A large-scale continuous culture method for mixed microalgae in salt lakes

Abstract

The present invention discloses a large-scale continuous culture method for mixed microalgae in salt lakes. The culture process requires the selection of advantageous mixed microalgae species from Yuncheng Salt Lake, screening and filtration of saturated brine and reclaimed water, primary expansion culture, secondary expansion culture, and large-scale culture in Yuncheng Salt Lake. The mixed microalgae are Chlorella and Dunaliella, and the algae species used are all obtained from the brine of Yuncheng Salt Lake. The culture medium is saturated brine and reclaimed water from Yuncheng Salt Lake, and the saturated brine and fresh water used are required to be filtered with a 200-mesh sieve. The molar ratio of sodium ions to magnesium ions in saturated brine is (0.5-1.5):1.0. The pH range of the culture medium is 7.0-9.0, and the salinity of the culture medium is 8. o Be / ~10 o Be / The supplements are soybean meal fermentation liquid, urea, and KH2PO4. The present invention effectively increases the biomass of microalgae and realizes the large-scale production of microalgae.

Description

Technical Field

[0001] The invention relates to the field of microalgae cultivation biotechnology, in particular to a large-scale continuous cultivation method for salt lake mixed microalgae. Background Art

[0002] Microalgae are a type of autotrophic plant found on land and in the oceans. They are nutrient-rich and have high photosynthetic efficiency. There are approximately 30,000 documented species of microalgae. Phototrophy and heterotrophy are two methods of producing microalgae.

[0003] Driven by light energy, microalgae can absorb CO2, convert that energy into chemicals stored within their cells, and simultaneously release oxygen. Microalgae's cellular metabolism produces products such as polysaccharides, proteins, oils, and pigments, all of which hold great promise for development in areas such as food, medicine, and liquid fuels. Consequently, microalgae cultivation is becoming increasingly widespread. However, existing microalgae cultivation methods typically cultivate a single algal species, resulting in high culture medium preparation costs and low microalgae cultivation efficiency.

[0004] The search for prior art on mixed microalgae and microalgae culture is as follows:

[0005] Patent application publication number CN112410225A (A Microalgae Cultivation Method) provides a microalgae cultivation method. This method primarily specifies the microalgae expansion process and general parameters such as the culture medium and light intensity. However, it does not specify the most critical microalgae species. The algae species is crucial, as optimal cultivation conditions vary for different species. The invention also fails to address the cultivation of mixed microalgae.

[0006] Patent application publication number CN114133039A (A method for treating municipal sewage using mixed microalgae) provides a method for treating municipal sewage using mixed microalgae, wherein the mixed microalgae are Chlorella and Scenedesmus, but no mixed culture of Chlorella and Dunaliella has been reported.

[0007] Patent application publication number CN 111607523A (A High-yield Industrialized Microalgae Cultivation Method) discloses a high-yield industrialized microalgae cultivation method. This method primarily specifies parameters such as the microalgae expansion facility site, temperature control, and CO2 generation and release devices. However, it does not specify the microalgae species used for cultivation. The invention also fails to address the cultivation of mixed microalgae. Summary of the Invention

[0008] To address the shortcomings of existing technologies, the present invention provides a large-scale continuous culture method for mixed salt lake microalgae. The microalgae species are all derived from the dominant algae species of Yuncheng Salt Lake, allowing the local algae to adapt to the brine environment of Yuncheng Salt Lake without domestication. The culture medium uses saturated brine and recycled water from Yuncheng Salt Lake, making it a widely available and cost-effective source. The cultured mixed microalgae do not require harvesting, and can be directly used as live bait for local Artemia culture. This method not only improves Artemia quality but also significantly reduces bait cost, resulting in significant economic benefits.

[0009] To achieve the above object, the present invention is implemented by adopting the following technical solutions:

[0010] A large-scale continuous culture method for mixed microalgae in salt lakes comprises the following steps:

[0011] Step 1: separating the dominant mixed microalgae species in the brine of Yuncheng Salt Lake; the mixed microalgae species include Chlorella and Dunaliella.

