HEPES Buffer CAS 7365-45-9

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  • Appearance: Colorless liquid
  • Assay: 99. 0%min
  • Stock: In stock
  • Sample: Available
  • Zhishang Chemical: HEPES buffer supplement

Basic Info of HEPES Buffer

What is HEPES Buffer?

HEPES buffer(4 – (2-hydroxyethyl) – 1-piperazine ethanesulfonic acid) is a zwitterionic sulfonic acid buffer; One of good’s 20 buffers. Hepes buffer is widely used in cell culture, mainly because it can maintain physiological pH value better than bicarbonate buffer, which is also commonly used in cell culture, although the concentration of carbon dioxide (produced by aerobic respiration) changes. Lepe Zuniga et al. An unwanted photochemical process in which hepes buffer produces hydrogen peroxide when exposed to ambient light is reported, which is not a problem in bicarbonate based cell culture buffer. Therefore, it is strongly recommended to place the solution containing hepes buffer in the dark as far as possible to prevent oxidation.

HEPES buffer has the following characteristics:

p K a1 (25 °C) = 3
p K a2 (25 °C) = 7.5
Useful pH range = 2.5 to 3.5 or 6.8 to 8.2

The negligible metal ion binding of hepes buffer makes it an ideal buffer for enzymes that may be inhibited by metal chelation.

HEPES Buffer Uses

  1. HEPES buffer(4-hydroxyethylpiperazineethanesulfonic acid) is a non-ionic amphoteric buffer, highly polar, inert to a variety of chemical reagents and enzymes, does not participate in and does not interfere with the process of biochemical reactions, and is resistant to enzymatic chemical reactions. There is no inhibitory effect on the reaction, so it can be specially used for the research of organelles and highly volatile, pH-sensitive proteins and enzymes.
  2. Cell-cell adhesion, short-term cell aggregation culture, buffer for washing tissues and cells.
  3. Patent CN201310477071.6 discloses a method for preparing porous zinc oxide microspheres by using HEPES buffer(4-hydroxyethylpiperazine ethanesulfonic acid) as a template. The method is simple to operate, and the prepared microspheres have the advantages of uniform size and hierarchical pore structure, etc., and can be used for the production of catalysts and gas sensing elements.
  4. Patent CN201080052103.2 uses HEPES buffer(4-hydroxyethylpiperazine ethanesulfonic acid) and its derivatives to treat cancer pain and suppress the side effects after chemotherapy.
  5. Hydroxyethylpiperazineethanesulfonic acid is a common component of buffers in animal and human cell cultures. Studies have confirmed that HEPES buffer(4-hydroxyethylpiperazine ethanesulfonic acid) has minimal cytotoxicity among all known buffers. The FDA (US Food and Drug Administration) has approved piperazine-based zwitterionic molecules as food additives.

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Application of HEPES Buffer

The enhancement effect of 4-hydroxyethylpiperazineethanesulfonic acid(HEPES buffer) on tuna peptide xanthine oxidase inhibitor, the specific experimental steps are as follows:

  1. Prepare 150mM 4-hydroxyethylpiperazineethanesulfonic acid(HEPES buffer), Tris-HCl, and PB buffer at 37°C with pH 7.4, and store at 4°C until use;
  2. Prepare the required sample (tuna peptide) with the above-mentioned 150 mM buffer, and the sample concentration is 40 mg/ml;
  3. Add 50 μL tuna peptide or 50 μL buffer (blank) and 150 μL xanthine in sequence to the 96-well microtiter plate, and do 3 parallels for each sample, incubate at 37°C for 5 min, add 50 μL xanthine oxidase, and read once in 30 s Absorbance value, a total of 50 times 25min. After reading, the reaction was terminated with 80 μL of HCl, and the reaction solution was diluted 10 times with the corresponding buffer and passed through a 0.25 μm aqueous membrane to measure the uric acid concentration;
  4. Measure the content of uric acid in the sample after the reaction by high performance liquid chromatography, and quantify it with a uric acid standard;
  5. Calculate the inhibition rate of xanthine oxidase, and the calculation method is as follows:

    I=(CB-CS)/CB×100%

    In the formula: I represents the inhibition rate of xanthine oxidase, CB is the peak area of the uric acid peak in the liquid phase spectrum after the reaction in the blank group (without adding the peptide), and CS represents the peak area of the uric acid peak in the liquid phase spectrum after the reaction in the sample group (with the peptide added).

In this method, the preparation process of tuna peptides is as follows: the tuna fish meat is crushed and mixed with deionized water according to the mass ratio of material to liquid of 1:1, the pH value is adjusted to 7.0, and compound protease and papain are respectively added according to 0.6-1.2% of the mass of tuna. , enzymolysis at 50-60°C for 4-7 hours, after the enzymolysis is completed, the temperature is raised to 95°C for 15 minutes to inactivate the enzyme, cooled, centrifuged, and the supernatant is concentrated and spray-dried to obtain tuna peptides.

In 4-hydroxyethylpiperazine ethanesulfonic acid buffer(HEPES buffer), Tris buffer and PB buffer, the content of uric acid produced by xanthine oxidase catalyzed by xanthine has almost no difference, while HEPES can significantly increase the xanthine content of tuna polypeptide oxidase inhibition rate, this result indicates that 4-hydroxyethylpiperazine ethanesulfonic acid(HEPES buffer) can enhance the inhibitory activity of polypeptide xanthine oxidase.

Preparation of HEPES Buffer

References

  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”. Cryobiology45 (1): 33–48.
  2. Lepe-Zuniga JL, Zigler JS, Gery I (October 1987). “Toxicity of light-exposed Hepes media”Journal of Immunological Methods103 (1): 145.
  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 Cellular & Developmental Biology21 (5): 282–7.

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