KR20030076133A - Developmentof culture medium of phytoplanktons (microalgae) - Google Patents

Abstract

PURPOSE: A method for development of a medium for culturing phytoplankton(microalgae) is provided, thereby effectively increasing the production yield of the phytoplankton(microalgae). CONSTITUTION: A method for development of a medium KEP for culturing phytoplankton(microalgae) comprises adding bacteria mineral water into a conventional medium for phytoplankton, wherein the conventional medium for phytoplankton is selected from BBM 10%, f/2, Chu 10 and GB11; the amount of bacteria mineral water added is 3%; the bacteria mineral water is prepared by fermenting a mixture of porcine urine and minerals of rock pellet using aerobic bacteria in a fermenter with aeration.

Description

Development of culture medium of phytoplankton (microalgae) {Developmentof culture medium of phytoplanktons (microalgae)}

Unlike terrestrial higher plants, which are transformed only by genetic recombination technology, phytoplankton is sensitive to physiological and biochemical metabolism depending on growth environment and culture conditions. It is a microalga with excellent adaptability. Microalgae have high productivity in water systems (freshwater, seawater), and are used as a source of pollutants (nutrient salts and heavy metals) in wastewater, as well as raw materials for fish feed, health and alternative foods, food additives, pharmaceuticals and cosmetics. It is used. In particular, the green algae Scendesmus spp. Used in the present invention is a universal species that inhabits anywhere in the world, and is one of the most predominant species in freshwater. Conventionally, a continuous culture method of continuously injecting nutrients (minerals and vitamins, etc.) is used to mass-culture these phytoplankton. This method is inexpensive due to the hassle of continuously adding the culture medium and the cost of maintaining the continuous culture equipment because phytoplankton intake of nutrients in the culture medium intensifies the depletion of nutrient sources over time. Not only fall, but also physiological and biochemical activity of phytoplankton is lowered to improve the adaptability to the newly introduced culture medium has the disadvantage of low industrial application efficiency.

To maximize the added value of phytoplankton for a long period of time (at least 1 month) without the need for continuous nutrient infusion by using KEP culture to compensate for the shortcomings of the continuous culture and to increase the industrial efficiency of phytoplankton. do. By using KEP culture medium than when using conventional commercial culture medium (BBM 10%, f / 2, Chu 10, BG11, etc.), phytoplankton does not occur and the general pigmentation phenomenon that appears as a depletion of minerals in subcultures does not occur. Cell division of cells occurs, the physiological activity showing the photosynthetic efficiency of photosynthetic photosystem II is extremely high, and the aging of cells, which are stagnant in the cell growth cycle, is prolonged. Secondary biochemical metabolites (constituent amino acids) such as chlorophyll a , carotenoids (α & β-carotene, astaxanthin, lutein) and essential amino acids are also synthesized up to 300% higher than phytoplankton grown in commercial cultures. Fatty acids were synthesized lower than phytoplankton grown in commercial culture. This result is of high industrial added value because phytoplankton can be used as an important biological resource that can be developed into medicines such as anticancer and immune enhancing agents, health supplements, cosmetics, beverage additives, natural seasonings and feed.

Figure 1 shows the change in cell growth rate of phytoplankton ( Scendesmus spp.) Cultured in three kinds of cultures-Control: commercial culture (BBM10%); KEP: culture medium containing 3% of fermented pork urine (biologically active water) in a commercial culture solution; WW: A culture solution containing 3% of normal porcine urine that has not been fermented to the commercial culture solution.

Figure 2 shows the photosynthetic efficiency of photosystem II, in which phytoplankton ( Scedesmus spp.) Cultured in three different cultures are represented by chlorophyllometer (Phyto-PAM).

3 shows changes in ATP (adenosine triphosphate) concentrations of phytoplankton ( Scendesmus spp.) To be cultured in three different cultures.

4 shows the diversity of chlorophyll concentrations of phytoplankton ( scenedesmus spp.) Cultured in three different cultures.

Figure 5 shows the types and concentrations of carotenoids of phytoplankton ( scenedesmus spp.) Cultured in three different cultures (As: astaxanthin, astaxanthin; Lu: lutein, lutein; Al-crt: α-carotene, carotene; Bt -crt: Bt-crt: β-carotene; TC: total carotenoid).

6 is a change in the concentration of fatty acids of phytoplankton ( scenedesmus spp.) Cultured in three kinds of culture medium.

7 shows phytoplankton (left: Scenedesmus. Quadricauda ; right: S. spinosus ) -chromosomes cultured in KEP culture for 51 days with very good chromosomes.

8 shows phytoplankton (left: cenedesmus spinosus ; S. acutus ) -chromosomes incubated in control cultures for 51 days, with no apparent nuclear appearance.

9 shows phytoplankton pigment changes according to different cultures for 34 days (No. 1-Control culture; No. 2-KEP culture: darker green than other 2 cultures, which can be visually identified as chlorophyll synthesis is much higher). No. 3 -WW Culture Solution)

Bioactivity of 0.05%, 1%, 2%, 3%, 5%, 7%, 9% in commercial culture (BBM 10%; pH = 6.8) (Stein 1973) (Table 1), respectively, for the development of KEP cultures Bacteria Mineral Water (Reference: Hiroshi N. 1998. Bacteria Saves the Earth-Challenges in BMW Technology. Blue Peace, 216 pp.) The same growth conditions (light cycle: 14h-m / 10h-) Cancer; brightness: 115 μmol m ⁻¹ s ⁻¹ ; temperature: 24 ° C.), Scenedesmus spp. (Phytoplankton mixed with Scenedesmus acutus, S. spinosus, and S. quadricauda ) After about two months of incubation, the cells of Scendesmus spp. Grown in KEP culture solution (pH = 6.6) with 3% bioactive water Growth occurs most vigorously (FIG. 1). Biologically active water is recycled livestock wastewater obtained by fermenting pig urine with rock pellet minerals and aerobic bacteria by aeration tank. The most optimal concentration is 3% bioactive water added to commercial culture. . In order to compare the effect of this KEP culture solution, the culture of Scenedesmus spp. Was attempted by simultaneously using a control medium, which is a commercial medium, and a culture medium (WW: Waste Water) to which non-fermented general livestock wastewater was added. As shown, the growth period growth rate (0.12) of the KEP culture solution is the highest, and three times higher than the growth period growth rate (0.4) of the control culture.