[0012] The Artemia culture water is filtered through a 200-mesh silk screen, and the filtered sample is the initial sample; after the initial sample is allowed to stand, the algae float up, and a circle of green color is attached to the liquid surface. The green algae seeds are inoculated into the culture medium and expanded to the million level per milliliter.

[0013] The culture medium is a nutrient base determined by the laboratory based on the saturated brine used to prepare Artemia aquaculture water: salinity 8 o Be / ~10 o Be / ; NaNO3 0.2g / L~0.4g / L, urea amount is 0.5g / L~1.0g / L, KH2PO40.01g / L~0.3g / L, NaHCO30.2g / L~1.0g / L; pH value 7.0~9.0.

[0014] The culture medium is saturated brine from Yuncheng Salt Lake, with a molar ratio of sodium ion to magnesium ion of (0.5-1.5):1.0, a pH of 7.0-9.0, and a salinity of 8. o Be / ~10 o Be / .

[0015] Step 2: The advantageous mixed microalgae species obtained in step 1 are cultured in multiple stages in sequence. The brine and fresh water used in the culture medium are required to be filtered with a 200-mesh sieve and sterilized.

[0016] The nutrient medium for expansion is: salinity 8 o Be / ~10 o Be / ; NaNO3 0.2g / L~0.4g / L; urea amount is 0.5g / L~1.0g / L; KH2PO4 0.01g / L~0.3g / L; pH value 7.0~9.0.

[0017] The saturated brine of Yuncheng Salt Lake was used for expansion, in which the molar ratio of sodium ion to magnesium ion was (0.5-1.5):1.0, pH was 7.0-9.0, and salinity was 8. o Be / ~10 o Be / .

[0018] Stirring method: Place the nano aeration plate at the bottom of the pool and let air flow into the water in the form of microbubbles.

[0019] Step 3: Take the mixed microalgae expanded in step 2 above and carry out mixed culture in a pond in Yuncheng Salt Lake to increase the biomass.

[0020] (1) Preparation before microalgae cultivation

[0021] Pond cleaning

[0022] Before filling the breeding pond with water, drain the pond water, clean the pond bottom mud, expose it to the sun, use the quicklime dry method to clean the pond, and reinforce and prevent leakage on the pond bank.

[0023] Watering and fertilization

[0024] Water inlet: The initial water inlet is saturated brine for preparing Artemia breeding water, with a depth of 25-35cm. It is left for 20 days and the breeding pond is disinfected and sterilized with high-concentration brine.

[0025] Fertilization: Based on a 100-mu aquaculture pond, after the initial water filling and 20 days of fertilization, fermented soybean meal is selected as the organic fertilizer, and 50-150 kg of fermented soybean meal is applied per mu. The water content is 0.6-0.8 m3, and 5-15 kg of urea and 2-10 kg of KH2PO4 are applied per mu. The salinity is adjusted to 8. o Be / ~10 o Be / , pH value 7.0 ~ 9.0; the incoming and replenishing water are filtered through a filter screen, and the screen is required to be above 200 mesh.

[0026] (2) Vaccination

[0027] Mixed microalgae species are taken from the expansion pond and inoculated into the microalgae culture pond. A continuous culture method is adopted. When the concentration of the microalgae cultured in the microalgae culture pond reaches the million level per milliliter, the mixed microalgae liquid is injected into the Artemia culture pond. Each pond is injected with 3cm to 5cm at a time, and at least 0.4m of algae liquid remains. Then, water is added to the microalgae culture pond to 0.6m to 0.8m, and nutrients are supplemented as needed to achieve continuous culture.

[0028] Further preferably, in step 1, the nutrient medium: salinity 9 o Be / ;NaNO3 0.34g / L, urea 0.8g / L, KH2PO4 0.03g / L, NaHCO3 0.8g / L.

[0029] Further preferably, in step 2, the culture medium for expansion is a nutrient base determined in the laboratory based on the saturated brine used to prepare Artemia aquaculture water: salinity 9 o Be / ;NaNO3 0.34g / L, urea 0.8g / L, KH2PO4 0.01g / L.

[0030] Further preferably, in step 1 and step 2, the saturated brine of Yuncheng Salt Lake has a molar ratio of sodium ions to magnesium ions of 1.0, a pH of 8.0, and a salinity of 9. o Be / .

