Lewis M. Brown
Solar Energy Research Institute
Mordhay Avron of the Weizmann Institute, Rehovot, Israel reported on the biotechnology of ß-carotene production in the unicellular alga Dunaliella bardawil. The production of this pigment was enhanced at high light intensity. Basic research on this organism has led to a successful industrial application. A new industry has emerged which produces and markets ß-carotene for its anti-cancer activity. A demand for ß-carotene from natural sources such as Dunaliella is supported by the research which indicates that the cisisomers from natural sources such as Dunaliella are more effective than the all-trans form as anti-cancer agents.
Lewis M. Brown of SERI (Golden, Colorado) reported that there are many other potential high value products that could be produced from microalgae in addition to ß-carotene. The biosynthesis of some of these may be enhanced by concentrated solar photons. It was also pointed out that concentrated solar photons have potential in the development of advanced photobioreactors that take advantage of flashing light or other considerations to increase product yield. The overall conclusion of the session was that this is a relatively unexplored area that would benefit from an infusion of research funds for development.
THE BIOTECHNOLOGY OF CULTIVATING DUNALIELLA RICH IN BETA CAROTENE: FROM BASIC RESEARCH TO INDUSTRIAL PRODUCTION
Wiezmann Institute of Science
The unicellular alga Dunaliella exists in several ill-defined species. In hypersaline lakes, the Dunaliella strain which predominates is often red, rather than green, in color, due to massive accumulation of a single pigment, beta-carotene. Of the many strains of the genus Dunaliella described, only two strains, Dunaliella bardawil and Dunaliella salina Teod., have been shown to possess the capacity to produce large amounts of beta-carotene, when cultivated under appropriate conditions. Electron micrographs of Dunaliella bardawil indicate that the massive amounts of beta-carotene are located in a large number of chloroplastic, lipoidal globules located in the interthylakoid space of the chloroplast. Under appropriate cultivation, more than 10% of the dry weight of D. bardawil is accounted for by beta-carotene.
The rate and extent of beta-carotene accumulation in D. bardawil is determined by the conditions under which it is cultivated. Under standard laboratory cultivation conditions, little beta-carotene is synthesized and the algae appear green in color. However, when the light intensity is increased much beyond the intensity required for normal growth, and when the rate of growth is limited, beta-carotene is accumulated to the highest levels.
Beta-Carotene accumulation protects against the deleterious effects of high-intensity irradiation. Thus, strains which do not accumulate beta-carotene, or beta-carotene-poor D. bardawil die when exposed to high irradiation under limiting growth conditions, while beta-carotene-rich D. bardawil survive under the same conditions. This may explain the previously mentioned predominance of D. bardawil or D. salina Teod. over green strains of Dunaliella in some saline lakes in nature, where low concentrations of algae are exposed to high solar irradiation under nutrient-limiting conditions.
The ability of selected strains of Dunaliella to produce beta-carotene by photosynthetic conversion of CO2 stimulated research aimed at probing the possible industrial production of this commercially valuable product. Since these algae grow at high salinity in a complete inorganic medium, few, if any, natural competitors and predators interfere with large-scale cultivation. Furthermore, the lack of a cell wall makes this alga, in contrast with most, easily digestible by most animal species.
Indeed, large-scale cultivation of Dunaliella is carried out today in Israel, in the United States, and in Australia for the commercial production of an algal meal rich in beta-carotene, purified beta-carotene, and other natural products.
Beta-carotene has been used both as a food coloring agent and as a source of vitamin A in animal feed. Adding dry Dunaliella or an extract of the algae high in beta-carotene to a vitamin A deficient chick diet indeed showed that it is an excellent source of the vitamin and, in addition, a yolk-color enhancing agent.
Recent epidemiological and oncological studies suggest that normal to high levels of beta-carotene in the body may protect against cancer development in humans. The studies provide evidence that people with above average beta-carotene intakes have a lower incidence of several types of cancer[5–7].
Natural beta-carotene, as found in D. bardawil and in many fruits and vegetables, contains about 50% of all-trans beta-carotene, with the rest
composed of a few other beta-carotene isomers[8,9]. It has not been resolved at present whether the beneficial effects of beta-carotene in the diet are correlated with this variability. However, it is clear that the beta-carotene isomer mixture, as present in Dunaliella , accumulated to a much larger extent in the liver of chicks and rats than the synthetic 99% all-trans beta-carotene, which presently accounts for most of the commercially available product.
Intensive cultivation of Dunaliella may be the first commercially successful example of the use of selected algae for the biological conversion of solar energy into products of commercial interest. This approach has so far been hampered by the economic difficulties inherent in the production of a relatively inexpensive product (i.e., feed) and by the required, relatively sophisticated technology. The high-priced product, beta-carotene, accounts for the renewed commercial interest in algal cultivation. After extraction of the beta-carotene from the concentrated Dunaliella, there remains a glycerol and protein-rich algae meal, which was shown to serve as an excellent source of feed for fish, fowl, and rats. We thus have in the Dunaliella cultivation system the aquaculturist's and ecologist's ideal facility: a production unit which removes CO2 from the air; produces oxygen, useful chemical products, and feed; and leaves no polluting residue requiring disposal.
