HEPES Buffer Preparation and Recipe

Details:

HEPES is a kind of biological buffer, and its development and production is still in its infancy. It has been widely used in biochemical products, clinical detection, in vitro diagnostic reagents, biological research, pharmaceutical and other fields. This paper introduces the production methods.

HEPES synthesis route

1. piperazine was reacted with ethylene oxide to form hydroxyethyl piperazine;

2. Sodium chloroethane was produced by 1,2-dichloroethane and sodium sulfite;

3. HEPES was produced by reaction of hydroxyethyl piperazine with sodium chloroacetate.

HEPES

HEPES synthesis steps

1. piperazine hexahydrate was dissolved in ethanol to form a transparent and colorless solution. Under the condition of 80 ℃ insulation, ethylene oxide was added while reflux was carried out. After the reaction, ethanol was removed by vacuum distillation, and the residue was separated and purified by vacuum separation. The distillate hydroxyethyl piperazine was collected at 125-127 degrees boiling range.

2. add anhydrous sodium sulfite and distilled water to the flask, heat it in a water bath outside, and add 1,2-dichloroethane at this temperature when the reaction liquid reaches 90 degrees, and then continue to return. Then, the reaction liquid is concentrated in 70 degree water bath until the white solid is completely separated. The solid and 95% alcohol are suspended, heated and returned, and the insoluble matter is filtered out while the heat is applied. The supernatant was placed at 0 degree, and then filtered with a burette funnel the next day to collect the white Flake sodium chloroacetate crystal.

3. add hydroxyethyl piperazine, sodium chloroethane sulfonate and water into the flask, and heat preservation and stirring reaction in 110 degree oil bath; Add sodium hydroxide to make pH 9-10, and return to the end of reaction. After cooling, the salt was purified by h+ column of ion exchange resin; The crude HEPES products were obtained by decompressure concentration of chromatography liquid, decolorization and filtration of activated carbon, concentration of clear liquid, pouring into boiling alcohol, adjusting pH with glacial acetic acid, and cooling by alcohol precipitation. Then, HEPES products can be obtained by recrystallization, cleaning and drying.

HEPES is easy to dissolve in water and difficult to dissolve in other solvents. It does not precipitate with polyvalent metal ions. The dissociation is rarely affected by its concentration, temperature, ion composition or salt content of the medium. Desheng is a manufacturer of biological buffer, which can provide high quality HEPES, Tris and other buffers.