[0031] Further preferably, in step 3, the amount of quicklime used per mu is 80-120 kg. Fertilization: Based on a 100 mu aquaculture pond, apply 10 kg of fermented soybean meal, 1 kg of urea, and 5 kg of KH2PO4 per mu, and adjust the salinity to 8. o Be / , pH 8.0.

[0032] The beneficial effects of the present invention are as follows:

[0033] 1. The raw materials of the present invention are locally sourced, the cost is low, the process is simple, and it is easy to implement. It has good economic and social benefits and can be promoted and applied to promote the healthy development of microalgae cultivation and aquaculture.

[0034] 2. The salt lake mixed microalgae cultured in the present invention is a natural green and environmentally friendly biological bait with good activity, comprehensive nutrition, and easy absorption, which can effectively improve the utilization rate and conversion rate of biological feed.

[0035] 3. By cultivating salt lake mixed microalgae through the present invention, the nitrogen, phosphorus and organic matter content in the salt lake can be effectively reduced, the BOD value in the salt lake water body can be reduced, and the salt lake water quality can be purified, thereby achieving obvious salt lake ecological benefits.

[0036] The invention has a reasonable design. The culture medium is selected from saturated brine and reclaimed water of Yuncheng Salt Lake, which are reasonably prepared and then supplemented with nutrients. By adjusting the content of mineral components in the culture medium and stabilizing the pH environment, a suitable growth environment is provided for the mixed microalgae, thereby effectively increasing the biomass of the microalgae and realizing large-scale production of the microalgae. The invention has good practical application value. DETAILED DESCRIPTION

[0037] The technical solutions in the embodiments of the present invention are described clearly and completely below, and those skilled in the art can implement them according to the description.

[0038] 1. Determination of dominant microalgae species

[0039] Through the investigation of the plankton resource system of Yuncheng Salt Lake, the following algae were found, see Table 1.

[0040] Table 1 Algae in Yuncheng Salt Lake

[0041]

[0042]

[0043] May to August is the peak season for Artemia. The salinity of Artemia aquaculture water is 12-15. o Be / The microalgae in aquaculture water are primarily Dunaliella and Chlorella, which Artemia feed on. Dunaliella is a single-celled phytoplankton with an oval, elliptical, or pear-shaped body, 14 to 22 μm long and 3 to 14 μm wide. Chlorella is a single-celled algae with a diameter of 3 to 8 μm. Ultimately, Dunaliella and Chlorella were determined to be the dominant microalgae species.

[0044] 2. Basis for determining culture medium salinity:

[0045] Table 2 Aquaculture salinity and algae growth

[0046]

[0047] Less than 8 o Be / Algae larger than 50μm can easily grow in brine. Artemia is a filter feeder, and its suitable food particles must be smaller than 50μm. Algae larger than 50μm cannot be eaten by Artemia.

[0048] From the investigation of plankton and biological environment in the brine of Yuncheng Salt Lake, it is known that 5-15 o Be / The brine contains both Dunaliella and Chlorella. The salinity of the water used to cultivate microalgae is 8 o Be / ~10 o Be / The reasons are: first, this salinity can inhibit the growth of large algae; second, the algae liquid with this salinity enters the Artemia breeding pond, which can inhibit the growth of large algae; third, this salinity liquid enters the Artemia breeding pond, which can replenish the evaporated breeding water; fourth, Dunaliella and Chlorella can grow faster under this salinity.

[0049] 3. Isolation, purification and expansion of dominant algae species

[0050] (1) Isolation, purification and expansion of Dunaliella

[0051] The dilution separation method and the plate separation method were used to separate Dunaliella species from Artemia culture water.

[0052] The culture medium is a nutrient base determined by the laboratory based on the saturated brine used to prepare Artemia aquaculture water: salinity 9 o Be / NaNO3 0.34g / L, KH2PO4 0.03g / L, NaHCO3 0.8g / L; pH 8.0. Sterilize the culture medium in a high-temperature autoclave. Use this medium to purify and propagate Dunaliella salina.

