02 Determination of Protein Concentration

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[Music] when dissolved every substance has the property to absorb part of light passing through the solution the phenomenon is called light absorption every chemical compound absorbs light at a strictly fixed wavelength in this way the respective substance can be identified a decimal logarithm of the ratio of the two intensities is called extinction

absorbance or optical density of the solution the extinction is directly proportional to the concentration of the substance in the sample this linear dependence is called beer-lambert law and it is used to determine substance concentrations the device for determining protein concentration is called spectrophotometer the sample is in a cuvette a beam of light at a fixed

wavelength for the specific substance passes through the qubit the spectrophotometer directly measures the absorbance at a given concentration light at a different wavelength is used to determine protein concentration proteins containing aromatic amino acids absorb light at a wavelength of 218 nanometers and their concentration can be measured directly in the ultraviolet part of the spectrum

the concentration of coloured proteins as haemoglobin myoglobin plastocyanin cytochromes flavo proteins etc is determined at different wavelengths in the visible part of the spectrum concentration of colorless proteins can be measured spectrophotometric allele after carrying out a colored reaction as the biruté assay sodium hydroxide is added to a protein solution then a drop

of copper sulfate is added to this alkaline solution in the presence of peptide bonds a violet color it seems which proves that the solution contains proteins the biruté reaction is a qualitative reaction proving the presence of Katies the intensity of the violet colors depends on the protein concentration in the sample this solution can be studied photo metrically