Method for improving algae metabolism and lutein synthesis

Abstract

The invention provides a method for improving algae metabolism and lutein synthesis, and relates to the technical field of bioengineering, and the method comprises the following steps: selecting microalgae seeds; inoculating the microalgae seeds into a conical flask of a Basal culture medium; culturing the conical flask under a low-light condition for a predicted number of days; repeating the low-light culture process to obtain a seed solution; and inoculating the obtained seed solution into a conical flask of a Basal culture medium, and continuously culturing to obtain the microalgae. According to the method provided by the invention, growth and lutein accumulation in the process of heterotrophic culture or fed-batch heterotrophic fermentation culture and the like can be effectively promoted after the selected seeds are subjected to low-light pre-culture, so that algae metabolism is accelerated, and the yield of lutein is increased.

Description

Technical Field

[0001] The invention relates to the technical field of bioengineering, and in particular to a method for improving algae metabolism and lutein synthesis. Background Art

[0002] As an ancient photosynthetic organism, microalgae can accumulate high value-added products such as lipids, proteins and pigments. These nutritional products are widely used in food, health care, health and cosmetics industries. Lutein is a primary carotenoid that plays a vital role as an auxiliary component of light absorption and non-photochemical quenching in the microalgae photosystem. Its natural color and effective eye protection properties make it widely used in feed additives and health products. Microalgae are considered to be a very promising source of lutein because they grow very fast, have a high lutein yield, are highly adaptable to various environments, and are not restricted by seasons, compared to the only commercial natural lutein source, marigold petals.

[0003] In the current cultivation of microalgae, although the photoautotrophic mode is suitable for lutein accumulation and has low cost, the low biomass accumulation is still the main factor hindering its high lutein production. The heterotrophic mode can achieve high-density cultivation of microalgae, and the maximum biomass of some algae species can reach 200g / L, which is dozens to hundreds of times that of autotrophic. However, due to the lack of light, the photosynthetic activity is low, and the lutein content is significantly lower than that of the autotrophic mode. Summary of the invention

[0004] In view of the shortcomings of the prior art described above, the object of the present invention is to provide a method for improving algae metabolism and lutein synthesis, which is used to solve the problem in the prior art that during the cultivation of microalgae, due to lack of light, the photosynthetic activity is low and the lutein content is significantly lower than that in the autotrophic mode.

[0005] To achieve the above-mentioned object and other related objects, the present invention provides a method for improving algae metabolism and lutein synthesis, comprising the following steps: selecting microalgae seeds; inoculating the microalgae seeds into a conical flask of Basal culture medium; culturing the conical flask for a predetermined number of days under low light conditions; repeating the low light culture process to obtain a seed solution; inoculating the obtained seed solution into a conical flask of Basal culture medium, and continuing to culture to obtain microalgae.

[0006] In one embodiment of the present invention, the selected microalgae seeds are C. protothecoides CS-41.

[0007] In one embodiment of the present invention, the microalgae seeds are inoculated into a conical flask of Basal culture medium, comprising: inoculating the microalgae seeds at 10% (v / v) into a 500 mL conical flask containing 200 mL of fresh modified Basal culture medium, and placing the flask in a shaking fermentation culture at 28° C. and 180 rpm.

[0008] In one embodiment of the present invention, the low light condition is 20-40 μmol / m 2 / s, the light quality of the low light is white light, and the estimated number of days for cultivation is 2 to 3 days.

[0009] In one embodiment of the present invention, the obtained seed liquid is inoculated into a conical flask of Basal culture medium and culture is continued, including: inoculating the obtained seed liquid into a 500mL conical flask containing 200mL of fresh improved Basal culture medium, and placing it in a shaking fermentation in the dark at 28°C and 180rpm to continue culture.

[0010] In one embodiment of the present invention, the culture is continued by shaking fermentation at 28° C. and 180 rpm, and further comprising:

[0011] During the culture process, glucose and urea were continuously added.

[0012] In one embodiment of the present invention, the mass concentration of the added glucose is 10 g / L, and the mass concentration of the added urea is 0.9 g / L.