[0053] (2) Isolation, purification and expansion of Chlorella

[0054] The dilution separation method and the plate separation method were used to separate Dunaliella species from Artemia culture water.

[0055] The culture medium is a nutrient base determined by the laboratory based on the saturated brine used to prepare Artemia aquaculture water: salinity 9 o Be / NaCO₃ 0.02g / L, NaNO₃ 2.0g / L, KH₂PO₄ 0.02g / L, urea 0.8g / L; pH 6.0. The culture medium is sterilized in a high-temperature autoclave. This medium is used to purify and propagate Chlorella vulgaris.

[0056] (3) Enrichment and laboratory expansion of mixed algae

[0057] Filter the Artemia culture water through a 200-mesh silk screen. The filtered sample is the initial sample. After the initial sample is left to stand for a period of time, algae will float to the surface, and a circle of green algae attached to the liquid surface will be inoculated into the culture medium and expanded to a million per milliliter.

[0058] The culture medium is a nutrient base determined by the laboratory based on the saturated brine used to prepare Artemia aquaculture water: salinity 9 o Be / NaNO3 0.34 g / L, urea 0.8 g / L, KH2PO4 0.03 g / L, NaHCO3 0.8 g / L; pH 7.0-9.0. Sterilize the culture medium in a high-temperature autoclave and use it for expansion.

[0059] Table 3 Data of the test on separation of dominant algae species and purification of nutrient medium

[0060]

[0061] 4. Culture medium selection

[0062] The composition of the culture medium is directly related to the effectiveness of large-scale outdoor cultivation. Open-air microalgae cultivation requires large quantities of high-salinity brine. Adding salt to fresh water can also achieve the required salinity, but this significantly increases cultivation costs. The present invention uses saturated brine, the same brine used to prepare Artemia culture water, to prepare the culture medium. Because saturated brine has a strong inhibitory effect on predators, it can be used directly in microalgae cultivation without disinfection. Furthermore, the brine contains a wealth of nutrients that essentially meet the growth needs of microalgae, requiring only the addition of elements such as nitrogen, phosphorus, and carbon. The brine and fresh water used in the culture medium must be filtered through a 200-mesh sieve.

[0063] 5. The large-scale continuous culture method of salt lake mixed microalgae described in the present invention requires the selection of advantageous mixed microalgae species from Yuncheng Salt Lake, screening and filtration of saturated brine and reclaimed water, primary expansion culture, secondary expansion culture, and large-scale culture in Yuncheng Salt Lake during the culture process.

[0064] Among them, the mixed microalgae are Chlorella and Dunaliella, and the ratio of Chlorella to Dunaliella is 1-10:10-1. All algae species used are obtained from the brine of Yuncheng Salt Lake. The culture medium is saturated brine and reclaimed water from Yuncheng Salt Lake. The saturated brine and fresh water used are required to be filtered with a 200-mesh sieve. The saturated brine is chloride ion, sodium ion, sulfate ion, and magnesium ion, and the molar ratio of sodium ion to magnesium ion is (0.5-1.5):1.0. The pH range of the culture medium is 7.0-9.0, and the salinity of the culture medium is 8 o Be / ~10 o Be / The supplements were soybean meal fermentation broth, urea, and KH2PO4.

[0065] The specific embodiments are as follows:

[0066] A large-scale continuous culture method for mixed microalgae in salt lakes, comprising the following steps:

[0067] Step 1: Isolate the dominant mixed microalgae species in the brine of Yuncheng Salt Lake; the mixed microalgae species include Chlorella and Dunaliella.

[0068] Filter the Artemia culture water through a 200-mesh silk screen. The filtered sample is the initial sample. After the initial sample is left to stand for a period of time, algae will float to the surface, and a circle of green algae attached to the liquid surface will be inoculated into the culture medium and expanded to a million per milliliter.

[0069] Among them, the culture medium is the nutrient base determined by the laboratory based on the saturated brine used to prepare Artemia culture water: salinity 9 o Be / NaNO₃ 0.34 g / L, urea 0.8 g / L, KH₂PO₄ 0.03 g / L, NaHCO₃ 0.8 g / L; pH 7.0-9.0. Sterilize the culture medium in an autoclave and use it for expansion.

