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chemical structure of HEPES
IUPAC name
2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid
Other names
  • 7365-45-9 ☑Y
3D model (JSmol)
  • Interactive image
  • CHEBI:42334 ☑Y
  • 22278 ☑Y
ECHA InfoCard 100.028.098
PubChem CID
  • 23831
RTECS number TY2900000
  • RWW266YE9I ☑Y
Molar mass 238.3012 g/mol
Appearance white crystalline powder
Density Not applicable
Melting point >234-238°C (453-457K)
40 g/100 ml (20°C)
Acidity (pKa) 3 (pKa1),
7.5 (pKa2)
Main hazards Irritant.
Safety data sheet External MSDS
R-phrases (outdated) R36, R37, R38.
S-phrases (outdated) S26, S36.
NFPA 704
Flammability code 0: Will not burn. E.g., water Health code 1: Exposure would cause irritation but only minor residual injury. E.g., turpentine Reactivity code 1: Normally stable, but can become unstable at elevated temperatures and pressures. E.g., calcium Special hazards (white): no codeNFPA 704 four-colored diamond
Flash point Non-flammable
Except where otherwise noted, data are given for materials in their standard state (at 25 °C [77 °F], 100 kPa).
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Infobox references

HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) is a zwitterionic organic chemical buffering agent; one of the twenty Good's buffers. HEPES is widely used in cell culture, largely because it is better at maintaining physiological pH despite changes in carbon dioxide concentration (produced by cellular respiration) when compared to bicarbonate buffers, which are also commonly used in cell culture. The dissociation of water decreases with falling temperature, but the dissociation constants (pK) of many other buffers do not change much with temperature. HEPES is like water in that its dissociation decreases as the temperature decreases. This makes HEPES a more effective buffering agent for maintaining enzyme structure and function at low temperatures.[1] Lepe-Zuniga et al. reported a phototoxicity of HEPES when exposed to ambient light by the production of hydrogen peroxide,[2][3] which is not a problem in bicarbonate-based cell culture buffers. It is therefore strongly advised to keep HEPES-containing solutions in darkness as much as possible.

HEPES has the following characteristics:

  • pKa1 (25 °C) = 3
  • pKa2 (25 °C) = 7.5
  • Useful pH range = 2.5 to 3.5 or 6.8 to 8.2

See also


  1. ^ Baicu SC, Taylor MJ (2002). "Acid-base buffering in organ preservation solutions as a function of temperature: new parameters for comparing buffer capacity and efficiency". Cryobiology. 45 (1): 33–48. doi:10.1016/S0011-2240(02)00104-9. PMID 12445548.
  2. ^ Lepe-Zuniga JL, Zigler JS, Gery I (October 1987). "Toxicity of light-exposed Hepes media". J. Immunol. Methods. 103 (1): 145. doi:10.1016/0022-1759(87)90253-5. PMID 3655381.
  3. ^ Zigler JS, Lepe-Zuniga JL, Vistica B, Gery I (May 1985). "Analysis of the cytotoxic effects of light-exposed HEPES-containing culture medium". In Vitro Cell. Dev. Biol. 21 (5): 282–7. doi:10.1007/BF02620943. PMID 4019356.
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