In order to prove whether the cells of Scenedesmus spp. Grown in KEP culture medium dominate physiologically and biochemically, the existing analytical methods are used as follows. A chlorophyllometer (Phyto-PAM: Walz) was used to measure photosynthetic efficiency, and the concentration of ATP (adenosine triphosphate) representing bioenergy was measured using a luminometer (TD-2O / 2O; Turner Designs, 2000). Chlorophyll concentration was measured using the conventional method (De Loura et al ., 1987), and carotenoids were analyzed by Kim and Lee (1998) using HPLC (SPD-M6A; Simadu). do. Fatty acid analysis is performed by a gas chromatograph instrument (HP 6890 Series) by Kim et al. , 1996.

The cells grown in commercial culture and WW culture of Scendesmus spp. Start aging after 21 days, but Scendesmus spp. Cultured in KEP culture starts aging at 41 (Fig. 1). Figure 2 shows that the photosynthetic efficiency of Scendesmus spp. Grown in KEP culture solution is much higher over the incubation period (FIG. 2), and the concentration of ATP is highest after 51 days of incubation (FIG. 3). After 31 days of culture, the chlorophyll α concentration and total chlorophyll of Scenedesmus spp. Grown in KEP culture were the highest (FIG. 4). Particularly, the color of Scenedesmus spp., Which grew in KEP culture from 31st day of culture, had high chlorophyll accumulation and dark green color, whereas the cells of Scenedesmus spp. Grown in commercial culture medium and WW culture medium had chlorophyll decomposed and the whole sample turned yellow. The phenomenon of discoloration appears.

Chlorophyll and α, β-carotene, astaxanthin, lutein and carotenoids, which are currently studied as antioxidants and anticancer agents, also have the highest Scenedesmus spp. Grown in KEP culture (Fig. 5). Fatty acid constituting one lipid shows the lowest concentration of Scenedesmus spp. Grown in KEP culture (FIG. 6). Also spp Scenedesmus grown in KEP culture the total concentration (336 mg / L) and concentration (119 mg / L) of the essential amino acids in the amino acid composition of the protein is Scenedesmus spp grown in a commercial culture medium (amino acids the total concentration:.. 124 mg Essential amino acid: 44 mg / L), about 300% higher (Tables 2 and 3).

<References>

De Loura IC, Dubacq JP, Thomas JC (1987) Plant Physiol . 83: 838--843.

Kim MK, Dubacq JP, Thomas JC, Giraud G (1996) Phytochemistry 43: 49-55.

Kim MK, Lee HW (1998) Algae 13: 151--155.

Stein J (1973) Handbook of Phycological Methods. Culture Methods and Growth Measurements, 448 pp. Cambridge: Cambridge University Press.

Turner Designs (2000) A TD-20 / 20 luminometer method for ATP measurements.

Turner Designs Biotechnology pp.1--4.

The KEP culture solution can be applied to all phytoplankton as well as Scenedesmus spp., And it may have a synergistic effect by varying the growth conditions (light quantity, photoperiod, mineral quality, temperature, growth hormone, etc.) at the time of culture. In addition, synergistic effects can be created by developing a variety of KEP modified KEP culture medium. In addition to the above analytes, biochemical analysis of bioactive substances and immunomodulators (bioassay) to reveal the effects of KEP culture medium as important biological resources that can be developed as anti-cancer effects, health supplements, cosmetics, beverage additives, natural seasonings and feed. Available.

Claims (1)

Development of KEP culture solution and KEP modified culture solution by adding 3% of fermented broth (bioactive water) to commercial cultures (BBM 10%; CHU-10; f2, etc.). When used for subculture, it is more economical than continuous culture because it does not need to add continuous culture like continuous culture. More physiological and biochemical efficiency results (about 300%). Development of phytoplankton grown in KEP culture solution developed in the present invention as medicines, dietary supplements, cosmetics, beverages and food additives, natural seasonings and feed. TABLE 1 Table 1 lists the composition of BBM (Bold's Basal Medium) 10% (Ref. Stein J. 1973. Handbook of Phycological Methods.Cultural Methods and Growth Measurements, 448 pp. Cambridge: Cambridge University Press) TABLE 2 Table 2 shows the amino acid concentrations of phytoplankton ( scenedesmus spp.) Cultured in three different cultures (Asp: aspartic acid; Thr: threonine; Ser: serine; Glu: glutamic acid; Pro: proline; Gly: glycine; Ala Alanine; Cys: cystine; Val: valine; Met: methionine; Ile: isoleucine: Leu: leucine; Tyr: tyrosine; Phe: phenylalanine; His: histidine; Lys: lysine; Arg: arginine) TABLE 3 Table 3 shows the essential amino acid concentrations of phytoplankton ( scenedesmus spp.) Cultured in three different cultures.