Requirement of Amino Acids

Countries and regions that have established dietary reference intakes for indispensable amino acids

Germany, Austria, Switzerland
South Korea

(As of March 2023)
Source: National Institute of Health and Nutrition, Japan: Summary of dietary reference intakes

Indispensable amino acid requirements (WHO)

Amino acid Requirement References
(mg/kg/day) (mg/kg/day) Methods Number
Lysine 30 29 24-hour indicator amino acid oxidation method 1)
31 24-hour indicator amino acid oxidation method 2)
Leucine 39 37.3 24-hour [13C] leucine balance study 3)
39.6 24-hour [13C] leucine balance study 4)
Isoleucine 20 20 Based on the amino acid composition of body protein, calculated from a value of 39 mg/kg/day for leucine requirement. 5)
Valine 26 26 Same as above 5)
Threonine 15 15 24-hour [13C] leucine balance study 6)
15 24-hour [13C] leucine balance study 7)


25 - The value is close to the midpoint of a range of requirement estimates all of which have some considerable uncertainty. -
Tryptophan 4 4

Based on an average of values derived from a variety of approaches.

One of these is the indicator amino acid oxidation method.


15 - Rounded sum value -
Methionine 10 10.4 Calculated from obligatory nitrogen loss (47.7 mg/kg/day) and total sulfur amino acid content of tissue protein (the mean of liver and muscle protein) 10) - 12)
Cysteine 5 4.1 Same as above 10) - 12)
Histidine 10 8 - 12 Since it has not been possible to determine histidine requirement experimentally, the conclusion of the previous report was endorsed. 13)
Total indispensable amino acids 184
  1. Kurpad AV et al., Lysine requirements of healthy adult Indian subjects, measured by an indicator amino acid balance technique. Am J Clin Nutr. 73(5):900-7, 2001.
  2. Kurpad AV et al., Lysine requirements of healthy adult Indian subjects receiving long-term feeding, measured with a 24-h indicator amino acid oxidation and balance technique., Am J Clin Nutr. 76(2):404-12, 2002.
  3. Kurpad AV et al., Daily requirement for and splanchnic uptake of leucine in healthy adult Indians. Am J Clin Nutr. 74(6):747-55, 2001.
  4. Kurpad AV et al., Leucine requirement and splanchnic uptake of leucine in chronically undernourished adult Indian subjects. Am J Clin Nutr. 77(4):861-7, 2003.
  5. Widdowson EM et al., Body composition of the fetus and infant. Nutrition and Metabolism of the Fetus and Infant. London, Nijhoff, pp 169–177, 1979.
  6. Borgonha S et al., Threonine requirement of healthy adults, derived with a 24-h indicator amino acid balance technique. Am J Clin Nutr. 75(4):698-704, 2002.
  7. Kurpad AV et al., Threonine requirements of healthy Indian men, measured by a 24-h indicator amino acid oxidation and balance technique. Am J Clin Nutr. 76(4):789-97, 2002.
  8. Lazaris-Brunner G et al., Tryptophan requirement in young adult women as determined by indicator amino acid oxidation with L-[13C]phenylalanine. Am J Clin Nutr. 68(2):303-10, 1998.
  9. Rand WM et al., Meta-analysis of nitrogen balance studies for estimating protein requirements in healthy adults. Am J Clin Nutr. 77(1):109-27, 2003.
  10. Paul AA et al., First supplement to McCance and Widdowson's The Composition of Foods. London, Her Majesty's Stationery Office, 1980.
  11. ExPASy proteomics server. Swiss institute of Bioinformatics
  12. Millward DJ, An adaptive metabolic demand model for protein and amino acid requirements. Br J Nutr. 90(2):249-60, 2003.
  13. Energy and protein requirements. Report of a Joint FAO/WHO/UNU Expert Consultation. Genebva, World Health Organization, 1985 (WHO Technical Report Series, No. 724)