[0013] In one embodiment of the present invention, the improved Basal medium includes KH2PO4, MgSO4·7H2O, EDTA·Na2, H3BO3, Mg CaCl2·2H2O, FeSO4·7H2O, ZnSO4·7H2O, MnCl2·4H2O, CuSO4·5H2O, Co(NO3)2·6H2O, and MoO3.

[0014] In one embodiment of the present invention, the mass concentration ratio of KH2PO4, MgSO4·7H2O, EDTA·Na2, H3BO3, mgCaCl2·2H2O, FeSO4·7H2O, ZnSO4·7H2O, MnCl2·4H2O, CuSO4·5H2O, Co(NO3)2·6H2O, and MoO3 is 1.25:1.0:0.5:0.1142:0.111:0.0498:0.0882:0.0142:0.0157:0.0049:0.0071.

[0015] In one embodiment of the present invention, the initial pH of the improved Basal medium is 6.1.

[0016] As described above, a method of improving algae metabolism and lutein synthesis of the present invention has the following beneficial effects: by selecting C. protothecoides CS-41 algae species as culture seeds, C. protothecoides CS-41 can have a higher lutein accumulation under the same culture conditions. Before culturing the microalgae seed liquid, the growth and lutein accumulation in heterotrophic culture or fed-batch heterotrophic fermentation culture can be effectively promoted by using low-light pre-culture, thereby accelerating algae metabolism and increasing lutein production. That is, compared with dark pre-culture and high-light pre-culture, the algae cell carbon availability and carbon efficiency of the heterotrophic fermentation experimental group after low-light pre-culture are the highest, thus promoting cell growth and providing more substrates for carotenoid accumulation; similarly, the culture strategy of low-light-induced seed liquid post-heterotrophic fermentation is also applicable to fed-batch heterotrophic fermentation. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 The present invention is a flow chart of the method for improving algal metabolism and lutein synthesis.

[0018] Figure 2 It is a schematic diagram showing the lutein content of C. protothecoides CS-41 algae cultured in high-light thin tube, high-light thick tube, low-light thin tube and low-light thick tube PBR in a preferred embodiment of the present invention.

[0019] Figure 3 Shown is a schematic diagram showing the growth curve of C. protothecoides CS-41 under different light qualities in a preferred embodiment of the present invention.

[0020] Figure 4 Shown is a schematic diagram showing the final lutein production of C. protothecoides CS-41 cultured under different light qualities in a preferred embodiment of the present invention.

[0021] Figure 5 Shown is the growth curve of heterotrophic fermentation of C. protothecoides CS-41 after dark pre-culture, and the consumption and replenishment curves of glucose and urea in a preferred embodiment of the present invention.

[0022] Figure 6 Shown is the growth curve of heterotrophic fermentation of C. protothecoides CS-41 after low-light pre-culture in a preferred embodiment of the present invention. DETAILED DESCRIPTION

[0023] The following is an explanation of the embodiments of the present invention by specific examples, and those skilled in the art can easily understand other advantages and effects of the present invention from the contents disclosed in this specification. The present invention can also be implemented or applied through other different specific embodiments, and the details in this specification can also be modified or changed in various ways based on different viewpoints and applications without departing from the spirit of the present invention. It should be noted that the features in the following embodiments and the embodiments can be combined with each other without conflict. It should also be understood that the terms used in the embodiments of the present invention are intended to describe specific embodiments, rather than to limit the scope of protection of the present invention. The test methods for which specific conditions are not specified in the following examples are usually carried out under conventional conditions or according to the conditions recommended by the manufacturers.

[0024] It should be noted that the structures, proportions, sizes, etc. illustrated in the drawings of this specification are only used to match the contents disclosed in the specification for people familiar with this technology to understand and read, and are not used to limit the limiting conditions for the implementation of the present invention, so they have no substantial technical significance. Any modification of the structure, change in the proportional relationship, or adjustment of the size should still fall within the scope of the technical content disclosed by the present invention without affecting the effects and purposes that can be achieved by the present invention. At the same time, the terms such as "upper", "lower", "left", "right", "middle" and "one" quoted in this specification are only for the convenience of description, and are not used to limit the scope of the implementation of the present invention. The change or adjustment of their relative relationship should also be regarded as the scope of the implementation of the present invention without substantially changing the technical content.

