Estimation of protein by lowry method referene
Answers
The method combines the reactions of copper ions with the peptide bonds under alkaline conditions (the Biuret test) with the oxidation of aromatic protein residues. The Lowry method is based on the reaction of Cu+, produced by the oxidation of peptide bonds, with Folin–Ciocalteu reagent (a mixture of phosphotungstic acid and phosphomolybdic acid in the Folin–Ciocalteu reaction). The reaction mechanism is not well understood, but involves reduction of the Folin–Ciocalteu reagent and oxidation of aromatic residues (mainly tryptophan, also tyrosine). Experiments have shown that cysteine is also reactive to the reagent. Therefore, cysteine residues in protein probably also contribute to the absorbance seen in the Lowry assay.[4] The result of this reaction is an intense blue molecule known as heteropolymolybdenum Blue.[5] The concentration of the reduced Folin reagent (heteropolymolybdenum Blue) is measured by absorbance at 660 nm.[6] As a result, the total concentration of protein in the sample can be deduced from the concentration of tryptophan and tyrosine residues that reduce the Folin–Ciocalteu reagent.
The method combines the reactions of copper ions with the peptide bonds under alkaline conditions (the Biuret test) with the oxidation of aromatic protein residues. The Lowry method is based on the reaction of Cu+, produced by the oxidation of peptide bonds, with Folin–Ciocalteu reagent (a mixture of phosphotungstic acid and phosphomolybdic acid in the Folin–Ciocalteu reaction). The reaction mechanism is not well understood, but involves reduction of the Folin–Ciocalteu reagent and oxidation of aromatic residues (mainly tryptophan, also tyrosine). Experiments have shown that cysteine is also reactive to the reagent. Therefore, cysteine residues in protein probably also contribute to the absorbance seen in the Lowry assay.[4] The result of this reaction is an intense blue molecule known as heteropolymolybdenum Blue.[5] The concentration of the reduced Folin reagent (heteropolymolybdenum Blue) is measured by absorbance at 660 nm.[6] As a result, the total concentration of protein in the sample can be deduced from the concentration of tryptophan and tyrosine residues that reduce the Folin–Ciocalteu reagent.