Determination of collagen content in recombinant human collagen dressing by high performance liquid chromatography

Objective To establish an HPLC method for the determination of type Ⅲ collagen in dressings for quality control of the product.

Methods A Agilent Eclipse XDB-C18 column(4.6×150 nm, 5 μm)was used with 0.1% TFA solution and 0.1% TFA acetonitrile solution as mobile phase at 30 ℃, the flow rate was 1.0 mL·min⁻¹, the detection wavelength was 214 nm.

Results The method was sensitive, after hydrolysis,for dressings with recombinant type Ⅲ humanized collagen content greater than 0.1mg/mL, The linear range was 0.1−1.2 mg/mL （R²=0.9 994）for type Ⅲ collagen, the average recovery was 90.91%~92.92%. The LOD and LOQ was 0.05 mg/mL and 0.1 mg/mL. After enzymatic hydrolysis, The linear range was 0.05~1.0 mg/mL （R²=0.9995）for type Ⅲ collagen, Can be used to guide the detection of low concentration dressing proteins.

Conclusion The method is rapid, accurate and reproducible, and can be used to determine the content of type Ⅲ collagen in dressings.