[0025] Microalgae can be cultivated in three modes: autotrophic, heterotrophic and mixotrophic.

[0026] Photoautotrophic culture is a cultivation mode in which microalgae use solar energy to convert inorganic carbon into organic matter. Photoautotrophy is the most common growth mode of microalgae and is also considered the most environmentally friendly and economical way to cultivate microalgae. Since no organic carbon source is added during the culture, photoautotrophic culture has a lower risk of contamination compared to heterotrophic or mixed culture modes. In addition, the photoautotrophic mode is more conducive to the accumulation of pigments in microalgae cells. However, photoautotrophic microalgae have a lower growth rate and cell density, which results in lower total lutein production and increases the cost of harvesting biomass.

[0027] Although only applicable to some algae species, heterotrophic culture mode can be carried out in conventional fermenters in the dark. This mode is relatively cheap, easy to manage, safe and has a fairly high biomass productivity. In order to achieve high-density culture of microalgae, multiple studies have currently adopted a variety of culture strategies including batch fermentation and fed-batch fermentation. Another advantage of heterotrophic culture is that it is easy to scale up, and the productivity of laboratory mass production can be continuously replicated on an industrial scale, which simplifies the optimization process of large-scale cultivation. However, despite the ability to obtain higher biomass, heterotrophically cultured microalgae usually have a low pigment content due to the lack of light stimulation.

[0028] Mixotrophy is a mode that combines photoautotrophy and heterotrophy to utilize both inorganic and organic carbon sources. Although some scientists believe that the specific growth rate of microalgae cultured in a mixed trophic mode is approximately equal to the sum of the specific growth rates in photoautotrophy and heterotrophy, others believe that the metabolic mechanism of microalgae cells in a mixed trophic mode is more complex due to the potential synergy between photosynthesis and aerobic respiration. For example, Zhang et al. proposed that under mixed trophic culture, photosynthesis and aerobic respiration of Zofia suffruticosa have a subtle coordination of energy and carbon metabolism, where mixed trophic culture has higher biomass productivity and the largest biomass dry weight than the sum of photoautotrophy and heterotrophy. Under mixed trophic culture, carbon dioxide exhaled by aerobic respiration can be captured and reused by photosynthesis, thereby increasing biomass productivity. However, due to light limitations, when the cell density reaches a certain value, the microalgae cells in the reactor are similar to heterotrophic culture. Although the mixed trophic mode shows a higher biomass than the heterotrophic mode in laboratory batch culture, the mixed trophic mode is not cost-effective in industrial applications because the additional light will make the system more complex and lead to higher costs.

[0029] Based on the defects of the above-mentioned three culture modes of autotrophy, heterotrophy and mixotrophy, the present invention provides a method for improving algal metabolism and lutein synthesis. By selecting C.protothecoides CS-41 algae species as culture seeds, C.protothecoides CS-41 can have a higher lutein accumulation under the same culture conditions. Before culturing the microalgae seed liquid, the growth and lutein accumulation in heterotrophic culture or fed-batch heterotrophic fermentation culture can be effectively promoted by using low-light pre-culture. In other words, compared with dark pre-culture and high-light pre-culture, the algae cell carbon availability and carbon efficiency of the heterotrophic fermentation experimental group after low-light pre-culture are the highest, thus promoting cell growth and providing more substrates for carotenoid accumulation; similarly, the culture strategy of low-light-induced seed liquid post-heterotrophic fermentation is also applicable to fed-batch heterotrophic fermentation.

[0030] See also Figure 1In one embodiment of the present invention, the present invention provides a method for improving algal metabolism and lutein synthesis, comprising the following steps:

[0031] Step S10: selecting microalgae seeds;

[0032] Step S20: inoculating the microalgae seeds into a conical flask of Basal culture medium;

[0033] Step S30: culturing the conical flask under low light conditions for a predetermined number of days;

[0034] Step S40: repeating the low-light culture process to obtain seed solution;

[0035] Step S50: inoculating the obtained seed solution into a conical flask of Basal culture medium, and continuing to culture to obtain microalgae.

