Microalgae culture medium based on plant extract and preparation method thereof

Abstract

The invention relates to a microalgae culture medium based on plant extracts and a preparation method thereof, and belongs to the technical field of algae culture media, the microalgae culture medium specifically comprises the following raw materials: blueberry peel extracts, cellulose nanofibers, a carbon source, a system stabilizer, inorganic salt, trace elements and sterile seawater; the carbon source is obtained by mixing glycerol and pretreated cane molasses according to a mass ratio of 1: (2-2.5). In the preparation method, processes in different components are coupled, the cellulose nanocrystals are extracted in an acid environment, and meanwhile, the extraction amount of the cellulose nanocrystals and the activity of anthocyanin and the like are regulated and controlled through the pH value, so that the characteristics of stability, oxidation resistance and the like of the components in the extraction process are maintained. The cane molasses is pretreated, and then the cane molasses and glycerol are jointly used as a carbon source, and the cane molasses and the blueberry peel extract have a synergistic effect in the aspect of promoting microalgae growth.

Description

Technical Field

[0001] This application relates to the technical field of algal culture media, and more specifically, to a microalgal culture medium based on plant extracts and a preparation method thereof. Background Art

[0002] Currently, microalgal culture has become a core solution in the fields of aquaculture, environmental governance, and green energy by integrating ecological services, resource recycling, and the development of high-value-added products. In the future, with the deep integration of synthetic biology and the circular economy, microalgae will play an irreplaceable role in global sustainable development. Most existing microalgal culture media use a single carbon source such as glucose or glycerol, resulting in a problem of mismatch between the absorption rate and metabolic pressure, which easily leads to the limitation of carbon source supply by light and CO2 diffusion, and it is difficult to meet the growth requirements of microalgae.

[0003] In addition, when agricultural waste such as cane molasses is directly used as a carbon source, components such as cellulose and pectin are not decomposed, hindering the absorption of microalgae.

[0004] Based on the above statements, this application provides a microalgal culture medium based on plant extracts and a preparation method thereof. Summary of the Invention

[0005] To solve the problems raised in the background art, this application provides a microalgal culture medium based on plant extracts and a preparation method thereof.

[0006] This application provides a microalgal culture medium based on plant extracts and a preparation method thereof, adopting the following technical solution:

[0007] A microalgal culture medium based on plant extracts, comprising the following raw materials in parts by mass:

[0008] 0.5 - 2 parts of blueberry peel extract, 0.01 - 0.05 parts of cellulose nanofibers, 20 - 40 parts of carbon source, 1 - 5 parts of system stabilizer, 6.52 - 7.74 parts of inorganic salts, 0.002 - 0.003 parts of trace elements, and 1000 parts of sterile seawater;

[0009] The carbon source is obtained by mixing glycerol and pretreated cane molasses in a mass ratio of 1:(2 - 2.5).

[0010] Furthermore, the inorganic salts include the following components:

[0011] 5 - 6 parts of NH4Cl, 1.3 - 1.4 parts of K2HPO4, 0.2 - 0.3 parts of MgSO4·7H2O, 0.01 - 0.02 parts of CaCl2·2H2O, 0.01 - 0.02 parts of Fe-EDTA.

[0012] Further, the trace element is at least one of ZnSO4, MnCl2, and CuSO4.

[0013] Further, the blueberry peel extract is specifically prepared by the following steps:

[0014] A1. Freeze-dry and crush the blueberry peels to obtain freeze-dried blueberry peel powder. Add it to deionized water, add cellulase to the system, and adjust the pH value of the system to 2 - 3. Treat it at 45 - 50 °C for 30 - 60 minutes, and then concentrate it under vacuum; Add the concentrated system to an ethanol solution, stir for 10 - 15 minutes, then raise the temperature of the system to 45 - 50 °C, perform ultrasonic-assisted extraction and then filter to collect the filtrate;

[0015] A2. Perform ultrafiltration separation on the filtrate obtained in step A1, and then concentrate the separated filtrate under vacuum to obtain the blueberry peel extract.

