Characterization of astaxanthin esters in Haematococcus pluvialis by liquid chromatography–atmospheric pressure chemical ionization mass spectrometry

Abstract

After first being analyzed by HPLC, 4 free carotenoids, 15 astaxanthin monoesters, 12 astaxanthin diesters, and 3 astacin monoesters in Haematococcus pluvialis were identified by liquid chromatography–atmospheric pressure chemical ionization mass spectrometry (LC–(APCI)MS). Identification of each compound was based on the characteristic fragment ions of the positive ion mode, negative ion mode, and MS 2. Astaxanthin esters were identified based on the loss of one or two fatty acids. In a positive ion mode, astaxanthin monoesters had characteristic fragment ions at m/ z 597 [M+H−fatty acid] + and m/ z 579 and 561 that resulted from a continuous loss of water. The relative intensity of m/ z 579 in MS 2 amounted to more than 80% of that of the molecular ion. In astaxanthin diesters, the intensity of m/ z 561 occasionally was equal to that of m/ z 579, but in general the former, amounting to 50 to 60% or more of the molecular ion, was stronger than the latter, which decreased to 20 to 30% of the molecular ion. In addition, a set of compounds with maximum absorbance at 400 nm, detected by high-performance liquid chromatography–diode array detector (HPLC–DAD), had strong characteristic fragment ions at m/ z 871 and 593 in the positive ion mode MS 2. They were presumed to be linolenic acid or an isomer of ω-6-γ-linolenic acid esters of astacin.