A method for preparing a bio-flocculating agent

Abstract

This invention belongs to the field of Baijiu (Chinese liquor) brewing and relates to a method for preparing a bioflocculator, comprising the following steps: S1, expansion culture: adding the seed liquid of flocculating microorganisms to a culture medium containing brewing organic wastewater for expansion culture to obtain a bioflocculating bacterial solution; S2, extraction: extracting the bioflocculating bacterial solution using a combination of freeze-thaw and acidification methods to obtain the bioflocculator. Using high-concentration organic wastewater from brewing to prepare a flocculating culture medium can rapidly obtain MBF, realizing the resource utilization of high-concentration wastewater while simultaneously obtaining MBF with high flocculation effect.

Description

Technical Field

[0001] This invention belongs to the field of liquor brewing technology and relates to a method for preparing a biological flocculant. Background Technology

[0002] Microbial flocculants (MBF) are natural organic polymers secreted by microorganisms that have flocculation activity. They can cause non-degradable solid suspended particles and colloidal particles to coagulate and precipitate. They are biodegradable, safe, efficient, and do not cause secondary pollution, making them a green water treatment agent.

[0003] However, most MBF preparation processes are currently quite complex. First, microorganisms capable of producing flocculation functions need to be screened, and then purified and cultured. The whole process is time-consuming. Furthermore, the culture of MBF-producing bacteria usually uses glucose, galactose, starch, etc. as organic carbon sources and yeast extract, beef extract, peptone, etc. as organic nitrogen sources, resulting in high MBF production costs.

[0004] In addition, the carbon and nitrogen sources for preparing biological flocculants currently include molasses wastewater, washing wastewater, sugarcane bagasse, biohydrogen production wastewater, straw powder, pig manure, and malt roots. These organic wastewaters with high carbon and nitrogen content can be used as culture media for MBF production, which can not only greatly reduce the production cost of MBF, but also make wastewater resources. However, some pretreatment of the wastewater is required to make it easier for floc-producing bacteria to absorb and utilize it, and at the same time obtain MBF with better flocculation effect.

[0005] Cellar bottom water and pot bottom water are high-concentration organic wastewater generated during the brewing of baijiu (Chinese liquor). Belonging to the category of organic wastewater from the food processing industry, they are rich in starch, reducing sugars, organic acids, esters, and alcohols. These substances are difficult to degrade, and direct discharge into wastewater treatment plants not only increases the difficulty of wastewater treatment but also wastes resources. However, this wastewater contains high levels of organic carbon, nitrogen, and various inorganic salts. Utilizing the nutrients in this high-concentration organic wastewater, rich in nutrients required for microbial growth and metabolism, as a flocculant culture medium can not only enable resource utilization of the wastewater but also reduce the production cost of microbial flocculants.

[0006] Currently, there are no reported methods for rapidly preparing MBF using high-concentration organic wastewater from brewing (cellar water, pot bottom water). Therefore, using high-concentration organic wastewater from brewing as an inexpensive substrate for the preparation of microbial flocculants is of great significance for promoting resource utilization and reducing pollution. Summary of the Invention

[0007] The purpose of this invention is to provide a method for preparing a bioflocculant, which uses high-concentration organic wastewater from brewing to prepare a flocculant-generating culture medium, thereby rapidly obtaining a bioflocculant (MBF). This method enables the resource utilization of high-concentration wastewater while simultaneously obtaining a bioflocculant (MBF) with high flocculation effect, reducing the production cost and time of the bioflocculant (MBF).

[0008] In a first aspect, the present invention provides a method for preparing a bioflocculant, comprising the following steps:

[0009] S1. Expanded culture: The seed liquid of flocculating microorganisms is added to a culture medium containing brewing organic wastewater for expanded culture to obtain bioflocculating bacterial liquid;

[0010] S2. Extraction: The bioflocculating bacterial solution is extracted using a combination of freeze-thaw and acidification methods to obtain the bioflocculating agent.

[0011] In some implementations, in step S2, the freeze-thaw process includes freezing with liquid nitrogen and then thawing with a constant-temperature water bath.

[0012] Preferably, the volume ratio of the liquid nitrogen to the bioflocculant liquid is (45-55):(8-12);

[0013] Preferably, the temperature of the constant temperature water bath is 28-35℃.

[0014] In some embodiments, in step S2, the acidification includes acidification extraction using hydrochloric acid;

[0015] Preferably, the concentration of the hydrochloric acid is 10-30%;

[0016] Preferably, the volume ratio of the hydrochloric acid to the bioflocculant liquid is (4-6):(8-12);

[0017] Preferably, the hydrochloric acid acidification treatment time is 4-6 minutes.

[0018] In some embodiments, in step S1, the floc-forming microorganisms are screened from activated sludge and / or aerobic sludge from a brewery wastewater treatment plant.

[0019] Preferably, the activated sludge is fresh sediment or suspended solids separated from the secondary sedimentation tank of a brewing wastewater treatment plant.

[0020] Preferably, the aerobic sludge is: biological contact oxidation-oxidation method A from a brewery wastewater treatment plant. 2 O 2 The mud-water mixture obtained from the secondary aerobic tank in the process section.

[0021] In some embodiments, the screening method includes: weighing fresh activated sludge and / or aerobic sludge into sterile distilled water, shaking, taking the supernatant and adding it to a culture medium, and then shaking and culturing to obtain the seed liquid of the floc-producing microorganisms;

[0022] Preferably, the volume ratio of the supernatant to the culture medium is (8-12):(145-155);

[0023] Preferably, the oscillation speed is 145-155 r / min; the oscillation time is 3-5 h;

[0024] Preferably, the conditions for the shaking culture are: 28-32℃, shaking speed of 140-150 r / min, and culture for 35-38 h;

[0025] Preferably, when the sludge is called activated sludge, the culture medium is Gao's No. 1 liquid culture medium;

[0026] Preferably, when the sludge is aerobic sludge, the culture medium is LB medium.

[0027] In some implementations, in step S1, the brewing organic wastewater includes at least one of cellar bottom water and pot bottom water;

[0028] Preferably, the conditions of the bottom water in the pit include: pH 4-5; COD content 150,000-160,000 mg / L; BOD5 content 37,000-38,000 mg / L; NH3-N content 3,000-4,000 mg / L; TP content 2,000-2,500 mg / L; and TN content 7,000-8,000 mg / L.

