Rapid chromatographic determination of caseins in milk with photometric and fluorimetric detection using a hydrophobic monolithic column

Abstract

Reverse-phase liquid chromatographic methods using a hydrophobic C18 monolithic column and on-line photometric and fluorimetric detection for the determination of the major casein (CN) proteins in milk are presented. The separation of αs1-CN, αs2-CN, β-CN and κ-CN was achieved in only five minutes. Fluorimetric detection enabled better analytical results than photometric detection. Thus, the dynamic ranges of the calibration graphs and detection limits obtained using fluorimetric detection were (mgmL−1): αs1-CN (0.74–10.0, 0.22), αs2-CN (0.15–10.0, 0.045), β-CN (0.68–10.0, 0.20) and κ-CN (0.21–10.0, 0.06). The analytical features of the photometric method, which does not allow the quantification of β-casein, were (mgmL−1): αs1-CN (1.5–9.0, 0.45), αs2-CN (1.4–10.0, 0.43) and κ-CN (0.4–9.0, 0.12). Precision data, expressed as relative standard deviation, ranged between 0.6% and 5.3% for the fluorimetric method and between 2.4% and 6.2% for the photometric method. Both methods were applied to the analysis of three different milk samples, obtaining recoveries in the ranges of 86.6–103.2% and 92.0–106.5% using fluorimetric and photometric detection, respectively.