MSN charities paid for the move. His licentiate did not qualify him for a Harvard scholarship or internship, so he worked as a night porter, cleaning bedpans and urinals, and took odd jobs besides.

William Rosen, in his history of antibiotics, dates the arrival in the United States to 1923. Subbarow earned the tropical medicine diploma in June 1924 and joined Harvard Medical School, where Professor Otto Folin assigned him to Fiske, who had been set the task of measuring phosphorus accurately in cell, nerve and bone tissue.

How the phosphorus test worked

A colorimetric test adds chemicals to a sample and reads the amount of the target substance from the colour that develops. Lakshna Mahajan and colleagues, in an Indian Journal of Clinical Biochemistry editorial, say Subbarow chose aminonaphtholsulfonic acid as the reducing agent in the reaction. It was an isomer, a different arrangement of the same atoms, that was easier to prepare than the one he first tried and could be bought from the dye industry. They report that the colour developed in under a minute and that the test tolerated interfering substances at ten times the level that the older reagent hydroquinone allowed.

The method held its place for decades. In his Nobel lecture of 11 December 1953, Fritz Lipmann still described measuring phosphate by “the ordinary Fiske-Subbarow procedure”. Subbarow took his PhD at the Harvard commencement in June 1929, according to Bharti and Marfatia.

Phosphocreatine in 1927, ATP in 1929

With a reliable phosphate test, Fiske and Subbarow could ask what form phosphorus takes in muscle. Science carried their paper “The nature of the ‘inorganic phosphate’ in voluntary muscle” on 22 April 1927. Mahajan and colleagues summarise the finding: much of the phosphate in muscle filtrates was not free phosphate but a compound containing creatine and phosphate in equal amounts, which broke down when muscle contracted and was rebuilt during recovery. That compound is phosphocreatine. Beerelli Seshi and Marshall Lichtman describe it as supplying energy to muscle and brain at times of high demand.

ATP, adenosine triphosphate, is the molecule cells use to carry energy from where it is made to where it is spent. Seshi and Lichtman call it the principal energy source for metabolic processes, and Mahajan and colleagues date its characterisation to 1929. Two papers that year bear on who was first. Karl Lohmann published “Über die Pyrophosphatfraktion im Muskel” (“On the pyrophosphate fraction in muscle”) in the journal Naturwissenschaften in August 1929. Fiske and Subbarow’s note “Phosphorus compounds of muscle and liver” appeared in Science on 18 October 1929.

Mahajan and colleagues write that textbooks cite Lohmann as the discoverer of ATP, that his work is thought to have been influenced by earlier discussions with Fiske, and that he also used the Fiske-Subbarow technique.

A faculty post that never came, and Lederle in 1940

Harvard did not give Subbarow a regular faculty position. Rosen puts the reason down to immigration status: as an immigrant from British India he had to register as an alien throughout his career, and Rosen calls the denial of tenure the most consequential result. Kutikuppala and colleagues, in a 2025 review in Annals of Medicine and Surgery, attribute it to institutional and racial bias and also cite his unconventional research approach and lack of formal credentials.

In 1940 he joined Lederle Laboratories at Pearl River, New York, a company that the agricultural chemicals maker American Cyanamid had owned since 1930. Rosen calls him its director of research. Bharti and Marfatia report that Lederle offered $14,000 a year and that Subbarow said he would accept half if the company built a research building for him.

Folic acid, aminopterin and the first leukaemia remissions

Folic acid is a B vitamin, also called vitamin B9. On 31 August 1945 Science published “Synthesis of a compound identical with the L. casei factor isolated from liver” by Robert Angier and 15 co-authors, among them Subbarow, listed seventh. Bharti and Marfatia credit a Lederle team under him with a method to isolate and synthesise folic acid on an industrial scale.

Sidney Farber, a pathologist at Boston Children’s Hospital, then asked whether blocking the vitamin might slow leukaemia. Kutikuppala and colleagues write that Farber had given folic acid to children with leukaemia and seen tumour growth speed up, and that he persuaded Subbarow and colleagues to develop a folic acid antagonist, a chemical that interferes with the vitamin’s action. Subbarow’s group synthesised aminopterin. On 3 June 1948 Farber, L K Diamond and co-authors reported in the New England Journal of Medicine temporary remissions in children with acute leukaemia treated with it. Bharti and Marfatia say the paper acknowledged Subbarow and his team, and Seshi and Lichtman credit his work with the first remissions in childhood acute leukaemia.