[0016] Further preferably, the blueberry peel extract is specifically prepared by the following steps:

[0017] A1. Freeze-dry and crush the blueberry peels and sieve them through a 40 - 60 mesh sieve to obtain freeze-dried blueberry peel powder. Add it to deionized water according to a mass ratio of 1 g:10 mL, add cellulase to the system, and use 0.1 mol / L HCl to adjust the pH value of the system to 2 - 3. Treat it at 45 - 50 °C for 30 - 60 minutes, and then concentrate it under vacuum to 1 / 5 of the original volume. Add the concentrated system to a 50 - 70 wt% ethanol solution according to a mass ratio of 1:5, stir at a rate of 30 - 90 rpm at room temperature for 10 - 15 minutes, then raise the temperature of the system to 45 - 50 °C, perform ultrasonic-assisted extraction 3 times, each time for 1 hour, filter and combine the filtrates, and collect the filtrate at the same time;

[0018] A2. Perform ultrafiltration separation on the filtrate obtained in step A1, and then concentrate the separated filtrate under vacuum to 1 / 5 of the original volume to obtain the blueberry peel extract.

[0019] During the above reaction process, the freeze-dried blueberry peel powder is treated with a cellulase solution in an acidic environment, then the system is concentrated and mixed with an ethanol solution to extract the anthocyanin components in the peels, and then a blueberry peel extract containing components such as cellulose nanocrystals and anthocyanins is obtained after ultrafiltration.

[0020] Further, in step A1, the usage amount of the cellulase solution is 40 - 50 FPU / g (compared with the freeze-dried blueberry peel powder).

[0021] Further, in steps A1 and A2, the temperature of vacuum concentration is 30 - 40 °C, and the vacuum degree is -0.08 Mpa.

[0022] Further, in step A2, the ultrafiltration membrane used has a molecular weight cut-off of 100 kDa, and the operating pressure is 0.2 - 0.3 MPa.

[0023] Further, the pretreated cane molasses is specifically prepared by the following steps:

[0024] B1. Treat the cane molasses with cellulase and pectinase to obtain enzymatically hydrolyzed cane molasses;

[0025] B2. Subsequently, remove impurities from the enzymatically hydrolyzed cane molasses by EDTA flocculation to obtain the pretreated cane molasses.

[0026] More preferably, the pretreated cane molasses is specifically prepared by the following steps:

[0027] B1. Dilute the cane molasses to 60 - 70 wt% of the total sugar concentration, treat it with cellulase and pectinase, set the system pH value to 4.5 - 5.0, the treatment temperature to 45 - 55 °C, and the treatment time to 20 - 30 hours to obtain enzymatically hydrolyzed cane molasses;

[0028] B2. Subsequently, add distilled water to the enzymatically hydrolyzed cane molasses at a mass ratio of 1:0.5, stir for 10 - 15 minutes, then add EDTA, set the EDTA concentration in the system to 0.2 - 0.3 (w / v)%, adjust the system pH value to 6.0 - 6.5, stir at a rate of 30 - 90 rpm at room temperature for 10 - 15 hours, centrifuge at a rate of 8000 rpm for 10 - 20 minutes, filter, and collect the supernatant to obtain the pretreated cane molasses.

[0029] During the above reaction process, cellulase and pectinase are used to pretreat the cane molasses to help with its early energy supply. Subsequently, EDTA is used for flocculation to remove impurities, and the heavy metal ions in the cane molasses are complexed by EDTA to eliminate their inhibitory effect on microalgae growth. At the same time, the pH value of the system is adjusted to weakly acidic to reduce the oxidation of phenolic substances in the molasses and maintain the stability of the enzymatic hydrolysis products. When used as a medium component later, it can also better improve the stability of the blueberry peel extract.

[0030] Further, in step B1, the usage amounts of cellulase, pectinase, and cane molasses are (35 - 45) FPU : (15 - 25) FPU : 1 g.

[0031] Further, the system stabilizer is at least one of alginate, humate, EDTA, sugar alcohols, cationic starch, chitosan and its complexes.

