Table 722

Production of Carotenoids in Bioreactors

Carotenoid

Produced Organism

P-Carotene Blakeslea trispora

P-Carotene Phycomyces blakesleeanus

Astaxanthin Haematococcus lacustris

Astaxanthin Xantophyllomyces dendrorhous

Some Relevant Information

Surfactants have been suggested to improve the production of P-carotene The mutant cars produces 100 times more pigment than wild type (up to 5 mg/g dry mycelium) Illumination enhances the astaxanthin production; a two-stage process is proposed to produce higher carotenoid levels With the mutant JB2, an increment in the carotenoid production, up to 2.3 times, is obtained (1.54 |g astaxanthin/mg dry weight)

Source: Adapted from Delgado-Vargas et al. (2000).2

with chemical synthesis and new strategies are sought for better carotenoid production. A single plasmid with five essential genes has been constructed, from GGPP synthase to P-carotene hydroxylase under a strong promoter. Additionally, two aspects are considered to overproduce carotenoids; the first is the channeling into carotenoids and the second the availability of membranes for the storage of these lipidic compounds. Thus, carotenoid production in Escherichia coli was raised to 289 |g/g d.w. of zeaxanthin.146 In another study, E. coli was transformed with the DXP-synthase (DXPS) genes of Bacillus subtilus and Synechocystis sp.; transformed cells increased their lycopene and ubiquinone-8 levels. Thus, the generation of DXP by this pathway is enough to elevate the isoprenoid production. This technology transferred to plants could be used to improve crop flavor, fragrance, and color.147

It has been found that mixotrophic cultivation and suitable irradiation of Hae-matococcus pluvialis have resulted in a fairly good yield of astaxanthin (up to 40 mg/l; 43 mg/g cell d.w.) within a reasonable time, under laboratory conditions. However, to compete with synthetic astaxanthin, suitable scaling-up is required. In addition, large-scale production in open ponds has proved unsatisfactory because of severe contamination problems, with bacteria or protozoa; it has been proposed that a selective medium might overcome this difficulty but the problem has not been solved. It is also known that the most critical factors in carotenoid production are temperature and light; most authors have employed temperatures in the range of 20 to 25°C for astaxanthin formation. Light conditions range between 20 and 430 pmol quanta/m2/s and usually fluorescent light sources are used; higher irradiation levels yield higher carotenoid concentrations. Today, many authors have adopted a two-stage approach in the cultivation of H. pluvialis for carotenoid production. During the vegetative/green phase, low irradiances are utilized and much higher irradiation (ten times) is employed during aplanospore formation and astaxanthin biosynthesis.114

Other algae employed in P-carotene production are Dunaliella salina and Spirulina platensis. D. salina is cultivated in open ponds with highly saline brines, whereas S. platensis is successfully cultivated in highly alkaline (pH > 9.2) waters.114

The cultivation of Xantophyllomyces dendrorhous is quite simple but yields obtained by various strains are usually less than 500 ^g/g d.w. of cells. Overproducing strains have been obtained by mutagenesis and production of up to 5 mg/g has been reached. However, biomass production is low and strains have shown instability; thus, commercialization has not been possible.114 Interestingly, some strains require the use of two-stage cultures; X. dendrorhous is most efficient in pigment production if in the initial stage cell growth is favored with low C/N ratio levels, and in the second stage, where production takes place, a high C/N ratio is used (Table 7.22).

Lutein sales as feed additive in the United States are about U.S. $150 million/year; consequently new sources of lutein have been proposed. In particular, Mureillopsis spp. has been suggested as a lutein source; it produces up to 35 mg/l culture. It has been observed that low N2 and high photon flux density levels induce the production of lutein. In a bioreactor production of 150 mg/m2/day has been reached, a value comparable with the production of astaxanthin by H. pluvialis and P-carotene by D. salina.148

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