Comparative analysis of astaxanthin and its esters in the mutant E1 of Haematococcus pluvialis and other green algae by HPLC with a C30 column

Abstract

A gradient reversed-phase high-performance liquid chromatography (HPLC) method using a C30 column was developed for the simultaneous determination of astaxanthin, astaxanthin monoesters and astaxanthin diesters in the green algae Chlorococcum sp., Chlorella zofingiensis, Haematococcus pluvialis and the mutant E1, which was obtained from the mutagenesis of H. pluvialis by exposure to UV-irradiation and ethyl methanesulphonate (EMS) with subsequent screening using nicotine. The results showed that the contents of total astaxanthins including free astaxanthin and astaxanthin esters ranged from 1.4 to 30.9 mg/g dry biomass in these green algae. The lower total astaxanthin levels (< 2 mg/g dry biomass) were detected in the green algae Chlorococcum sp. and C. zofingiensis. The higher total astaxanthin levels (>16 mg/g dry biomass) were found in the green alga H. pluvialis and its mutant E1. It is notable that the mutant E1 is found to have considerably higher amounts of total astaxanthin (30.9 mg/g) as compared to the wild strain of H. pluvialis (16.1 mg/g). This indicates that UV-irradiation and EMS compound mutagenesis with subsequent screening using nicotine is an effective method for breeding of a high-producing astaxanthin strain of H. pluvialis. In addition, the green alga C. zofingiensis had a remarkably higher percentage of astaxanthin diesters (76.3% of total astaxanthins) and a remarkably lower percentage of astaxanthin monoesters (18.0% of total astaxanthins) in comparison with H. pluvialis (35.5% for diesters and 60.9% for monoesters), the mutant E1 (49.1% and 48.1%) and Chlorococcum sp. (18.0% and 58.6%).

Potential health‐promoting effects of astaxanthin: A high‐value carotenoid mostly from microalgae

“…Therefore, astaxanthin in nature is either conjugated with proteins or esterified with one or two fatty acids to form monoester and diester forms [8,11]. In H. pluvialis, astaxanthins exist mainly as various astaxanthin esters formed by combining various fatty acids with different isomers of astaxanthin [6].…”

“…Peng et al [11] showed that the green microalga C. zofingiensis had a remarkably higher percentage of astaxanthin diesters in comparison with H. pluvialis with a higher percentage of astaxanthin monoesters. The esters of astaxanthin and adonixanthin, and free canthaxanthin were the major carotenoids in the alga Chlorococcum cells [3].…”

Preparation of (3R, 3′R)-astaxanthin monoester and (3R, 3′R)-astaxanthin from Antarctic krill (Euphausia superba Dana)

“…Other methods are based on conjecture. For example, in 2008 Peng and Xiang [15] analyzed the AM from green algae by HPLC using β-carotene as a standard. However, the inference was solely based on the higher polarity of AM than that of β-carotene.…”

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“…Therefore, astaxanthin in nature is either conjugated with proteins or esterified with one or two fatty acids to form monoester and diester forms, producing an array of colors in different organisms 14,16 . It is the chromophore in the blue, green, and yellow pigments of lobsters.…”

“…In the astaxanthin molecule,each double bond from the polyene chain may exist in two configurations as geometric isomers cis or trans 13,15,16 . Most carotenoids found in nature are predominantly all trans-isomers 12,13 .…”