[0036] In this embodiment, through step S10: selecting microalgae seeds, it can be seen that in the selection of microalgae seeds, in order to improve the synthesis of lutein, the lutein content is an indicator for selecting algae species. Of course, similarly, the index factors of biomass and specific production rate also need to be considered. Through steps S20 to S40: inoculating the microalgae seeds in a conical flask of Basal culture medium; culturing the conical flask under low light conditions for an expected number of days; repeating the low light culture process to obtain seed liquid, it can be seen that in the process of improving algae metabolism and lutein synthesis, the microalgae seeds inoculated in the conical flask of Basal culture medium can be cultured by low light induction, so that the low light pre-cultured seed liquid can effectively increase the maximum biomass, lutein content and final lutein yield of the cultured microalgae. After culturing the conical flask under low light conditions for an expected number of days, repeating this pre-culture process again can obtain a stable seed liquid, and through step S50: inoculating the obtained seed liquid in a conical flask of Basal culture medium, continuing to culture, and obtaining microalgae. It can be seen that after obtaining the seed solution, when continuing to culture in Basal medium, the culture can be carried out by heterotrophic fermentation culture method, and of course, it can also be cultured by fed-batch heterotrophic fermentation culture method. Compared with dark pre-culture and high-light pre-culture, the microalgae cultured in the above manner have higher carbon availability of algae cells in the heterotrophic fermentation experimental group after low-light pre-culture and have higher carbon efficiency.

[0037] In one embodiment of the present invention, the selected microalgae seeds are C. protothecoides CS-41.

[0038] See also Figure 2, showing the lutein content of C.protothecoides CS-41 cultured in high-light capillary tubes, high-light thick tubes, low-light capillary tubes, and low-light thick tube PBRs. Among them, C.protothecoides CS-41 showed higher lutein content under several culture conditions, reaching up to 6.84 mg / g in low-light thick tubes. Light intensity is an important factor affecting lutein accumulation, and low light is more conducive to lutein accumulation. Moreover, in the selection of carbon sources in the culture medium, whether it is an inorganic carbon source or an organic carbon source, C.protothecoides CS-41 algae is more suitable as a microalgae for producing lutein.

[0039] Of course, in order to achieve the purpose of inducing culture using low light, microalgae seeds can also be selected from P.kessleri HH2.

[0040] In one embodiment of the present invention, the microalgae seeds are inoculated into a conical flask of Basal medium, comprising: inoculating the microalgae seeds into a 500 mL conical flask containing 200 mL of fresh modified Basal medium at 10% (v / v), and placing the microalgae seeds in a 28°C, 180 rpm shaking table for fermentation. In this embodiment, the shaking table can be a constant temperature shaking table of model DHZ-DA, and the oscillator can be a vortex oscillator of model 88880018. Of course, this process can be a culture method of heterotrophic fermentation after illumination pre-culture.

[0041] Specifically, the present invention also provides a method for improving algal metabolism and lutein synthesis by heterotrophic fermentation after light pre-culture, comprising: inoculating 10% (v / v) microalgae seeds into a 500 mL conical flask filled with 200 mL of fresh improved Basal medium, placing it in a shaking fermentation flask at 28°C and 180 rpm, and culturing it under low light of 30 μmol / m 2 / s for 2-3 days. Repeat the pre-culture process again to obtain a stable seed solution. The obtained seed solution was inoculated into a 500 mL conical flask containing 200 mL of fresh modified Basal medium, placed in a shaking fermentation culture at 28°C and 180 rpm, and cultured in the dark for 5 days.

[0042] In one embodiment of the present invention, the low light condition is 20-40 μmol / m 2 / s, the light quality of the low light is white light, and the estimated number of days for cultivation is 2 to 3 days.

[0043] In this embodiment, when low light conditions are selected, 30 μmol / m 2When optimizing the light quality, a white light LED strip was wrapped around a 500 mL conical flask and fixed so that the light intensity at the bottom of the conical flask containing uninoculated culture medium was 30 μmol / m 2 / s The cultured seed solution was inoculated at 10% inoculation amount into 200 mL of sterilized modified Basal medium and a 500 mL conical flask wrapped with a light strip, and cultured at 28°C and 180 rpm.