[0029] Preferably, the conditions of the water at the bottom of the pot include: pH 3-4; COD content 5000-6000 mg / L; BOD5 content 1500-1600 mg / L;

[0030] In some embodiments, in step S1, the culture medium includes at least one of: a culture medium diluted with water from the bottom of a pit, a culture medium diluted with water from the bottom of a pot, a culture medium diluted with water from the bottom of a pit, and a culture medium derived from water from the bottom of a pot.

[0031] Preferably, the cellar bottom water dilution culture medium is a culture medium obtained by diluting the cellar bottom water;

[0032] Preferably, the dilution ratio of the water at the bottom of the pit is 3-5 times;

[0033] Preferably, the COD concentration in the culture medium diluted by the bottom water is 4000-6000 mg / L;

[0034] Preferably, the bottom water culture medium is made by directly using the bottom water of the pot as the culture medium;

[0035] Preferably, the COD concentration in the bottom water culture medium is 5500-5800 mg / L;

[0036] The exogenous culture medium diluted with the pit bottom water is obtained by adding additional nutrients to the culture medium diluted with the pit bottom water.

[0037] The exogenous culture medium for the bottom water of the pot is obtained by adding nutrients to the culture medium for the bottom water of the pot.

[0038] Preferably, the nutrient includes at least one of a carbon source, a nitrogen source, and an inorganic element;

[0039] Preferably, the nutrients include at least one of glucose, KH2PO4, K2HPO4, MgSO4, NaCl, and urea;

[0040] Preferably, the nutrients include: glucose 8-12 g / L, KH2PO4 1-5 g / L, K2HPO4 2-7 g / L, MgSO4 0.1-0.5 g / L, NaCl 0.5-3 g / L, and urea 0.1-2 g / L.

[0041] In some implementations, the scaling-up culture conditions in step S1 are: temperature 28-32℃, rotation speed 125-135r / min, and time 30-40h.

[0042] In some embodiments, in step S1, the volume ratio of the flocculating microbial seed solution to the culture medium is (3-8):100.

[0043] In a second aspect, the present invention provides a bioflocculant, which is prepared by any of the methods described above for preparing bioflocculants.

[0044] In summary, this application includes at least one of the following beneficial technical effects:

[0045] (1) By using water from the bottom of the pit and water from the bottom of the pot as inexpensive substrate culture medium to expand the culture of the selected bacterial liquid with flocculation activity, bio-flocculation agent (MBF) can be obtained quickly, realizing the resource utilization of high-concentration wastewater and obtaining bio-flocculation agent (MBF) with high flocculation effect, thereby reducing the production cost and production time of bio-flocculation agent (MBF).

[0046] (2) Using the water at the bottom of the cellar and the water at the bottom of the pot as inexpensive substrate culture medium, the selected bacterial liquid with flocculation activity was cultured in a large scale. The resulting microbial flocculant had a good flocculation effect, with a flocculation rate of over 87% and an average of 88.90%, which is 2.66% higher than the flocculation rate of the microbial flocculant obtained by culturing with traditional laboratory culture medium. The cost of preparing inexpensive substrate culture medium is low, and at the same time, the resource utilization of high-concentration organic wastewater from brewing is realized.

[0047] (3) The flocculant obtained by the extraction method of freeze-thaw combined with hydrochloric acid treatment has a better flocculation effect. Detailed Implementation

[0048] The following specific embodiments further illustrate the technical solution of the present invention. These specific embodiments do not represent a limitation on the scope of protection of the present invention. Non-essential modifications and adjustments made by others based on the concept of the present invention still fall within the scope of protection of the present invention.

[0049] 1. The culture medium preparation method of the present invention is as follows:

[0050] Preparation of LB medium: Weigh 10g of peptone, 5g of yeast powder, and 3g of sodium chloride, and add 1L of deionized water;

[0051] Preparation of PDA medium: Weigh 20g of glucose and 6g of potato flour, and add 1L of deionized water;

[0052] Preparation of YPD medium: Weigh 5g of yeast powder, 10g of peptone, and 10g of glucose, and add 1L of deionized water;

[0053] Preparation of Gao's No. 1 liquid culture medium: soluble starch (20g), KNO3 (1g), K2HPO4 (0.5g), MgSO4·7H2O (0.5g), NaCl (0.5g), FeSO4·7H2O (0.01g), agar 20g, pH=7.4-7.6.

[0054] 2. The relevant water quality indicators of the pit bottom water and pot bottom water of this invention were measured with reference to the "Surface Water Environmental Quality Standard" (GB3838-2002), and the results are as follows:

[0055] Table 1. Water quality characteristics of bottom water and cellar water (mg / L)

[0056] PH COD <![CDATA[BOD5]]> <![CDATA[NH3-N]]> TP TN Water at the bottom of the cellar 4.65 154352 37142 3440 2130 7892 Bottom water 3.26 5269 1592 15.5 18 46.3

[0057] 3. The "activated sludge" used in the embodiments of the present invention refers to the fresh sediment or suspended solids discharged from the secondary sedimentation tank of the brewing wastewater treatment plant after sedimentation. It is a residual activated sludge with a water content of about 80% and is brown.

[0058] The "aerobic sludge granules" used in this embodiment of the invention refer to: aerobic sludge granules from a brewing wastewater treatment plant are produced using the biological contact oxidation-oxidation method A in brewing wastewater treatment plants. 2 O 2 The mud-water mixture obtained from the secondary aerobic tank in the process section has a water content of 95%.