[0070] Step 2: Carry out primary and secondary expansion culture respectively for the dominant algae species determined in step 1. The brine and fresh water used in the culture medium are required to be filtered with a 200-mesh sieve and sterilized.

[0071] Primary expansion: 2 L of the mixed microalgae species cultured in the laboratory in step 1 was inoculated into a 400 L white plastic culture box for primary expansion outdoors.

[0072] Secondary expansion: Take 100L of mixed microalgae solution from the first-stage expansion and inoculate it to 100ml in a white plastic culture box. 2 Secondary expansion cultivation is carried out in the breeding pond.

[0073] Among them, the culture medium used for expansion is the nutrient base determined by the laboratory based on the saturated brine used to prepare Artemia culture water: salinity 9 o Be / ; NaNO3 0.34g / L, urea 0.8g / L, KH2PO4 0.01g / L; pH value 8.0.

[0074] Agitation method: Place the nano-aeration plate at the bottom of the bucket to introduce air into the water in the form of microbubbles. This improves photosynthesis efficiency, prevents algae from sinking and agglomerating, and reduces nutrient and gas gradients. It also replenishes CO2 and provides a carbon source.

[0075] The saturated brine of Yuncheng Salt Lake was used for expansion, with a molar ratio of sodium ion to magnesium ion of 1.0, a pH of 8.0, and a salinity of 9. o Be / .

[0076] Step 3: Take the mixed microalgae expanded in step 2 above and carry out mixed culture in a 100-mu pond in Yuncheng Salt Lake to increase the biomass.

[0077] (1) Preparation before microalgae cultivation

[0078] Pond cleaning

[0079] The aquaculture pond covers an area of ​​100 mu and is approximately 1.2 meters deep. Thirty days before filling, drain the pond and clean the bottom mud. After two weeks of exposure to the sun, use 100 kg of quicklime per mu for dry cleaning. The bank around the pond is reinforced and leak-proofed.

[0080] Watering and fertilization

[0081] Water inlet: The initial water inlet is saturated brine for preparing Artemia breeding water, with a depth of 25cm. It is left for 20 days and the breeding pond is disinfected and sterilized with high-concentration brine.

[0082] Fertilization: Fermented soybean meal is the organic fertilizer of choice. After the initial watering and 20 days of placement, apply 100 kg of fermented soybean meal per mu, replenish water to 0.6 m, apply 1 kg of urea and 5 kg of KH2PO4 per mu. Adjust the salinity to 8 o Be / , pH value 8.0. The inlet and replenishing water are filtered through a filter screen with a mesh size of 200 or above.

[0083] (2) Vaccination

[0084] In 100m 2 Take 15-30ml of mixed microalgae species from the secondary expansion pool 3 (This example selects 20m 3 ), and then inoculated into a 100-acre microalgae cultivation pond.

[0085] A continuous culture method is used. When the concentration of microalgae in the microalgae culture pond reaches millions per milliliter, the mixed microalgae solution is injected into the Artemia culture pond, with 5cm of solution injected into each pond at a time, leaving at least 0.4ml of solution remaining. The microalgae culture pond is then topped up with water to 0.6ml, and nutrients are supplemented as needed to achieve continuous culture.

[0086] (3) Daily management of microalgae cultivation

[0087] Appropriate fertilization: Apply urea, KH2PO4, and NaHCO3 solutions weekly. Apply less fertilizer or no fertilizer on rainy days, and apply plenty of fertilizer on sunny days. Apply fermented soybean meal once every month. A reasonable combination of inorganic and organic fertilizers is recommended.

[0088] Water quality measurement: Measure various physical and chemical indicators of water bodies every day, including daily average water temperature, pH, dissolved oxygen, ammonia nitrogen content, nitrite content and water color changes.

[0089] Agitation method: Mainly rely on the wind agitation of Yuncheng Salt Lake, combined with appropriate mechanical agitation. Since Dunaliella cells have no cell walls and are easily damaged, the agitation method should not be too violent. Wheel agitation is selected, stirring 3 to 4 times a day, each time for 1 hour.