[0032] A preparation method of a microalgae medium based on plant extracts specifically includes the following steps:

[0033] Mix glycerol and pretreated molasses according to the mass ratio of the formula parts to obtain a carbon source; add the system stabilizer to sterile seawater, stir at 30 - 90 rpm for 20 - 30 minutes at room temperature, sterilize at 121 °C and then cool to 40 - 50 °C, add the trace elements, inorganic salts and cellulose nanofiber components of the formula parts, cool the system to room temperature, add the blueberry peel extract, continue to stir for 20 - 30 minutes, and store at 0 - 4 °C to obtain a microalgae culture medium based on plant extracts.

[0034] In summary, the present application has the following beneficial effects:

[0035] 1. In the technical solution of the present invention, the blueberry peel extract is used as a component of the microalgae culture medium. The blueberry peel freeze-dried powder is treated with a cellulase solution in an acidic environment, and then the system is concentrated and mixed with an ethanol solution to extract the active ingredients such as anthocyanins in the peel. Subsequently, after ultrafiltration, a blueberry peel extract containing cellulose nanocrystals and anthocyanins and other components is obtained. By coupling the processes in different components, cellulose nanocrystals are extracted using an acidic environment, and at the same time, the extraction amount of cellulose nanocrystals and the stability of active ingredients such as anthocyanins during the extraction process are regulated by the pH value, maintaining its antioxidant and other properties.

[0036] 2. In the technical solution of the present invention, by pretreating cane molasses and then using it together with glycerol as a carbon source, it has a synergistic effect with the blueberry peel extract in promoting the growth of microalgae. On the one hand, the pretreatment step decomposes cane molasses into glucose and fructose components, which can be quickly absorbed by microalgae, significantly improving the absorption efficiency of microalgae for carbon sources and promoting rapid proliferation in the initial stage. Glycerol can be used as a slow-release carbon source in the system, slowly releasing the carbon skeleton through the glycerol kinase pathway to maintain the long-term stable growth of microalgae. After the pretreatment of cane molasses, the early metabolism is concentrated, and reducing sugars and organic acids are more likely to accumulate than before pretreatment, thereby causing the accumulation of reactive oxygen species. The compounded blueberry peel extract can reduce oxidative damage by scavenging free radicals, and the combination of the two can achieve a better microalgae culture effect. On the other hand, glycerol can regulate the osmotic pressure of microalgae cells and relieve high-salt or high-light stress; anthocyanins reduce photoinhibition by stabilizing the thylakoid membrane structure. At the same time, the co-pigmentation effect of anthocyanins and microalgae chlorophyll can enhance the light capture efficiency and improve the synthesis rate of photosynthetic products such as carbohydrates and lipids. The three work together to enhance the survival ability of microalgae under extreme conditions. Detailed implementation mode

[0037] Next, in combination with the embodiments of the present invention, the technical solutions in the embodiments of the present invention will be clearly and completely described. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the protection scope of the present invention.

[0038] All reagents involved in the specific implementation manner of this application are of chemical pure grade. Additionally:

[0039] Cane molasses: provided by Shandong Tianxiang Chemical Co., Ltd.

[0040] Cellulase: hemicellulase 0323, provided by Fujian Shengshi Jiatai Biotechnology Co., Ltd.

[0041] Pectinase: brand is Longkete.

[0042] Cellulose nanofibers: 10 - 50 nm wide, 0.5 - 3 μm long; purity ≥ 99.5%.

[0043] Example 1

[0044] A microalgae culture medium based on plant extracts, comprising the following raw materials in parts by mass:

[0045] 0.5 part of blueberry peel extract, 0.01 part of cellulose nanofibers, 20 parts of carbon source, 1 part of system stabilizer, 6.52 parts of inorganic salts, 0.002 part of trace elements, and 1000 parts of sterile seawater; the carbon source is obtained by mixing glycerol and pretreated cane molasses in a mass ratio of 1:2.