General methods used for determining protein and amino acid requirements

1. Nitrogen balance method

Concept Outline method
  • Protein/amino acids is by far the major nitrogen-containing substance in the body, so that gain or loss of nitrogen from the body can be regarded as synonymous with gain or loss of protein/amino acids.
  • It is implicit in the method that in the healthy subject, body nitrogen will be constant (in the adult) if the dietary intake of the specific test nutrient, such as an indispensable amino acid, is adequate. It follows from this that if body nitrogen is decreasing, then the diet is deficient.
  • It is assumed that maintenance of the body amino acid mass is synonymous with health.
  • All intakes and losses of nitrogen are quantified accurately to determine the difference between intakes and losses.
Advantages Disadvantages
  • Classical method
  • There is a possibility that the amount of ingestion that was not ingested, such as spillage and residue on plates, may be included in the amount of ingestion, and the amount of ingestion may be overestimated.
  • All losses cannot be quantified, and the loss may be underestimated.
  • As a result, amino acid requirements will tend to be underestimated.
  • It takes more than a week to adapt to the experimental diet.

2. Carbon balance method

Concept Outline method
  • The requirement of a specific indispensable amino acid in adults in the dietary intake of that amino acid which balances all routes of loss.
  • After intravenously infusion of a stable isotope labeled with the carboxyl group of a specific indispensable amino acid to be evaluated, measurements are made of the expiration rate of labeled carbon dioxide in the breath, and the oxidation rate of the amino acid is calculated.
  • There is the 24-h protocol, the fed-only protocol (3-5 h-infusion), and the short-term fasting/feeding protocol (3h-fasted + 5h-fed during 8h-infusion).
Advantages Disadvantages
  • The protocol for assessing carbon balance has been refined to such a degree that few serious potential sources of error remain.
  • Measurements of amino acid oxidation are generally of between 4 and 24 hours’ duration.
  • The subject is confined on a bed or chair, with limited activity, both because of the infusion lines and blood sampling, but also to prevent large changes in the rate of CO2 production.
  • The problem remains that the extent to which the experimental feeding protocol is representative of a normal day.
  • It is difficult to ensure that sufficient different intake levels are fed around the requirement level.

3. Indicator amino acid oxidation method

Concept Outline method
  • If one amino acid in the diet is below requirement (i.e. is limiting), then all other indispensable amino acids cannot be fully utilized for protein synthesis and the excess is therefore oxidized.
  • A stable isotope labeled with a carboxyl group of a fixed amount of an indicator amino acid is orally ingested, and an expiration rate of labeled carbon dioxide in the breath is measured, and an oxidation rate of the indicator amino acid at each intake of the test amino acid is calculated.
  • Each subject repeats this procedure 6 or 7 times with varying test amino acid intakes.
  • A “breakpoint” in the curve for oxidation of the “indicator” amino acid against the intake of the test amino acid is determined.
Advantages Disadvantages
  • There is no problem of giving nutritionally significant quantities because the tracer is separate from the test amino acid.
  • It is practically simple resulting from the lack of need to make balance measurements or for prior dietary adaptation to each of the varying intake levels.
  • The theoretical basis of the approach differs from other methods, and what is measured bears a different and quite complex relationship to the amino acid requirement measured by the traditional balance approaches.
  • In the human adult in overall balance, because net protein synthesis may be less intense than in a growing animal, the demonstration of a breakpoint is often much less convincing.
  • The problem remains that the extent to which the experimental feeding protocol is representative of a normal day.

4. Prediction from the obligatory nitrogen loss

Concept Outline method
  • The individual amino acids contribute to the obligatory nitrogen loss in subjects on a protein-free diet in proportion to their representation in whole-body protein.
  • Obligatory oxidative losses (mg/kg/day) = obligatory nitrogen losses (mg nitrogen/kg/day) x amino acid (mg/g tissue nitrogen)
  • Adjusted for the efficiency of utilization.
Advantages Disadvantages
  • Values could be derived for amino acids for which there is a lack of experimental data.
  • The requirement of only one amino acid can be identified in this way, i.e. the limiting amino acid, with the highest ratio of metabolic demand for maintenance to tissue protein content.


Protein and amino acid requirements in human nutrition. WHO Technical Reports Series 935.