Analysis of Peptides & Proteins / Spectro

Applications for Spectro

 
 
 

Analysis of Amino Acids by KBr Tablet Method

In the transmission method, light is irradiated on a measurement sample, and the light that passes through the sample is detected. However, the adequate technique (accessory device) differs depending on the sample shape. Among techniques used with the transmission method, the KBr tablet method is mainly used to measure solids and powders. The KBr tablet method is a necessary and indispensable procedure which has been standardized not only in the Japanese Pharmacopoeia (JP), but also in the United States Pharmacopeia (USP), standards of the American Society for Testing and Materials (ASTM). Special care is necessary in order to obtain a satisfactory spectrum by the KBr tablet method, including securing a uniform powder size and maintaining a constant pressure during tablet forming (pelletizing). In particular, the effect of moisture requires special attention. In this experiment, we conducted an analysis of amino acids, which easily absorb moisture, by the KBr tablet method. 

 
 
 
 
 
 
 
 

Protein Analysis Using FTIR - Secondary Structure Analysis of Bovine Serum Albumin Using Curve Fitting 

Proteins have local three-dimensional structures that are called secondary structures such as α- helices, β-sheets, β-turns, and random coils. These structures are created by the hydrogen bonding of C=O groups and N-H groups of peptide bonds within or in between polypeptide chains. The infrared absorption related to such secondary structures appears as a single broad peak at about 1650 cm-1 due to the overlapping of multiple absorption bands. This broad peak is referred to as the "amide I band" and is caused by the stretching vibrations of C=O groups in peptide bonds. By analyzing the amide I band, we can obtain information about the secondary structure of proteins. A method for determining the individual peaks of overlapping absorption bands is curve fitting (peak splitting). Curve fitting expresses the waveform of each absorption band as a calculated spectrum using approximate curves such as Lorentzian or Gaussian curves and optimizes the peak information (position, intensity, and full width at half maximum) of the approximate curves to minimize the difference between the actually measured spectrum and the calculated spectrum.

 
 
 
 
 
 
 
 
 
 
 
 
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