[0046] Among them, the blueberry peel extract is specifically prepared by the following steps:

[0047] A1. Freeze - dry the blueberry peel and then crush it through a 60 - mesh sieve to obtain blueberry peel freeze - dried powder. Add it to deionized water according to a mass ratio of 1 g:10 mL. Add cellulase to the system and adjust the pH value of the system to 2 with 0.1 mol / L HCl. Treat it at 45 °C for 30 minutes, then vacuum - concentrate it to 1 / 5 of the original volume. Add the concentrated system to a 50 wt% ethanol solution according to a mass ratio of 1:5. Stir at a rate of 30 rpm at room temperature for 10 minutes, then raise the system temperature to 45 °C, and perform ultrasonic - assisted extraction 3 times, with an ultrasonic power of 400 W and each time for 1 hour. Filter and combine the filtrates, and collect the filtrates at the same time; the usage amount of the cellulase solution is 40 FPU / g (compared with the blueberry peel freeze - dried powder);

[0048] A2. Ultrafilter the filtrate obtained in step A1. The molecular weight cut-off of the ultrafiltration membrane used is 100 kDa, and the operating pressure is 0.2 MPa. Subsequently, vacuum concentrate the separated filtrate to 1 / 5 of the original volume to obtain blueberry peel extract;

[0049] In steps A1 and A2, the temperature of vacuum concentration is 30 °C and the vacuum degree is -0.08 Mpa.

[0050] The inorganic salts used in the formula include the following components: 5 parts of NH4Cl, 1.3 parts of K2HPO4, 0.2 parts of MgSO4·7H2O, 0.01 parts of CaCl2·2H2O, 0.01 parts of Fe-EDTA.

[0051] The trace element used in the formula is ZnSO4.

[0052] The system stabilizer used in the formula is mannitol.

[0053] The pretreated cane molasses used in the formula is specifically prepared by the following steps:

[0054] B1. Dilute cane molasses to 60 wt% of the total sugar concentration with sterile seawater, and treat it with cellulase and pectinase. Set the system pH value to 4.5, the treatment temperature to 45 °C, and the treatment time to 20 hours to obtain enzymatically hydrolyzed cane molasses; the usage amounts of cellulase, pectinase and cane molasses are 35 FPU: 15 FPU: 1 g;

[0055] B2. Subsequently, add the enzymatically hydrolyzed cane molasses to distilled water according to a mass ratio of 1:0.5, stir for 10 minutes, then add EDTA. Set the EDTA concentration in the system to 0.2 (w / v)%, adjust the system pH value to 6.0, and stir at a rate of 30 rpm at room temperature for 10 hours. Centrifuge at a rate of 8000 rpm for 15 minutes, filter, and collect the supernatant to obtain pretreated cane molasses.

[0056] The preparation method of the aforementioned culture medium specifically includes the following steps:

[0057] Mix glycerol and pretreated molasses according to the mass ratio of the formula parts to obtain a carbon source; add the system stabilizer to sterile seawater, stir at 90 rpm at room temperature for 20 minutes, sterilize at 121 °C and then cool to 40 °C. Add the trace elements, inorganic salts and cellulose nanofiber components of the formula parts to the system, cool the system to room temperature, add blueberry peel extract, continue to stir for 20 minutes, and store at 4 °C to obtain a microalgae culture medium based on plant extracts.

[0058] Example 2

[0059] A microalgae culture medium based on plant extracts, comprising the following raw materials in parts by mass:

[0060] 1 part of blueberry peel extract, 0.03 part of cellulose nanofibers, 30 parts of carbon source, 3 parts of system stabilizer, 7.74 parts of inorganic salts, 0.003 part of trace elements and 1000 parts of sterile seawater; the carbon source is obtained by mixing glycerol and pretreated cane molasses according to a mass ratio of 1:2.3.