[0044] For optimal light quality selection for lutein accumulation, please refer to Figure 3 , showing the growth curve of C.protothecoides CS-41 under different light qualities. It can be found that the light quality has little effect on the maximum biomass. The growth rate of C.protothecoides CS-41 under white light conditions is the fastest, and the maximum biomass can reach 21.2g / L. Therefore, C.protothecoides CS-41 consumes glucose and urea faster under white light conditions. After reaching the plateau period, the biomass of algal cells under dark culture and white light culture is relatively stable, while the biomass under red light and blue light culture has decreased.

[0045] See also Figure 4 , showing the final lutein production of C.protothecoides CS-41 cultured under different light qualities. It can be seen that the order of lutein production is: white light > blue light > red light ≥ dark. C.protothecoides CS-41 can accumulate higher lutein content under white light. White light is the most suitable light quality for C.protothecoides CS-41 to accumulate lutein. White light has a balanced mixture of blue light and red light. Chlorophyll absorbs energy at 600-700nm, and lutein absorbs energy at 400-500nm. Therefore, algae cells cultured under white light have more light-harvesting antenna proteins to capture light energy, increasing the overall photosynthetic rate.

[0046] In one embodiment of the present invention, the obtained seed solution is inoculated into a conical flask of Basal medium, and further culture is performed, including: the obtained seed solution is inoculated into a 500 mL conical flask containing 200 mL of fresh improved Basal medium, and the culture is continued by shaking fermentation in a shaking table at 28°C and 180 rpm in the dark. In this embodiment, when continuing to culture, it can be a fed-batch heterotrophic fermentation culture after light pre-culture.

[0047] Specifically, the present invention also provides a method for improving algal metabolism and lutein synthesis by feeding heterotrophic fermentation after light pre-culture, comprising: inoculating 10% (v / v) seeds into a 500 mL conical flask containing 200 mL of fresh improved Basal medium, placing the flask in a shaking fermentation culturing at 28°C and 180 rpm, and culturing the flask under low light of 30 μmol / m 2 / s for 2-3 days. Repeat this pre-culture process again to obtain a stable seed solution. The obtained seed solution was inoculated into a 500 mL conical flask containing 200 mL of fresh modified Basal medium, and the initial inoculation amount of the three pre-culture modes was controlled to be the same. The fermentation culture was placed in a shaking incubator at 28°C and 180 rpm, and cultured in the dark for 12 days. Glucose and urea were added during the culture process to ensure the continuous growth of C. protothecoides CS-41.

[0048] In one embodiment of the present invention, the culture is continued by shaking fermentation at 28° C. and 180 rpm, further comprising:

[0049] During the culture process, glucose and urea were continuously added.

[0050] Do et al. conducted a central composite design and statistically modeled the biomass and lutein production of C. sorokiniana TH01. The study showed that under the optimal light intensity (75 μmol / m 2 / s, 10g / L sodium acetate concentration, N / P ratio of 35 and 2% CO2 conditions, high biomass concentration (26.21g / L) and lutein content (5.01mg / g) can be achieved simultaneously. Glycerol has also been tried in the mixed culture of Scenedesmus sp., but it performed poorly in increasing the lutein content (0.08%–0.1%) compared with the autotrophic mode. In heterotrophic and mixed culture modes, in order to maximize production, the focus is usually on achieving high biomass density rather than high lutein content. Therefore, in these culture modes, glucose is an optimal carbon source choice.