[0090] Water regulation: Due to the high daily evaporation rate of the Yuncheng Salt Lake, fresh water must be replenished promptly. If continuous rainfall or heavy rain reduces the salinity in the aquaculture pond, the brine salinity must be adjusted promptly.

[0091] Microscopic examination: Take water samples with a plankton net and use a microscope to observe whether there are other algae and harmful organisms in the water. If there are any, kill them with drugs to ensure the purity of the microalgae and prevent harm.

[0092] Keep records: record the fertilizers, medicines, water quality indicators and various conditions used that day.

[0093] (4) Prevention and control of algae pests

[0094] At low temperatures, microalgae do not grow vigorously, so the introduction of species in the early stages of cultivation can quickly make microalgae become the dominant population.

[0095] It competes with other algae and inhibits their growth. In the middle and late stages of cultivation, the microalgae have become the dominant species, and other algae and harmful organisms cannot reproduce in large numbers.

[0096] Microalgae pests primarily include rotifers, cladocerans, various insect larvae, and various protozoa that feed on microalgae. Aquatic insecticides such as trichlorfon and pyrethroids can be used to control these microalgae pests. However, it's important not to use a single pesticide for extended periods; instead, alternate between several to prevent drug resistance.

[0097] (5) Microalgae biomass index

[0098] Microscopic observation of the 100-acre microalgae cultivation pond revealed a concentration of microalgae reaching millions per milliliter, primarily consisting of Dunaliella and Chlorella. The water appeared emerald green to the naked eye.

[0099] The blood cell count method was used.

[0100] Preparation of Lugol's solution: Dissolve 6g of potassium iodide in 20ml of water. After it is completely dissolved, add 4g of iodine and shake thoroughly. After the iodine is fully dissolved, adjust the volume to 100ml (pack in a brown glass bottle).

[0101] Prepare the algae solution to be tested (the number of algae cells in each grid is preferably 5 to 10, and the algae solution to be tested does not need to be diluted), fix it with Lugol's solution, and add 10 to 15 ml of Lugol's solution to every 1000 ml of algae solution.

[0102] Take a clean blood cell counting chamber (25×16 type) and cover the counting area with a coverslip.

[0103] Use a 20 μl pipette to absorb the algae solution fixed with Lugol's solution and drip a small drop along the lower edge of the cover glass from the grooves on both sides of the middle platform of the counting chamber.

[0104] Let it sit for a while to allow the cells to settle on the counting plate and no longer drift with the liquid. Place the hemocytometer on the microscope stage and find the counting area under a low-power microscope first, then switch to a 400x microscope for counting.

[0105] Count the number of algal cells in the four corners and the central grid of the large square, and repeat the counting three times.

[0106] Calculation formula

[0107] Number of algal cells = (number of algal cells in 80 cells / 80) × 400 × 10 4 × dilution factor

[0108] Table 4 Algal cell count

[0109]

[0110] (6) Determination of microalgae harvesting and feeding methods

[0111] The main purpose of cultivating microalgae is to feed Artemia. The breeding site is very close to the Artemia breeding base. The algae liquid is directly pumped out through the circulating river into the Artemia breeding pond without special harvesting.

[0112] The salt lake mixed microalgae cultivation method of the present invention has been described through specific examples. Those skilled in the art may refer to the present invention and appropriately modify the composition and ratio of the ingredients to achieve other corresponding objectives. Such modifications do not depart from the scope of the present invention. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention are intended to be included within the scope of protection of the present invention.