[0061] Among them, the blueberry peel extract is specifically prepared by the following steps:

[0062] A1. Freeze-dry the blueberry peel and then crush it through a 60-mesh sieve to obtain freeze-dried blueberry peel powder. Add it to deionized water according to a mass ratio of 1 g:10 mL. Add cellulase to the system and adjust the pH value of the system to 2.5 with 0.1 mol / L HCl. Treat it at 50 °C for 60 minutes, then vacuum concentrate it to 1 / 5 of the original volume. Add the concentrated system to a 60 wt% ethanol solution according to a mass ratio of 1:5. Stir at a rate of 60 rpm at room temperature for 15 minutes, then raise the system temperature to 45 - 50 °C, and perform ultrasonic-assisted extraction 3 times with an ultrasonic power of 400 W for 1 hour each time. Filter and combine the filtrates, and collect the filtrates at the same time; the usage amount of the cellulase solution is 45 FPU / g (compared with the freeze-dried blueberry peel powder);

[0063] A2. Ultrafilter and separate the filtrate obtained in step A1. The molecular weight cut-off of the ultrafiltration membrane used is 100 kDa, and the operating pressure is 0.3 MPa. Then vacuum concentrate the separated filtrate to 1 / 5 of the original volume to obtain the blueberry peel extract;

[0064] In steps A1 and A2, the temperature of vacuum concentration is 35 °C and the vacuum degree is -0.08 Mpa.

[0065] The inorganic salts used in the formula include the following components: 6 parts of NH4Cl, 1.4 parts of K2HPO4, 0.3 part of MgSO4·7H2O, 0.02 part of CaCl2·2H2O, 0.02 part of Fe-EDTA.

[0066] The trace element used in the formula is ZnSO4.

[0067] The system stabilizer used in the formula is mannitol.

[0068] The pretreated cane molasses used in the formula is specifically prepared by the following steps:

[0069] B1. Dilute the cane molasses to a total sugar concentration of 65 wt% with sterile seawater, and treat it with cellulase and pectinase. Set the pH value of the system to 5.0, the treatment temperature to 50 °C, and the treatment time to 24 hours to obtain enzymatically hydrolyzed cane molasses; the usage amounts of cellulase, pectinase and cane molasses are 40 FPU:20 FPU:1 g;

[0070] B2. Subsequently, the enzymatically hydrolyzed cane molasses was added to distilled water at a mass ratio of 1:0.5. After stirring for 15 minutes, EDTA was added, and the concentration of EDTA in the system was set to 0.25 (w / v)%. The pH value of the system was adjusted to 6.0 and stirred at a rate of 90 rpm at room temperature for 12 hours, centrifuged at a rate of 8000 rpm for 15 minutes, filtered, and the supernatant was collected to obtain pretreated cane molasses.

[0071] The preparation method of the aforementioned medium specifically includes the following steps:

[0072] Mix glycerol and pretreated molasses according to the mass ratio of the formula parts to obtain a carbon source; add the system stabilizer to sterile seawater, stir at 90 rpm at room temperature for 25 minutes, sterilize at 121 °C and then cool to 45 °C, add the trace elements, inorganic salts and cellulose nanofiber components of the formula parts, cool the system to room temperature, add blueberry peel extract, continue to stir for 25 minutes, and store at 4 °C to obtain a microalgae medium based on plant extracts.

[0073] Example 3

[0074] A microalgae medium based on plant extracts, comprising the following raw materials in parts by mass:

[0075] 2 parts of blueberry peel extract, 0.05 part of cellulose nanofiber, 40 parts of carbon source, 5 parts of system stabilizer, 7.74 parts of inorganic salts, 0.003 part of trace elements and 1000 parts of sterile seawater; the carbon source is obtained by mixing glycerol and pretreated cane molasses according to a mass ratio of 1:2.5.

[0076] Among them, the blueberry peel extract is specifically prepared by the following steps:

[0077] A1. Freeze-dry the blueberry peel and crush it through a 60-mesh sieve to obtain blueberry peel freeze-dried powder. Add it to deionized water at a mass ratio of 1 g:10 mL. Add cellulase to the system and adjust the pH value of the system to 3 with 0.1 mol / L HCl. Treat it at 50 °C for 60 minutes, then vacuum concentrate to 1 / 5 of the original volume. Add the concentrated system to a 70 wt% ethanol solution at a mass ratio of 1:5. Stir at a rate of 90 rpm at room temperature for 15 minutes, then raise the system temperature to 50 °C, and perform ultrasonic-assisted extraction 3 times with an ultrasonic power of 400 W for 1 hour each time. Filter and combine the filtrates, and collect the filtrates at the same time; the usage amount of the cellulase solution is 50 FPU / g (compared with the blueberry peel freeze-dried powder);

[0078] A2. Ultrafiltration separation was performed on the filtrate obtained in step A1. The molecular weight cut-off of the ultrafiltration membrane used was 100 kDa, and the operating pressure was 0.3 MPa. Then, the separated filtrate was vacuum concentrated to 1 / 5 of the original volume to obtain blueberry peel extract;

[0079] In steps A1 and A2, the temperature of vacuum concentration is 40°C and the vacuum degree is -0.08 Mpa.