[0059] Example 1: Activated sludge + Gao's No. 1 liquid culture medium + pit bottom water diluted 3 times

[0060] A method for rapidly preparing bioflocculants from high-concentration organic wastewater from brewing includes the following steps:

[0061] S1. Expanded culture:

[0062] S1.1 Screening for microorganisms with flocculation activity: Microorganisms with flocculation activity were screened from the activated sludge of the brewing wastewater treatment plant (in other embodiments, the source of flocculating microorganisms may also be commercially available or other channels). The specific screening method is as follows: First, weigh soluble starch (20g), KNO3 (1g), K2HPO4 (0.5g), MgSO4·7H2O (0.5g), NaCl (0.5g), FeSO4·7H2O (0.01g), and agar (20g). A Gaoshi No. 1 liquid culture medium with a pH of 7.4 was prepared (in other embodiments, the Gaoshi No. 1 liquid culture medium may also be obtained from commercial purchase or other channels); then 10g of fresh activated sludge was weighed into a conical flask containing 90ml of sterile distilled water, and after shaking on a shaker for 4h at a shaking speed of 150r / min, 10ml of the supernatant was added to 150ml of Gaoshi No. 1 liquid culture medium, and placed at 30℃ and shaken at 145r / min for 36h to obtain the bacterial solution of floc-producing microorganisms;

[0063] S1.2 Cultivation: Dilute the bottom water of the brewing cellar three times and take 1L for sterilization to obtain the bottom water dilution culture medium; then add the bacterial solution of 5% flocculent microorganisms to the bottom water dilution culture medium for expansion culture. The culture conditions are 30℃, 130r / min, and 36h to obtain the bioflocculation bacterial solution.

[0064] S2. Extraction: Adjust the pH of the bioflocculating bacterial solution to about 7, sterilize it in a high-temperature sterilizer at 121℃ for 21 minutes, take 10 ml of the bioflocculating bacterial solution, freeze it with 50 ml of liquid nitrogen, thaw it in a constant temperature water bath at 30℃, and then add 5 ml of 20% hydrochloric acid for acidification treatment for 5 minutes to obtain bioflocculating agent 1.

[0065] Example 2: Activated sludge + Gao's No. 1 liquid culture medium + pit bottom water diluted 5 times

[0066] A method for rapidly preparing bioflocculants from high-concentration organic wastewater from brewing includes the following steps:

[0067] S1. Expanded culture:

[0068] S1.1 Screening for microorganisms with flocculation activity: Microorganisms with flocculation activity were screened from the activated sludge of the brewing wastewater treatment plant (in other embodiments, the source of flocculating microorganisms may also be commercially available or other channels). The specific screening method is as follows: First, weigh soluble starch (20g), KNO3 (1g), K2HPO4 (0.5g), MgSO4·7H2O (0.5g), NaCl (0.5g), FeSO4·7H2O (0.01g), and agar (20g). A Gaoshi No. 1 liquid culture medium with a pH of 7.4 was prepared (in other embodiments, the Gaoshi No. 1 liquid culture medium may also be obtained from commercial purchase or other channels); then 10g of fresh activated sludge was weighed into a conical flask containing 90ml of sterile distilled water, and after shaking on a shaker for 4h at a shaking speed of 150r / min, 10ml of the supernatant was added to 150ml of Gaoshi No. 1 liquid culture medium, and placed at 30℃ and shaken at 145r / min for 36h to obtain the bacterial solution of floc-producing microorganisms;

[0069] S1.2 Cultivation: Dilute the bottom water of the brewing cellar by 5 times and take 1L for sterilization to obtain the bottom water dilution culture medium; then add the bacterial solution of 5% flocculent microorganisms to the bottom water dilution culture medium for expansion culture. The culture conditions are 30℃, 130r / min, and 36h to obtain the bioflocculation bacterial solution.

[0070] S2. Extraction: Adjust the pH of the bioflocculating bacterial solution to about 7, sterilize it in a high-temperature sterilizer at 121℃ for 21 minutes, take 10 ml of the bioflocculating bacterial solution, freeze it with 50 ml of liquid nitrogen, thaw it in a constant temperature water bath at 30℃, and then add 5 ml of hydrochloric acid with a mass fraction of 20% for acidification treatment for 5 minutes to obtain bioflocculating agent 2.

[0071] Example 3: Activated sludge + Gao's No. 1 liquid culture medium + pit bottom water diluted 3 times + exogenous culture medium

[0072] A method for rapidly preparing bioflocculants from high-concentration organic wastewater from brewing includes the following steps:

[0073] S1. Expanded culture:

[0074] S1.1 Screening for microorganisms with flocculation activity: Microorganisms with flocculation activity were screened from the activated sludge of the brewing wastewater treatment plant (in other embodiments, the source of flocculating microorganisms may also be commercially available or other channels). The specific screening method is as follows: First, weigh soluble starch (20g), KNO3 (1g), K2HPO4 (0.5g), MgSO4·7H2O (0.5g), NaCl (0.5g), FeSO4·7H2O (0.01g), and agar (20g). A Gaoshi No. 1 liquid culture medium with a pH of 7.4 was prepared (in other embodiments, the Gaoshi No. 1 liquid culture medium may also be obtained from commercial purchase or other channels); then 10g of fresh activated sludge was weighed into a conical flask containing 90ml of sterile distilled water, and after shaking on a shaker for 4h at a shaking speed of 150r / min, 10ml of the supernatant was added to 150ml of Gaoshi No. 1 liquid culture medium, and placed at 30℃ and shaken at 145r / min for 36h to obtain the bacterial solution of floc-producing microorganisms;

[0075] S1.2 Cultivation: Dilute the bottom water of the brewing cellar three times, take 1L and add 10g of glucose, 2g of KH2PO4, 5g of K2HPO4, 0.2g of MgSO4, 1g of NaCl and 0.5g of urea, and then sterilize to obtain the cellar bottom water dilution + exogenous culture medium; then add the 5% ratio of the flocculating microbial bacterial solution to the cellar bottom water dilution + exogenous culture medium for large-scale cultivation. The cultivation conditions are 30℃, 130r / min, and 36h to obtain the bioflocculating bacterial solution;

[0076] S2. Extraction: Adjust the pH of the bioflocculating bacterial solution to about 7, sterilize it in a high-temperature sterilizer at 121℃ for 21 minutes, take 10 ml of the bioflocculating bacterial solution, freeze it with 50 ml of liquid nitrogen, thaw it in a constant temperature water bath at 30℃, and then add 5 ml of hydrochloric acid with a mass fraction of 20% for acidification treatment for 5 minutes to obtain bioflocculating agent 3.