Methotrexate is a modified form of aminopterin, according to Kutikuppala and colleagues. Bharti and Marfatia describe an internal memo in which Subbarow directed his team to look for a less toxic compound, which led to it. They report that on the early drugs leukaemia relapsed in all the patients whose disease had been held in check, and that most clinicians in India have prescribed methotrexate, which is cheap and effective for psoriasis. The patent law behind India’s low-cost medicines is covered in how India became the world’s pharmacy.

A filariasis drug and an antibiotic from soil

After the attack on Pearl Harbor in 1941, Bharti and Marfatia write, Subbarow added drugs against parasites to Lederle’s programme to protect American soldiers in tropical Asia. The result was diethylcarbamazine. A 1947 paper in the Journal of Laboratory and Clinical Medicine, “Experimental chemotherapy of filariasis III”, by Reginald Hewitt and five co-authors including Y Subbarow, tested a related compound against naturally acquired filarial infections in cotton rats and dogs. Filariasis is a disease caused by parasitic worms. Bharti and Marfatia say the World Health Organization recognises diethylcarbamazine as a principal drug for preventing and treating it.

Lederle was also hunting for new antibiotics. Rosen says Subbarow hired Benjamin Duggar, a plant pathologist then in his seventies, and that the two began a worldwide soil collection, with soldiers and sailors sending samples from places as far apart as the Caucasus and South America. Accounts of when Duggar arrived differ, from 1941 in Bharti and Marfatia to 1944 in Rosen.

A marker erected in 1959 at the University of Missouri’s Sanborn Field records the next step. Plot 23, planted with timothy grass since 1888, supplied one of a dozen soil samples that William Albrecht sent to Lederle in August 1945, and Duggar isolated the organism from it. Rosen calls it a yellow soil microbe, an actinomycete, and says it took nearly three years of testing before Duggar announced it, naming it Streptomyces aureofaciens, the “gold maker”, and the drug aureomycin. It acted against both Gram-positive and Gram-negative bacteria, the two broad groups that the Gram stain separates.

Credit is shared in the accounts. The Missouri marker names Duggar as the isolator. Seshi and Lichtman say Subbarow directed the research that produced the first tetracycline, the family of antibiotics to which aureomycin belongs. Bharti and Marfatia write that he took a backseat when Duggar presented aureomycin at a New York Academy of Sciences meeting, and that he died about a fortnight later. Somasekhar gives the date as 8 August 1948, aged 53, and Bharti and Marfatia give 9 August.

What the Nobel record shows

Subbarow did not receive a Nobel Prize, Seshi and Lichtman write. The Nobel Foundation’s nomination archive lists no nomination for him or for Fiske in its year-by-year lists for medicine from 1926 to 1953 (the archive has released medicine only to 1953) or for chemistry from 1926 to 1974. Lohmann appears in the medicine lists three times in 1952, each nomination from a different professor in Münster. The 1953 medicine prize went to Hans Krebs for the citric acid cycle and to Fritz Lipmann for coenzyme A.

Others read the gap as a failure of credit. His biographer S P K Gupta argued in January 2020 that Subbarow did not receive due credit, which BioVoice reports he linked partly to British scientific dominance and colonial constraints. Kutikuppala and colleagues write that the scientific community largely overlooked his work although textbooks include it. The priority record is mixed: Lohmann’s note came about two months before Fiske and Subbarow’s, textbooks name Lohmann, and Fiske, who shared the muscle work, also has no nomination in the archive. For an Indian-born biochemist who did win, see Har Gobind Khorana and the genetic code, and for another priority question in Indian science, CV Raman and the effect that bears his name.

How India has remembered him

The Government of India issued a Rs 1 commemorative stamp in 1995, the centenary year of his birth. M Somasekhar, a former associate editor of The Hindu BusinessLine, lists a bust at the Nizam Institute of Medical Sciences in Hyderabad, a statue at Guntur Medical College, a plaque and a library at Lederle’s research laboratory, a building block at the National Institute of Nutrition, and auditoriums at the University of Hyderabad and GITAM University in Visakhapatnam.

Guru Gobind Singh Indraprastha University in Delhi has held an annual Subbarow memorial lecture since 2002. In January 2020 its vice chancellor announced an endowment to make the lecture permanent, timed to Subbarow’s 125th birth anniversary, and Gupta announced a contribution of Rs 1 lakh (Rs 100,000) toward it.