[0080] The inorganic salts used in the formula include the following components: 6 parts of NH4Cl, 1.4 parts of K2HPO4, 0.3 part of MgSO4·7H2O, 0.02 part of CaCl2·2H2O, and 0.02 part of Fe-EDTA.

[0081] The trace element used in the formula is CuSO4.

[0082] The system stabilizer used in the formula is sodium alginate (9005-38-3, 99%).

[0083] The pretreated cane molasses used in the formula is specifically prepared by the following steps:

[0084] B1. Dilute the cane molasses with sterile seawater to 70 wt% of the total sugar concentration, treat it with cellulase and pectinase, set the system pH value to 5.0, the treatment temperature to 55°C, and the treatment time to 30 hours to obtain enzymatically hydrolyzed cane molasses; the usage amounts of cellulase, pectinase, and cane molasses are 45 FPU: 25 FPU: 1 g;

[0085] B2. Subsequently, add distilled water to the enzymatically hydrolyzed cane molasses according to a mass ratio of 1:0.5, stir for 15 minutes, then add EDTA, set the EDTA concentration in the system to 0.3 (w / v)%, adjust the system pH value to 6.5, stir at a rate of 90 rpm at room temperature for 15 hours, centrifuge at a rate of 8000 rpm for 20 minutes, filter, and collect the supernatant to obtain the pretreated cane molasses.

[0086] The preparation method of the foregoing culture medium specifically includes the following steps:

[0087] Mix glycerol and pretreated molasses according to the mass ratio of the formula parts to obtain a carbon source; add the system stabilizer to sterile seawater, stir at 90 rpm at room temperature for 30 minutes, sterilize at 121°C and then cool to 50°C, add the trace element, inorganic salt, and cellulose nanofiber components of the formula parts, cool the system to room temperature, add blueberry peel extract, continue to stir for 30 minutes, and store at 4°C to obtain a microalgae culture medium based on plant extracts.

[0088] Comparative Example 1

[0089] The difference between this comparative example and Example 1 is that in step A1, the system pH value is set to 5.0.

[0090] Comparative Example 2

[0091] The difference between this comparative example and Example 1 is that in step A1, the system pH value is set to 7.0.

[0092] Comparative Example 3

[0093] The difference between this comparative example and Example 1 is that in step A1, the pH value of the system is set to 8.0.

[0094] Comparative Example 4

[0095] The difference between this comparative example and Example 1 is that the blueberry peel extract in this comparative example is specifically prepared by the following steps:

[0096] A1. Freeze-dry the blueberry peel and then crush it through a 60-mesh sieve to obtain freeze-dried powder of blueberry peel. Add it to a 50 wt% ethanol solution according to a mass ratio of 1 g:10 mL, stir at a rate of 30 rpm at room temperature for 10 minutes, then raise the system temperature to 45°C, perform ultrasonic-assisted extraction 3 times with an ultrasonic power of 400 W for 1 hour each time, filter and combine the filtrates, and collect the filtrates at the same time;

[0097] A2. Perform nanofiltration separation on the filtrate obtained in step A1. The molecular weight cut-off of the nanofiltration membrane used is 300 Da, and the operating pressure is 0.4 MPa. Then vacuum-concentrate the separated filtrate to 1 / 5 of the original volume to obtain the blueberry peel extract.

[0098] Comparative Example 5

[0099] The difference between this comparative example and Example 1 is that glycerol is used instead of pretreated cane molasses as the carbon source in this comparative example.