[0077] Example 4: Activated sludge + Gao's No. 1 liquid culture medium + pit bottom water diluted 5 times + exogenous culture medium

[0078] A method for rapidly preparing bioflocculants from high-concentration organic wastewater from brewing includes the following steps:

[0079] S1. Expanded culture:

[0080] S1.1 Screening for microorganisms with flocculation activity: Microorganisms with flocculation activity were screened from the activated sludge of the brewing wastewater treatment plant (in other embodiments, the source of flocculating microorganisms may also be commercially available or other channels). The specific screening method is as follows: First, weigh soluble starch (20g), KNO3 (1g), K2HPO4 (0.5g), MgSO4·7H2O (0.5g), NaCl (0.5g), FeSO4·7H2O (0.01g), and agar (20g). A Gaoshi No. 1 liquid culture medium with a pH of 7.4 was prepared (in other embodiments, the Gaoshi No. 1 liquid culture medium may also be obtained from commercial purchase or other channels); then 10g of fresh activated sludge was weighed into a conical flask containing 90ml of sterile distilled water, and after shaking on a shaker for 4h at a shaking speed of 150r / min, 10ml of the supernatant was added to 150ml of Gaoshi No. 1 liquid culture medium, and placed at 30℃ and shaken at 145r / min for 36h to obtain the bacterial solution of floc-producing microorganisms;

[0081] S1.2 Cultivation: Dilute the bottom water of the brewing cellar 5 times, take 1L and add 10g glucose, 2g KH2PO4, 5g K2HPO4, 0.2g MgSO4, 1g NaCl and 0.5g urea, then sterilize to obtain the cellar bottom water dilution + exogenous culture medium; then add the 5% ratio of the flocculating microbial bacterial solution to the cellar bottom water dilution + exogenous culture medium for large-scale cultivation at 30℃, 130r / min for 36h to obtain the bioflocculating bacterial solution;

[0082] S2. Extraction: Adjust the pH of the bioflocculating bacterial solution to about 7, sterilize it in a high-temperature sterilizer at 121℃ for 21 minutes, take 10 ml of the bioflocculating bacterial solution, freeze it with 50 ml of liquid nitrogen, thaw it in a constant temperature water bath at 30℃, and then add 5 ml of hydrochloric acid with a mass fraction of 20% for acidification treatment for 5 minutes to obtain bioflocculating agent 4.

[0083] Example 5: Aerobic granules + LB medium + water from the bottom of the pit, diluted 3 times.

[0084] A method for rapidly preparing bioflocculants from high-concentration organic wastewater from brewing includes the following steps:

[0085] S1. Expanded culture:

[0086] S1.1 Screening for microorganisms with flocculation activity: Microorganisms with flocculation activity were screened from the aerobic sludge particles of the brewing wastewater treatment plant (in other embodiments, the source of flocculating microorganisms may also be commercially available or other channels). The specific screening method is as follows: First, weigh 10g of peptone, 5g of yeast powder, and 3g of sodium chloride, and add 1L of deionized water to prepare LB medium (in other embodiments, the source of LB medium may also be commercially available or other channels); then take 10ml of the aerobic sludge-water mixture and add it to 150ml of LB medium, place it at 30℃, and shake at 145r / min for 36h to obtain the bacterial solution of flocculating microorganisms;

[0087] S1.2 Cultivation: Dilute the bottom water of the brewing cellar three times and take 1L for sterilization to obtain the bottom water dilution culture medium; then add the bacterial solution of 5% flocculent microorganisms to the bottom water dilution culture medium for expansion culture. The culture conditions are 30℃, 130r / min, and 36h to obtain the bioflocculation bacterial solution.

[0088] S2. Extraction: Adjust the pH of the bioflocculating bacterial solution to about 7, sterilize it in a high-temperature sterilizer at 121℃ for 21 minutes, take 10 ml of the bioflocculating bacterial solution, freeze it with 50 ml of liquid nitrogen, thaw it in a constant temperature water bath at 30℃, and then add 5 ml of hydrochloric acid with a mass fraction of 20% for acidification treatment for 5 minutes to obtain bioflocculating agent 5.

[0089] Example 6: Aerobic granules + LB medium + pit bottom water diluted 5 times

[0090] A method for rapidly preparing bioflocculants from high-concentration organic wastewater from brewing includes the following steps:

[0091] S1. Expanded culture:

[0092] S1.1 Screening for microorganisms with flocculation activity: Microorganisms with flocculation activity were screened from the aerobic sludge particles of the brewing wastewater treatment plant (in other embodiments, the source of flocculating microorganisms may also be commercially available or other channels). The specific screening method is as follows: First, weigh 10g of peptone, 5g of yeast powder, and 3g of sodium chloride, and add 1L of deionized water to prepare LB medium (in other embodiments, the source of LB medium may also be commercially available or other channels); then take 10ml of the aerobic sludge-water mixture and add it to 150ml of LB medium, place it at 30℃, and shake at 145r / min for 36h to obtain the bacterial solution of flocculating microorganisms;

[0093] S1.2 Cultivation: Dilute the bottom water of the brewing cellar by 5 times and take 1L for sterilization to obtain the bottom water dilution culture medium; then add the bacterial solution of 5% flocculent microorganisms to the bottom water dilution culture medium for expansion culture. The culture conditions are 30℃, 130r / min, and 36h to obtain the bioflocculation bacterial solution.

[0094] S2. Extraction: Adjust the pH of the bioflocculating bacterial solution to about 7, sterilize it in a high-temperature sterilizer at 121℃ for 21 minutes, take 10 ml of the bioflocculating bacterial solution, freeze it with 50 ml of liquid nitrogen, thaw it in a constant temperature water bath at 30℃, and then add 5 ml of hydrochloric acid with a mass fraction of 20% for acidification treatment for 5 minutes to obtain bioflocculating agent 6.

[0095] Example 7: Aerobic granules + LB medium + water from the bottom of the pit diluted 3 times + exogenous culture medium

[0096] A method for rapidly preparing bioflocculants from high-concentration organic wastewater from brewing includes the following steps:

[0097] S1. Expanded culture:

[0098] S1.1 Screening for microorganisms with flocculation activity: Microorganisms with flocculation activity were screened from the aerobic sludge particles of the brewing wastewater treatment plant (in other embodiments, the source of flocculating microorganisms may also be commercially available or other channels). The specific screening method is as follows: First, weigh 10g of peptone, 5g of yeast powder, and 3g of sodium chloride, and add 1L of deionized water to prepare LB medium (in other embodiments, the source of LB medium may also be commercially available or other channels); then take 10ml of the aerobic sludge-water mixture and add it to 150ml of LB medium, place it at 30℃, and shake at 145r / min for 36h to obtain the bacterial solution of flocculating microorganisms;

