Determination of Collagen Protein Content in Liquid Phase

Collagen is an important protein that is widely present in various human tissues, such as skin, bones, muscle tendons, and ligaments. Accurate determination of its concentration is crucial for multiple fields such as biomaterial research, food industry, and pharmaceutical research.

 

Liquid Chromatography (LC) is an efficient and precise analytical technique for collagen concentration measurement, especially High-Performance Liquid Chromatography (HPLC) and Ultra-High Performance Liquid Chromatography (UHPLC) techniques. These techniques can separate, identify, and quantify collagen and its hydrolysis products in the samples. 

 

Common methods for collagen content determination include:

 

1. Sample Preparation

Collagen is extracted from the sample and preprocessed by centrifugation, filtration, or using Solid Phase Extraction (SPE) to remove impurities and purify the collagen.

 

2. Enzymatic Hydrolysis

Proteolytic enzymes are used to further break down collagen into peptides or amino acids, which helps to improve its resolution and detection sensitivity in LC.

 

3. Chromatographic Analysis

Select appropriate chromatographic columns (such as reverse phase columns) and mobile phases (such as water/acetonitrile or water/methanol gradient) to achieve effective separation of collagen or its hydrolysis products. Common detectors include Ultraviolet (UV) detectors, Fluorescence detectors, and Mass Spectrometry (MS) detectors. MS detectors are particularly suitable for the identification and quantification of specific peptides in complex samples.

 

4. Quantitative Analysis

By preparing a standard curve using standard substances of known concentration (such as specific collagen peptides or amino acids), accurate quantification of collagen content in the sample can be achieved. The content of collagen in the sample is calculated quantitatively by comparing the peak area or peak height of the peptide or amino acid with the standard curve.

 

The advantage of LC in determining collagen content is its high separation efficiency, high sensitivity, and good repeatability, making it an important tool for analyzing collagen and its hydrolysis products. The method is particularly suitable for the precise measurement of collagen content in complex biological samples, which is of great importance for quality control, functional evaluation, and research of collagen.