[0100] Comparative Example 6

[0101] The difference between this comparative example and Example 1 is that cane molasses is used instead of pretreated cane molasses in this comparative example.

[0102] Comparative Example 7

[0103] The difference between this comparative example and Example 1 is that the pretreated cane molasses in this comparative example is specifically prepared by the following steps:

[0104] Dilute the cane molasses to 60 wt% of the total sugar concentration with sterile seawater, treat it with cellulase and pectinase, set the pH value of the system to 4.5, the treatment temperature to 45°C, and the treatment time to 20 hours to obtain the pretreated cane molasses; the usage amounts of cellulase, pectinase, and cane molasses are 35 FPU:15 FPU:1 g.

[0105] Comparative Example 8

[0106] The difference between this comparative example and Example 1 is that the pretreated cane molasses in this comparative example is specifically prepared by the following steps:

[0107] Dilute cane molasses to a total sugar concentration of 60 wt% using sterile seawater, stir for 10 minutes, add EDTA, set the EDTA concentration in the system to 0.2 (w / v)%, adjust the pH value of the system to 6.0, and stir at a rate of 30 rpm at room temperature for 10 hours. Centrifuge at a rate of 8000 rpm for 15 minutes, filter, collect the supernatant, and obtain pretreated cane molasses.

[0108] Performance test

[0109] Now perform performance tests on the microalgae culture media prepared in Examples 1-3 and Comparative Examples 1-8 of this application.

[0110] System stability test:

[0111] Store the microalgae culture media prepared in different groups at 30 °C and a light intensity of 8000 lux for 12, 24, and 48 hours, and measure the changes in the pH value and the change rate of anthocyanin content after storage of the system. The specific test results are shown in Table 1 below:

[0112] Table 1

[0113]

[0114]

[0115] As shown by the results in Table 1 above: The microalgae culture media prepared in the three examples showed better stability, with only minor changes in the pH value and less degradation rate of anthocyanin. From the results in Comparative Examples 1-3, it can be seen that at the more suitable pH value given in the examples, the components in the blueberry peel extract can be better optimized, and the stability of anthocyanin is better. From the results in Comparative Example 4, it can be seen that using a suitable extraction method can regulate the components of the extract, and the anthocyanin content increases under a small cut-off volume, but the stability decreases. From the results in Comparative Examples 5-8, it can be seen that using pretreated cane molasses and glycerol as a composite carbon source can improve the stability of each component in the system, which is beneficial to reducing the system changes caused by carbon source decomposition.

[0116] Now detect the cultivation effects of the microalgae culture media prepared in Examples 1-3 and Comparative Examples 1-8. Taking Chaetoceros simplex, Pavlova viridis, and Platymonas sp. as examples, use the culture media prepared in different groups to culture the above microalgae respectively. The cultivation light intensity is 5000 lux, the cultivation temperature is 25 ± 1 °C, aerate 3 times a day, supplement the culture media once every 3 days, and the supplement amount is 10 wt% of the initial culture media. Harvest after 15 days of cultivation, and measure the concentration and biomass of the microalgae in different groups to judge the cultivation effects of different culture media. The specific detection results are shown in Table 2.

[0117] Table 2

[0118]

[0119]

[0120] Among the three strains selected for the above tests, the optimal growth light intensity of Chaetoceros unicornis is lower than the set test light intensity, the optimal growth light intensity of Platymonas subcordiformis is higher than the set test light intensity, and the set test light intensity is suitable for Pavlova viridis. It can be seen from the results in Table 2 that the algae cultivation effects in several embodiments are better than the samples in the comparative examples, among which the biomass of Platymonas subcordiformis is the highest. It can be seen from the results in the comparative examples that by pretreating cane molasses and then using it together with glycerol as a carbon source, it has a synergistic effect with blueberry peel extract in promoting the growth of microalgae.

[0121] In the description of the specification, the description with reference to terms such as "one embodiment", "example", "specific example", etc. means that the specific features, structures, materials or characteristics described in connection with the embodiment or example are included in at least one embodiment or example of the present invention. In this specification, the schematic representations of the above terms do not necessarily refer to the same embodiment or example. Moreover, the specific features, structures, materials or characteristics described can be combined in a suitable manner in any one or more embodiments or examples.