[0099] S1.2 Cultivation: Dilute the bottom water of the brewing cellar three times, take 1L and add 10g of glucose, 2g of KH2PO4, 5g of K2HPO4, 0.2g of MgSO4, 1g of NaCl and 0.5g of urea, and then sterilize to obtain the cellar bottom water dilution + exogenous culture medium; then add the bacterial solution of 5% flocculant microorganisms to the cellar bottom water dilution culture medium for large-scale cultivation at 30℃, 130r / min for 36h to obtain the bioflocculating bacterial solution;

[0100] S2. Extraction: Adjust the pH of the bioflocculating bacterial solution to about 7, sterilize it in a high-temperature sterilizer at 121℃ for 21 minutes, take 10 ml of the bioflocculating bacterial solution, freeze it with 50 ml of liquid nitrogen, thaw it in a constant temperature water bath at 30℃, and then add 5 ml of hydrochloric acid with a mass fraction of 20% for acidification treatment for 5 minutes to obtain bioflocculating agent 7.

[0101] Example 8: Aerobic granules + LB medium + water from the bottom of the pit, diluted 5 times.

[0102] A method for rapidly preparing bioflocculants from high-concentration organic wastewater from brewing includes the following steps:

[0103] S1. Expanded culture:

[0104] S1.1 Screening for microorganisms with flocculation activity: Microorganisms with flocculation activity were screened from the aerobic sludge particles of the brewing wastewater treatment plant (in other embodiments, the source of flocculating microorganisms may also be commercially available or other channels). The specific screening method is as follows: First, weigh 10g of peptone, 5g of yeast powder, and 3g of sodium chloride, and add 1L of deionized water to prepare LB medium (in other embodiments, the source of LB medium may also be commercially available or other channels); then take 10ml of the aerobic sludge-water mixture and add it to 150ml of LB medium, place it at 30℃, and shake at 145r / min for 36h to obtain the bacterial solution of flocculating microorganisms;

[0105] S1.2 Cultivation: Dilute the bottom water of the brewing cellar 5 times, take 1L and add 10g of glucose, 2g of KH2PO4, 5g of K2HPO4, 0.2g of MgSO4, 1g of NaCl and 0.5g of urea, and then sterilize to obtain the cellar bottom water dilution + exogenous culture medium; then add the bacterial solution of 5% flocculent microorganisms to the cellar bottom water dilution culture medium for large-scale cultivation. The cultivation conditions are 30℃, 130r / min, and 36h to obtain the bioflocculating bacterial solution;

[0106] S2. Extraction: Adjust the pH of the bioflocculating bacterial solution to about 7, sterilize it in a high-temperature sterilizer at 121℃ for 21 minutes, take 10 ml of the bioflocculating bacterial solution, freeze it with 50 ml of liquid nitrogen, thaw it in a constant temperature water bath at 30℃, and then add 5 ml of hydrochloric acid with a mass fraction of 20% for acidification treatment for 5 minutes to obtain bioflocculating agent 8.

[0107] Example of effect

[0108] Take the bioflocculating agents 1-8 obtained in Examples 1-8 respectively, add 95 ml of deionized water, stir at 700 r / min for 10 min using a magnetic stirrer, and then centrifuge at 7000 rpm for 15 min in a centrifuge. Take 5 ml of the supernatant to determine the flocculation activity and flocculation rate. The determination method is as follows:

[0109] Weigh 4g of kaolin and dissolve it in 1000ml of distilled water to prepare a kaolin suspension. Weigh 1g of NaCl and dissolve it in 100ml of distilled water to prepare a 1% solution. Take 100ml of the kaolin suspension in a 250ml beaker, adjust the pH to 6.5-8 with NaOH, add 10ml of the 1% NaCl solution and bacterial culture, and stir with a magnetic stirrer at 440r / min for 30s, then at 120r / min for 5 minutes. After standing for 10 minutes, take the supernatant and measure its absorbance at 550nm with a spectrophotometer. Use the kaolin suspension with added fermentation culture medium as a control. The flocculation rate is calculated using the formula: flocculation rate = (AB) / A × 100%, where A is the absorbance of the control supernatant at 550nm and B is the absorbance of the sample supernatant with added fermentation broth at 550nm.

[0110] The flocculation results were measured as follows:

[0111] Table 2. Flocculation Effect of Bioflocculants 1-8

[0112]

[0113] As shown in Table 2, the microbial flocculant obtained by using pit bottom water and pot bottom water as inexpensive substrate culture medium to expand the culture of the screened bacterial liquid with flocculation activity has a good flocculation effect, with flocculation rates exceeding 87% and averaging 88.90%. This is 2.66% higher than the flocculation rate of the microbial flocculant obtained by using traditional laboratory culture medium. The cost of preparing the inexpensive substrate culture medium is low, and at the same time, the resource utilization of high-concentration organic wastewater from brewing is realized.

[0114] Comparative Example 1 investigated the effects of different culture conditions and extraction methods of bacterial suspension on the flocculation activity of the bacterial suspension.

[0115] 1. First, microorganisms with flocculation activity were screened from the activated sludge and aerobic sludge of the brewery wastewater treatment plant. The specific method is as follows:

[0116] Quantitatively weigh 10g of fresh activated sludge into a conical flask containing 90ml of sterile distilled water. After shaking on a shaker for 4 hours at a shaking speed of 150r / min, take 10ml of the supernatant and add it to LB, PDA, YPD, or Gao's No. 1 liquid culture medium, with a total culture medium volume of 150ml. At the same time, directly take 10ml of the aerobic sludge-water mixture and add it to 150ml of LB, PDA, YPD, or Gao's No. 1 liquid culture medium respectively. Place it at 30℃ and shake at 145r / min for 36 hours to obtain the bacterial solution of flocculating microorganisms. Take the bacterial solutions from LB, PDA, YPD, and Gao's No. 1 liquid culture medium respectively for flocculation activity determination.

[0117] Table 3. Flocculation rates obtained from different culture media.

[0118]

[0119] 2. The bacterial solution was subjected to freeze-thaw and acidification extraction before the flocculation rate was measured:

[0120] Take 10 ml of the bacterial culture obtained in Comparative Example 1 above, freeze it with 50 ml of liquid nitrogen, thaw it in a constant temperature water bath at 30°C, add 5 ml of 20% hydrochloric acid for acidification for 5 min, then add 95 ml of deionized water, stir at 700 r / min for 10 min using a magnetic stirrer, and then centrifuge at 7000 rpm for 15 min in a centrifuge. Take 10 ml of the supernatant to determine the flocculation activity.