Claims

1. A large-scale continuous culture method for mixed microalgae from salt lakes, characterized by: The steps include: Step 1: Isolate the dominant mixed microalgae species from the brine of Yuncheng Salt Lake; the mixed microalgae species include Chlorella and Dunaliella; Filter the Artemia culture water with a 200-mesh silk screen. The filtered sample is the initial sample. After the initial sample is allowed to stand, algae will float up and a circle of green algae attached to the liquid surface will be inoculated into the culture medium and expanded to a concentration of one million per milliliter. The culture medium is a nutrient base determined by the laboratory based on the saturated brine used to prepare Artemia aquaculture water: salinity 8 o Be / ~10 o Be / ; NaNO3 0.2g / L~0.4g / L, urea amount is 0.5g / L~1.0g / L, KH2PO4 0.01g / L~0.3g / L, NaHCO3 0.2g / L~1.0g / L; pH value 7.0~9.0; The culture medium is saturated brine from Yuncheng Salt Lake, in which the molar ratio of sodium ion to magnesium ion is (0.5-1.5):1.0, the pH is 7.0-9.0, and the salinity is 8 o Be / ~10 o Be / ; Step 2: The advantageous mixed microalgae species obtained in step 1 are cultured in multiple stages in sequence. The brine and fresh water used in the culture medium are required to be filtered with a 200-mesh sieve and sterilized. The nutrient medium for expansion is: salinity 8 o Be / ~10 o Be / ; NaNO3 0.2g / L~0.4g / L; urea amount 0.5g / L~1.0g / L; KH2PO4 0.01g / L~0.3g / L; pH value 7.0~9.0; Among them, the saturated brine of Yuncheng Salt Lake is used for expansion, in which the molar ratio of sodium ion to magnesium ion is (0.5-1.5):1.0, the pH is 7.0-9.0, and the salinity is 8 o Be / ~10 o Be / ; Stirring method: Place the nano aeration plate at the bottom of the pool and introduce air into the water in the form of microbubbles; Step 3: Take the mixed microalgae expanded in step 2 above and perform mixed culture in a pond in Yuncheng Salt Lake to increase the biomass; (1) Preparation before microalgae cultivation ①Pond cleaning Before filling the aquaculture pond, drain the water, clean the pond bottom mud, expose it to the sun, use quicklime dry method to clean the pond, and reinforce the pond bank and prevent leakage; ② Watering and fertilization Water inlet: The initial water inlet is saturated brine for preparing Artemia culture water, with a depth of 25-35 cm. Leave it for 20 days and use high-concentration brine to disinfect and sterilize the culture pond; ③ Fertilization: Taking the area of ​​the breeding pond as 100 mu, after the initial water filling and 20 days, the organic fertilizer is fermented soybean meal, 50kg to 150kg of fermented soybean meal per mu, and the water is replenished to 0.6m~0.8m, and 5~15kg of urea and 2~10kg of KH2PO4 per mu are applied; adjust the salinity to 8 o Be / ~10 o Be / , pH value 7.0 ~ 9.0; the inlet and replenishing water are filtered through the filter screen, the screen is required to be above 200 mesh; (2) Vaccination Mixed microalgae species are taken from the expansion pond and inoculated into the microalgae culture pond. A continuous culture method is adopted. When the concentration of the microalgae cultured in the microalgae culture pond reaches the million level per milliliter, the mixed microalgae liquid is injected into the Artemia culture pond. Each pond is injected with 3cm to 5cm at a time, and the remaining algae liquid is at least 0.4m. Then, water is added to the microalgae culture pond to 0.6m to 0.8m, and nutrients are supplemented as needed to achieve continuous culture.

2. The method for large-scale continuous cultivation of mixed salt lake microalgae according to claim 1, characterized in that: In step 1, nutrient medium: salinity 9 o Be / ;NaNO3 0.34g / L, urea 0.8g / L, KH2PO4 0.03g / L, NaHCO3 0.8g / L.

3. The large-scale continuous culture method of salt lake mixed microalgae according to claim 2, characterized in that: In step 2, the culture medium used for expansion is the nutrient base determined by the laboratory based on the saturated brine used to prepare Artemia aquaculture water: the salinity is 9 o B e / ;NaNO3 0.34g / L, urea 0.8g / L, KH2PO4 0.01g / L.

4. The large-scale continuous culture method of salt lake mixed microalgae according to claim 3, characterized in that: In step 1 and step 2, the saturated brine of Yuncheng Salt Lake has a molar ratio of sodium ion to magnesium ion of 1.0, a pH of 8.0, and a salinity of 9. o Be / .

5. The large-scale continuous culture method of salt lake mixed microalgae according to claim 4, characterized in that: In step 3, the amount of quicklime used per mu is 100 kg.

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