[0122] The above content is only an example and illustration of the present invention. Those skilled in the art of the present technology can make various modifications or supplements to the described specific embodiments or use similar methods to replace them, as long as they do not deviate from the invention or exceed the scope defined by the claims of the present invention, they should all belong to the protection scope of the present invention.

Claims

1. A microalgae culture medium based on plant extracts, characterized in that, It includes the following raw materials in parts by mass: 0.5 - 2 parts of blueberry peel extract, 0.01 - 0.05 parts of cellulose nanofibers, 20 - 40 parts of carbon source, 1 - 5 parts of system stabilizer, 6.52 - 7.74 parts of inorganic salts, 0.002 - 0.003 parts of trace elements, and 1000 parts of sterile seawater; The carbon source is obtained by mixing glycerol and pretreated cane molasses in a mass ratio of 1:(2 - 2.5).

2. The microalgae culture medium based on plant extracts according to claim 1, characterized in that, The inorganic salts include the following components: 5 - 6 parts of NH4Cl, 1.3 - 1.4 parts of K2HPO4, 0.2 - 0.3 parts of MgSO4·7H2O, 0.01 - 0.02 parts of CaCl2·2H2O, 0.01 - 0.02 parts of Fe-EDTA.

3. The microalgae culture medium based on plant extracts according to claim 1, wherein, The blueberry peel extract is specifically prepared by the following steps: A1. Freeze-dry and crush the blueberry peel to obtain blueberry peel freeze-dried powder, add it to deionized water, add cellulase to the system, and adjust the pH value of the system to 2 - 3. Treat it at 45 - 50 °C for 30 - 60 minutes, and then concentrate it under vacuum; add the concentrated system to an ethanol solution, stir for 10 - 15 minutes, then raise the temperature of the system to 45 - 50 °C, perform ultrasonic-assisted extraction and then filter to collect the filtrate; A2. Ultrafilter and separate the filtrate obtained in step A1, and then concentrate the separated filtrate under vacuum to obtain blueberry peel extract.

4. The microalgae culture medium based on plant extracts according to claim 3, characterized in that, In step A1, the usage amount of the cellulase solution is 40 - 50 FPU / g.

5. A microalgae culture medium based on plant extracts according to claim 3, wherein, In steps A1 and A2, the temperature of vacuum concentration is 30 - 40 °C.

6. The microalgae culture medium based on plant extracts according to claim 3, characterized in that, In step A2, the molecular weight cut-off of the ultrafiltration membrane used is 100 kDa, and the operating pressure is 0.2 - 0.3 MPa.

7. A microalgae culture medium based on plant extracts according to claim 1, characterized in that, The pretreated cane molasses is specifically prepared by the following steps: B1. Treat cane molasses with cellulase and pectinase to obtain enzymatically hydrolyzed cane molasses; B2. Then remove impurities from the enzymatically hydrolyzed cane molasses by EDTA flocculation to obtain pretreated cane molasses.

8. A microalgae culture medium based on plant extracts according to claim 7, characterized in that, In step B1, the usage amounts of cellulase, pectinase, and cane molasses are (35 - 45) FPU:(15 - 25) FPU:1 g.

9. A microalgae culture medium based on plant extracts according to claim 1, wherein, The system stabilizer is at least one of alginate, humate, EDTA, sugar alcohol substances, cationic starch, chitosan substances, etc.

10. A method for preparing a microalgae culture medium based on plant extracts according to any one of claims 1-9, characterized in that, Specifically, it includes the following steps: Mix glycerol and pretreated molasses according to the mass ratio of the formula parts to obtain the carbon source; add the system stabilizer to sterile seawater, stir for 20 - 30 minutes, sterilize and then cool to 40 - 50 °C, add the trace elements, inorganic salts, and cellulose nanofiber components in the formula parts, cool the system to room temperature, add blueberry peel extract, continue to stir for 20 - 30 minutes, and store at 0 - 4 °C to obtain a microalgae culture medium based on plant extracts.

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