[0121] The method for determining flocculation activity is the same as above, and the flocculation results are as follows:

[0122] Table 4. Flocculation rates of different bacterial cultures obtained from liquid nitrogen freeze-thaw + acid extraction treatment.

[0123]

[0124] Comparing Tables 3 and 4, it can be seen that the flocculation effect of bacterial solutions directly cultured on different culture media without freeze-thaw + acidification extraction treatment was poor. After freeze-thaw + acidification extraction treatment, the flocculation rate of bacterial solutions in LB, PDA, YPD, and Gao's No. 1 liquid media all increased significantly, indicating that freeze-thaw + acidification extraction is beneficial to improving the flocculation rate of bacterial solutions. Among them, the aerobic granular sludge bacterial solution in LB medium and the sludge bacterial solution in Gao's No. 1 liquid medium showed better flocculation effects. Therefore, this invention will further explore the effect of extraction method on flocculation rate based on the aerobic granular sludge bacterial solution in LB medium and the sludge bacterial solution in Gao's No. 1 liquid medium.

[0125] 3. Investigate the effects of liquid nitrogen freeze-thaw or hydrochloric acid extraction alone on the flocculation rate.

[0126] (1) Liquid nitrogen freeze-thaw: Take 10 ml of sludge bacterial culture of Gao Shi No. 1 and aerobic granular bacterial culture of LB respectively, freeze them with 50 ml of liquid nitrogen, thaw them with 30℃ warm water, add 95 ml of deionized water, stir with a magnetic stirrer at 700 r / min for 10 min, centrifuge at 7000 rpm for 15 min, and take the supernatant to determine the flocculation activity.

[0127] (2) Hydrochloric acid extraction: Take 10 ml of sludge bacterial solution cultured by Gao Shi No. 1 and aerobic granular bacterial solution cultured by LB, add 5 ml of 20% hydrochloric acid for acidification treatment for 5 min, then add 95 ml of deionized water, stir at 700 r / min for 10 min using a magnetic stirrer, then centrifuge at 7000 rpm for 15 min, and take the supernatant to determine the flocculation activity.

[0128] The results are as follows:

[0129] Table 5. Flocculation effect of flocculants extracted by different extraction methods

[0130]

[0131]

[0132] Comparing Tables 4 and 5, it can be seen that the flocculation rate of freeze-thaw + hydrochloric acid extraction is much greater than that of freeze-thaw treatment alone, and it is also better than that of acidification treatment alone. This indicates that the freeze-thaw + hydrochloric acid synergistic extraction method is more conducive to improving the flocculation rate, and freeze-thaw or acidification extraction alone cannot achieve the technical effect of this scheme.

[0133] Comparative Example 2 investigated the effects of other different extraction methods on the flocculation activity of bacterial culture.

[0134] As can be seen from Comparative Example 1 above, the effects of different extraction methods on the flocculation rate of bacterial solutions from different sources of activated sludge and aerobic particles follow the same trend. Therefore, further research on extraction methods should only be conducted based on bacterial solutions from activated sludge.

[0135] Quantitatively weigh 10g of fresh activated sludge into a conical flask containing 90ml of sterile distilled water. Shake on a shaker for 4 hours at a shaking speed of 150r / min. Take 10ml of the supernatant and add it to LB, PDA, YPD, or Gao's No. 1 liquid culture medium (150ml total). Incubate at 30℃ with shaking at 145r / min for 36 hours. Then, extract the sludge as follows:

[0136] (1) Centrifugation method: Take 10 ml of bacterial culture from LB, PDA, YPD and Gao's No. 1 liquid culture medium respectively, place them in a centrifuge and centrifuge at 7000 rpm for 15 min, and take the supernatant to determine the flocculation activity.

[0137] (2) The treatment method is centrifugation followed by the addition of ethanol:

[0138] Take 10 ml of bacterial culture from LB, PDA, YPD, and Gao's No. 1 liquid culture medium respectively, and centrifuge at 7000 rpm for 10 min. Take the supernatant, add 2 times the volume of ethanol at 4℃, let stand for 4 h, centrifuge at 6000 rpm for 10 min, collect the supernatant and measure the flocculation effect.

[0139] (3) The treatment method of freeze-thaw + acidification + addition of ethanol was adopted: 10 ml of bacterial culture was taken and frozen with 50 ml of liquid nitrogen. After thawing, it was thawed in a constant temperature water bath at 30℃. Then, 5 ml of hydrochloric acid with a mass fraction of 20% was added for acidification treatment for 5 min. Then, 95 ml of deionized water was added and stirred at 700 r / min for 10 min using a magnetic stirrer. The supernatant was taken and 2 times the volume of ethanol was added at 4℃ and allowed to stand for 4 h. Then, it was centrifuged at 7000 rpm for 15 min in a centrifuge. 10 ml of the supernatant was taken to determine the flocculation activity.

[0140] The results are as follows:

[0141] Table 6. Flocculation effects measured by other different treatment methods

[0142]

[0143] As can be seen from the table above, the flocculation effect of the bacterial solution measured by the freeze-thaw + acidification treatment method is the best, and the flocculation rate is higher than that of the direct centrifugation and centrifugation + ethanol treatment methods. Therefore, the freeze-thaw + acidification treatment method of the present invention is simple, does not consume ethanol, and has a good flocculation effect.

[0144] As can be seen from the above comparative examples 1 and 2, under the freeze-thaw + acidification extraction method, the activated sludge + Gao's medium has the best bacterial flocculation rate. Therefore, further research on the effects of different thawing methods and acidification extraction methods on bacterial flocculation activity will be carried out on the basis of activated sludge + Gao's medium.

[0145] Comparative Example 3 investigated the effects of different thawing methods on the flocculation activity of bacterial cultures.

[0146] (1) Natural thawing at room temperature: 10 ml of activated sludge + Gao's culture medium was frozen with 50 ml of liquid nitrogen and then thawed naturally at room temperature. 5 ml of 20% hydrochloric acid was added for acidification treatment for 5 min, followed by 95 ml of deionized water. The mixture was stirred at 700 r / min for 10 min using a magnetic stirrer and then centrifuged at 7000 rpm for 15 min. 10 ml of the supernatant was taken to determine the flocculation activity.