[0051] Nitrogen source is one of the most important components for lutein biosynthesis and biomass accumulation. Under nitrogen-free conditions, the lutein content in Zofia spp. decreased significantly under high light conditions. However, as the initial nitrate concentration increased from 4.4 mM to 13.2 mMs, the maximum lutein content of Desmodesmus sp. F51 increased significantly from 3.76 mg / g to 4.97 mg / g

[38] . Similarly, in Chlorella vulgaris, supplementation with nitrate significantly increased the lutein content to 10.5 mg / g. Therefore, an adequate nitrogen source is essential for the accumulation of lutein during microalgae cultivation. Nitrate, urea and ammonium salts are the most commonly used nitrogen sources in microalgae cultivation. A study showed that the nitrogen utilization efficiency of C. sorokiniana MB-1-M12 was as follows: urea > ammonium > nitrate. Compared with nitrate and uric acid, ammonium salts can be used immediately for algal growth and have lower energy consumption. However, culturing algae with ammonium salts will reduce the pH value due to the release of H+ ions during cell growth. Xie et al. adopted a dual carbon source cooperation strategy (simultaneous use of NaHCO3 and CO2) to solve the pH change during ammonium consumption, thereby achieving a lutein production of 5.22 mg / L / day. At the same order, the nitrogen cost of lutein production is: urea < ammonium < nitrate. From an economic point of view, urea is the best choice for industrial lutein production compared with other nitrogen sources.

[0052] In this embodiment, during the cultivation of microalgae, high-concentration substrate flow fermentation is a relatively time-saving and labor-saving fermentation method, that is, a relatively high level of substrate is added to the initial culture medium, and when glucose is observed to be exhausted, a large amount of carbon and nitrogen sources, namely glucose and urea, are supplemented again. This method can alleviate the substrate limitation caused by the low substrate concentration when the algae cells grow rapidly in the early stage. Specifically, glucose is a carbon source that is more suitable for the microalgae acoustic field, and 10g / L glucose with the highest substrate utilization rate is selected as the subsequent culture carbon source concentration.

[0053] See also Figure 5 and 6 , Figure 5 Figure 2 shows the growth curve of heterotrophic fermentation of C. protothecoides CS-41 after dark pre-culture, the consumption and replenishment curves of glucose and urea, Figure 6 The growth curve of heterotrophic fermentation of C.protothecoides CS-41 after low-light pre-culture, and the consumption and replenishment curves of glucose and urea. It can be seen that the final biomass obtained by fed-batch fermentation after dark pre-culture is 24.90g / L, while the final biomass obtained by fed-batch fermentation after low-light pre-culture is 30.85g / L. Low-light pre-culture seeds and fed-batch fermentation effectively increased the final biomass and growth rate of C.protothecoides CS-41.

[0054] Specifically, the mass concentration of the added glucose is 10 g / L, and the mass concentration of the added urea is 0.9 g / L.

[0055] In one embodiment of the present invention, the improved Basal medium includes KH2PO4, MgSO4·7H2O, EDTA·Na2, H3BO3, Mg CaCl2·2H2O, FeSO4·7H2O, ZnSO4·7H2O, MnCl2·4H2O, CuSO4·5H2O, Co(NO3)2·6H2O, and MoO3.

[0056] Preferably, the mass concentration ratio of KH2PO4, MgSO4·7H2O, EDTA·Na2, H3BO3, mgCaCl2·2H2O, FeSO4·7H2O, ZnSO4·7H2O, MnCl2·4H2O, CuSO4·5H2O, Co(NO3)2·6H2O, and MoO3 is 1.25:1.0:0.5:0.1142:0.111:0.0498:0.0882:0.0142:0.0157:0.0049:0.0071.

[0057] In this embodiment, the modified Basal medium ( / L): 1.25g KH2PO4, 1.0g MgSO4·7H2O, 0.5g EDTA·Na2, 114.2mg H3BO3, 111mg CaCl2·2H2O, 49.8mg FeSO4·7H2O, 88.2mg ZnSO4·7H2O, 14.2mg MnCl2·4H2O, 15.7mg CuSO4·5H2O, 4.9mg Co(NO3)2·6H2O, 7.1mg MoO3. Compared with Kuhl medium and BG11 medium, the modified Basal medium is more conducive to the growth of C. protothecoides CS-41 algae species.

[0058] Preferably, the initial pH of the improved Basal medium is 6.1.

