Sample mixtures

Various astaxanthin-containing samples were deesterified individually and merged to examine cross-interactions. The individual samples contained either 240 μg of H. pluvialis powder (Golden Peanut), 16 μg of astaxanthin monopalmitate, or 12 μg of free astaxanthin. Maximum 4.5% w/w of the H. pluvialis powder were considered to be free astaxanthin by measurements. Astaxanthin monopalmitate consists of about 69.4% w/w of free astaxanthin. Thus, the final extracts of the samples contained 4.2 to 4.7 μg/mL of free astaxanthin. Moreover, three samples were prepared, two containing mixtures of two of the single solutions, and one sample containing all three. In these, the individual samples were represented at identical amounts as in the samples containing only one of the analytes.

After enzymolysis and recovery in petroleum ether, three aliquots of 600 μL were taken from each sample. They were prepared either with 63 μL pure acetone, 31.5 μL pure acetone and 31.5 μL of astaxanthin standard solution containing 3.6 μg free astaxanthin, or with 63 μL of astaxanthin standard solution containing 7.2 μg free astaxanthin. Furthermore, 600 μL pure petroleum ether were enriched with acetone and astaxanthin like the samples. The experiment was performed once.