[0147] (2) Thawing in a constant temperature water bath at 30℃: 10ml of activated sludge + Gao's culture medium was frozen with 50ml of liquid nitrogen and then thawed in a constant temperature water bath at 30℃. 5ml of 20% hydrochloric acid was added for acidification treatment for 5min, followed by the addition of 95ml of deionized water. The mixture was stirred at 700r / min for 10min using a magnetic stirrer and then centrifuged at 7000rpm for 15min. 10ml of the supernatant was taken to determine the flocculation activity.

[0148] (3) Natural thawing at room temperature: 10 ml of activated sludge + Gao's culture medium was frozen with 50 ml of liquid nitrogen, thawed by sonication for 10 min, 5 ml of 20% hydrochloric acid was added for acidification for 5 min, then 95 ml of deionized water was added, and the mixture was stirred at 700 r / min for 10 min with a magnetic stirrer, and then centrifuged at 7000 rpm for 15 min. 10 ml of the supernatant was taken to determine the flocculation activity.

[0149] (4) Natural thawing at room temperature: 10 ml of activated sludge + Gao's culture medium was frozen with 50 ml of liquid nitrogen, thawed with cold water, acidified with 5 ml of 20% hydrochloric acid for 5 min, then 95 ml of deionized water was added, and stirred at 700 r / min for 10 min with a magnetic stirrer, and then centrifuged at 7000 rpm for 15 min. 10 ml of the supernatant was taken to determine the flocculation activity.

[0150] The results are as follows:

[0151] Table 7. Flocculation Effects Measured by Different Thawing Methods

[0152]

[0153] As can be seen from the table above, using a 30℃ water bath for thawing results in a slightly better flocculation effect than ultrasonic thawing, which is better than natural room temperature thawing and cold water thawing.

[0154] Comparative Example 4 investigated the effects of different acidification extraction methods on the flocculation activity of bacterial culture.

[0155] (1) Flocculation effect of hydrochloric acid treatment at different concentrations

[0156] The bacterial culture medium was prepared by adding 5% of the culture medium to 150 ml of inexpensive substrate medium and culturing it under the following conditions: 30-35℃, 130-145 rpm, for 36 h. The inexpensive substrate medium was diluted 5 times with water from the bottom of a pit, the pH was adjusted to 5-7, and then sterilized in a high-temperature autoclave at 121℃ for 21 min. After culturing the bacterial culture for 36-48 h, 10 ml of the culture was taken respectively and acidified with 5 ml of hydrochloric acid at 5%, 10%, 20%, and 30% concentrations. Then, 95 ml of deionized water was added, and the mixture was stirred at 700 rpm for 10 min using a magnetic stirrer, followed by centrifugation at 7000 rpm for 15 min. The supernatant was then used to determine the flocculation activity, and the results are as follows:

[0157] Table 8. Flocculation effects obtained from treatment with different concentrations of hydrochloric acid.

[0158]

[0159] As shown in the table above, the flocculation effect obtained by treatment with 20% hydrochloric acid is better than that obtained by treatment with other concentrations of hydrochloric acid.

[0160] (2) Flocculation effect of different acid treatments

[0161] Take 10 ml of the bacterial solution prepared by high-temperature sterilization in (1), and acidify it for 5 min each with 5 ml of 20% nitric acid, 20% hydrochloric acid, 20% formic acid, and 20% acetic acid. Then add 95 ml of deionized water, stir at 700 r / min for 10 min using a magnetic stirrer, and centrifuge at 7000 rpm for 15 min. Take the supernatant and measure the flocculation activity. The results are as follows:

[0162] Table 9. Flocculation effects obtained from treatment with different acids of the same concentration.

[0163]

[0164] As shown in Table 9, the flocculation effect obtained by acidification extraction with 20% hydrochloric acid is the best.

[0165] Comparative Example 5 investigated the effect of different bacterial suspension addition amounts on bacterial suspension flocculation activity.

[0166] Take 10ml, 5ml, and 2ml of sludge bacterial culture cultured in Gao's No. 1 liquid culture medium, respectively, and quickly freeze them with 50ml of liquid nitrogen. Then thaw them with 30℃ warm water, add 90ml of deionized water, stir with a magnetic stirrer at 700r / min for 10min, and centrifuge at 7000rpm for 15min. Take the supernatant to determine the flocculation activity and flocculation rate.

[0167] The results are as follows:

[0168] Table 10. Flocculation effect of adding different amounts of bacterial solution to the extracted flocculant

[0169]

[0170] Table 10 shows that adding 10ml, 5ml, and 2ml of sludge bacterial solution resulted in high flocculation rates exceeding 80%, indicating that a range of 2-10ml of sludge bacterial solution yields good results. However, adding 5ml of sludge bacterial solution resulted in a flocculation rate of 88.52%, higher than that of adding 10ml and 2ml. This demonstrates that the amount of sludge bacterial solution added significantly affects the flocculation effect. Therefore, the optimal flocculant dosage is 5ml, which is relatively small. Thus, the flocculating microorganisms in the brewing environment were identified as originating from the sludge in the brewing wastewater treatment plant and the activated sludge in the aerobic tank. Flocculating bacterial solution was obtained using LB medium and Gao's No. 1 liquid medium, and the optimal treatment method for extracting the flocculant was determined to be freeze-thaw + hydrochloric acid treatment, with an addition amount of 5ml of flocculant.

[0171] Comparative Example 6 investigated the effect of pit bottom water with different COD concentrations as an expansion culture medium on the flocculation activity of bacterial culture.

[0172] The bottom water was diluted 30.8 times, 308 times, and 10 times, respectively, reducing the COD concentration to 5 g / L, 0.5 g / L, and 15.4 g / L. The bacterial solution cultured with activated sludge and Gao's No. 1 medium was added at a ratio of 5% to the expansion medium with different COD concentrations for expansion culture. The culture conditions were 30℃ and 130 rpm, and cultured for 36 h and 72 h, respectively. 10 ml of the bioflocculating bacterial solution was taken from each solution, frozen with 50 ml of liquid nitrogen, thawed in a constant temperature water bath at 30℃, acidified with 5 ml of 20% hydrochloric acid for 5 min, then 95 ml of deionized water was added, and the mixture was stirred at 700 rpm for 10 min using a magnetic stirrer, followed by centrifugation at 7000 rpm for 15 min. 5 ml of the supernatant was taken to determine the flocculation activity using the same method as above. The flocculation results are as follows:

[0173] Table 11. Flocculation effects obtained from culturing water at different dilution ratios at the bottom of the pit.