1. Ben-Amotz, A., and M. Avron. 1981. Glycerol and Beta-Carotene Metabolism in the Halotolerant Alga Dunaliella. A Model System for Biosolar Energy Conversion. Trends in Biochem. Sci. 6:297–299.
2. Ben-Amotz, A., A. Shaish, and M. Avron. 1989a. The Mode of Action of the Massively Accumulated Beta-Carotene D. bardawil in Protecting the Alga Against Damage by Excess Irradiation. Plant Physiol. 91:1040–1043.
3. Ben-Amotz, A., and M. Avron. 1990. The Biotechnology of Cultivating the Halotolerant Algae Dunaliella for Industrial Products. Trends in Biotechnology 8:121–126.
4. Ben-Amotz, A., S. Edelstein, and M. Avron. 1986. Use of the Beta-Carotene Rich Alga Dunaliella bardawil as a Source of Retinol. British Poultry Science 27:613–619.
5. Peto, R., R. Doll, J.D. Buckley, and M.B. Sport. 1981. Can Dietary Beta-Carotene Materially Reduce Human Cancer Rates? Nature 290:201–207.
6. Menkes, M.S., G.W. Comstock, J. Vuilleumier, K.J. Helsing, A.A. Rider, and R. Brookmeyer. 1986. Serum Beta-Carotene, Vitamin A and E, Selenium and the Risk of Lung Cancer. New Engl. J. Med. 315:1250–1254.
7. Ziegler, R.G. 1989. A Review of Epidemiological Evidence that Carotenoids Reduce the Risk of Cancer. J. Nutr. 199:116–122.
8. Ben-Amotz, A., A. Katz, and M. Avron. 1982. Accumulation of Beta-Carotene in Halotolerant Algae: Purification and Characterization of Beta-Carotene-Rich Globules from Dunaliella bardawil (Chlorophyceae). J. Phycol. 18:529–537.
9. Ben-Amotz, A., A. Lers, and M. Avron. 1988. Stereoisomers of Beta-Carotene and Phytoene in the Alga D. bardawil. Plant Physiol. 86:1286–1291.
10. Ben-Amotz, A., S. Mokady, S. Edelstein, and M. Avron. 1989b. Bioavailability of a Natural Isomer-Mixture as Compared with Synthetic All-Trans Beta-Carotene in Rats and Chicks. J. Nutr. 119:1013–1019.
PRODUCTION POTENTIAL OF BIOCHEMICALS FROM ALGAE AND OTHER BIOTECHNOLOGICAL INNOVATIONS ENABLED BY HIGHER SOLAR CONCENTRATION
Lewis M. Brown
Biotechnology Research Branch
Solar Energy Research Institute
The approach of using multiple-sun light intensity to increase the yield of various products from autotrophically grown algae is relatively unexplored. There are some possibilities for advanced photobioreactor design that might result in increased culture density and hence lower culture volume. However, culture productivity and product yield (typically) do not increase linearly with light intensity, but rather saturate at fairly low light intensities for most algae in the range of 15–20% of full sun (300–500 µmol quanta m-2 s-1) depending on species and on growth irradiance. Instantaneous measurements of photo-synthesis indicate that algae grown under higher light intensities are saturated at higher light intensities and have higher photosynthetic rates than algae grown under lower light intensities. There is no obvious advantage to growing algae at full sun let alone multiple suns because no increase in productivity is typically realized. However, it may be possible to grow algae in more concentrated culture at multiple-sun irradiances if smaller culture volumes are desired. Such culture conditions may be economical for high-value products (e.g. pharmaceuticals, pigments), but are unlikely to be economical for low-to mid-value products (fuel and commodity chemicals). Another problem with utilization of concentrated solar photon irradiances is photoinhibition. The photosynthetic process is severely inhibited at full-sun or multiple-sun irradiance. While there are probably multiple sites affected by high light intensity, it is thought that a primary site for photoinhibition is at the QB protein of photosystem II. Thus, there is a limit to the amount of light that algae can be exposed to without cellular damage taking place. Some workers have proposed that higher rates of photosynthesis can be achieved in flashing light compared to continuous light. Such effects are more clearly seen at > 10 Hz, and similar phenomena are sometimes claimed for lower rates of flashing, although such effects are by no means well accepted. Also, there has been no practical application of these results especially with concentrated solar photon sources. Much of the work has been done outside of the United States, and even the research that has been performed in the United States has been in the basic science (biochemistry) of photoinhibition. Application of concentrated solar photon technology for algal biotechnology is unlikely to take place in the United States as there is no support for such work. Leadership in this area is with Japan with its strong research programs in photobioreactor design, application of concentrated solar photon intensities, and search for high-value bioactive products from algae. Efforts in Israel have been an early demonstration of the potential of this approach.