[0059] In summary, the present invention discloses a method for improving algae metabolism and lutein synthesis. By selecting C.protothecoides CS-41 algae species as culture seeds, C.protothecoides CS-41 can have a higher lutein accumulation under the same culture conditions. Before culturing microalgae seeds, etc., low-light pre-culture can effectively promote growth and lutein accumulation in heterotrophic culture or fed-batch heterotrophic fermentation culture, thereby accelerating algae metabolism and increasing lutein production. That is, compared with dark pre-culture and high-light pre-culture, the algae cell carbon availability and carbon efficiency of the heterotrophic fermentation experimental group after low-light pre-culture are the highest, thus promoting cell growth and providing more substrates for carotenoid accumulation; similarly, the culture strategy of low-light induction of seed liquid and then heterotrophic fermentation is also applicable in fed-batch heterotrophic fermentation. Therefore, the present invention effectively overcomes the various shortcomings of the prior art and has a high industrial utilization value.

[0060] The above embodiments are merely illustrative of the principles and effects of the present invention, and are not intended to limit the present invention. Anyone familiar with the art may modify or alter the above embodiments without departing from the spirit and scope of the present invention. Therefore, all equivalent modifications or alterations made by a person of ordinary skill in the art without departing from the spirit and technical concept disclosed by the present invention shall still be covered by the claims of the present invention.

Claims

1. A method for improving algal metabolism and lutein synthesis, characterized in that: The steps include: Selecting microalgae seeds; Inoculating the microalgae seeds into a conical flask containing Basal medium; Incubate the conical flask under low light conditions for a predetermined number of days; Repeat the low-light culture process to obtain seed solution; The obtained seed solution is inoculated into a conical flask of Basal culture medium, and the culture is continued to obtain microalgae.

2. The method for improving algal metabolism and lutein synthesis according to claim 1, characterized in that: The selected microalgae seeds were C. protothecoides CS-41.

3. The method for improving algal metabolism and lutein synthesis according to claim 1, characterized in that: The microalgae seeds are inoculated in a conical flask of Basal medium, including: The microalgae seeds were inoculated into a 500 mL conical flask filled with 200 mL of fresh modified Basal medium at 10% (v / v), and placed in a shaking fermentation culture at 28° C. and 180 rpm.

4. The method for improving algal metabolism and lutein synthesis according to claim 1, characterized in that: The low light condition is 20-40 μmol / m 2 / s, the light quality of the low light is white light, and the estimated number of days for cultivation is 2 to 3 days.

5. The method for improving algal metabolism and lutein synthesis according to claim 1, characterized in that: The obtained seed solution is inoculated into a conical flask of Basal medium and culture is continued, including: The obtained seed solution was inoculated into a 500 mL conical flask containing 200 mL of fresh improved Basal medium, and the culture was continued by shaking fermentation in a shaker at 28° C. and 180 rpm in the dark.

6. The method for improving algal metabolism and lutein synthesis according to claim 5, characterized in that: The culture was continued by shaking the fermentation in a shaking incubator at 28°C and 180 rpm, and further comprising: During the culture process, glucose and urea were continuously added.

7. The method for improving algal metabolism and lutein synthesis according to claim 6, characterized in that: The mass concentration of the added glucose is 10 g / L, and the mass concentration of the added urea is 0.9 g / L.

8. The method for improving algal metabolism and lutein synthesis according to claim 3 or 5, characterized in that: The modified Basal medium includes KH2PO4, MgSO4·7H2O, EDTA·Na2, H3BO3, Mg CaCl2·2H2O, FeSO4·7H2O, ZnSO4·7H2O, MnCl2·4H2O, CuSO4·5H2O, Co(NO3)2·6H2O, and MoO3.

9. The method for improving algal metabolism and lutein synthesis according to claim 8, characterized in that: The mass concentration ratios of KH2PO4, MgSO4·7H2O, EDTA·Na2, H3BO3, mgCaCl2·2H2O, FeSO4·7H2O, ZnSO4·7H2O, MnCl2·4H2O, CuSO4·5H2O, Co(NO3)2·6H2O, and MoO3 are 1.25:1.0:0.5:0.1142:0.111:0.0498:0.0882:0.0142:0.0157:0.0049:0.0071.

10. The method for improving algal metabolism and lutein synthesis according to claim 1, characterized in that: The initial pH of the modified Basal medium was 6.1.

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