[0174]

[0175] In this invention, after diluting the original bottom water (COD 154 g / L) by 3 or 5 times, the COD is effectively reduced to about 50 g / L or 30 g / L. As shown in the table above, the bacterial culture cultured after diluting the bottom water by 5 times has the best flocculation effect. However, the bacterial culture cultured at a lower concentration (COD concentration between 0.5 g / L and 5 g / L) has a lower flocculation effect than the bacterial culture cultured in a larger culture medium with a higher concentration of bottom water.

[0176] Comparative Example 7 investigated the effects of different types of seed culture media on the flocculation activity of bacterial cultures.

[0177] Gao's No. 1 liquid medium, Czapek's medium, beef extract peptone medium (MRS), and general fermentation medium (SB) were prepared separately. 10g of fresh activated sludge was quantitatively weighed into an Erlenmeyer flask containing 90ml of sterile distilled water. After shaking on a shaker for 4 hours (shaking speed 150r / min), 10ml of the supernatant was added to each of the three media for incubation at 30℃ and 130r / min for 36 hours. The samples from the different media were then compared. The bacterial culture was added at a ratio of 5% to a 5-fold diluted culture medium from the bottom of the pit and cultured under the conditions of 30℃ and 130r / min for 36h. After culturing, 10ml of the flocculant culture was taken, frozen with 50ml of liquid nitrogen, thawed in a constant temperature water bath at 30℃, acidified with 5ml of 20% hydrochloric acid for 5min, and then 95ml of deionized water was added. The mixture was stirred at 700r / min for 10min using a magnetic stirrer and then centrifuged at 7000rpm for 15min. 10ml of the supernatant was taken to determine the flocculation activity.

[0178] Czapek's medium preparation method: sucrose 6g, FeSO4 0.002g, K2PO4 1.0g, (NH4)2SO4 1.0g, KCl 0.5g, NaNO3 3.0g, MgSO4 0.5g, agar 15g, 1L deionized water, pH 7.0;

[0179] Preparation method of beef extract peptone medium: 3.0g beef extract, 10.0g peptone, 5.0g NaCl, 1L deionized water;

[0180] General fermentation medium preparation method: 3.2% tryptone, 2% yeast extract, 0.5% NaCl.

[0181] The flocculation results are as follows:

[0182] Table 12 Flocculation effect of different types of seed culture media

[0183]

[0184] As can be seen from the table above, Gao's No. 1 liquid culture medium is more effective than other types of culture media for culture and screening.

[0185] Comparative Example 8 investigated the effects of soy sauce brewing wastewater and brewing cellar bottom water as inexpensive culture media on bioflocculants.

[0186] Weigh 10g of fresh activated sludge into a conical flask containing 90ml of sterile distilled water. Shake on a shaker for 4 hours (shaking speed 150r / min). Take 10ml of the supernatant and add it to Gaoshi No. 1 liquid culture medium for cultivation. The cultivation conditions are 30℃ and 130r / min. After culturing for 36 hours, seed culture is obtained. This seed culture is added at a ratio of 5% to 5 times the volume of pit bottom water culture medium and soy sauce brewing wastewater for cultivation (take 150ml of each and sterilize in a high-temperature sterilizer at 121℃ for 21min). The culture conditions were 30℃, 130r / min, and 36h. After culture, flocculating bacterial solution was obtained. 10ml of flocculating bacterial solution was frozen with 50ml of liquid nitrogen, thawed in a constant temperature water bath at 30℃, and then acidified with 5ml of 20% hydrochloric acid for 5min. Then, 95ml of deionized water was added, and the mixture was stirred at 700r / min for 10min using a magnetic stirrer. Finally, it was centrifuged at 7000rpm for 15min. 10ml of the supernatant was taken to determine the flocculation activity.

[0187] Soy sauce brewing wastewater has a COD of approximately 900-1000 mg / L and NH4+. + -N is approximately 80-100 mg / L, and TN is approximately 300-400 mg / L.

[0188] Table 13 Flocculation effect of flocculant cultures using different types of inexpensive substrates

[0189]

[0190] As shown in the table above, compared with using soy sauce brewing wastewater as a cheap substrate to prepare bio-flocculating agents, the bio-flocculating agents cultured in brewing cellar bottom water culture medium have a better flocculation rate without the addition of supplementary carbon and nitrogen sources.

[0191] It is understood that the present invention has been described through some embodiments, and those skilled in the art will recognize that various changes or equivalent substitutions can be made to these features and embodiments without departing from the spirit and scope of the invention. Furthermore, under the teachings of the present invention, these features and embodiments can be modified to adapt to specific situations and materials without departing from the spirit and scope of the invention. Therefore, the present invention is not limited to the specific embodiments disclosed herein, and all embodiments falling within the scope of the claims of this application are within the protection scope of the present invention.

Claims

1. A method for preparing a bioflocculant, characterized by, Includes the following steps: S1. Expanded culture: The seed liquid of flocculating microorganisms is added to a culture medium containing brewing organic wastewater for expanded culture to obtain bioflocculating bacterial liquid; The flocculating microorganisms were screened from activated sludge and / or aerobic sludge from a brewing wastewater treatment plant. The screening method included: weighing fresh activated sludge and / or aerobic sludge into sterile distilled water, shaking it at a speed of 145-155 r / min for 3-5 h, adding the supernatant to a culture medium, and culturing it under the following conditions: 28-32℃, shaking at a speed of 140-150 r / min, for 35-38 h; thus obtaining the seed culture of the flocculating microorganisms. When the sludge is activated sludge, the culture medium in the screening method is Gao's No. 1 liquid culture medium; when the sludge is aerobic sludge, the culture medium in the screening method is LB culture medium. The culture medium for the expanded culture is a diluted culture medium of pit bottom water, a culture medium of pot bottom water, a diluted exogenous culture medium of pit bottom water, or an exogenous culture medium of pot bottom water. The diluted culture medium of pit bottom water is a culture medium obtained by diluting the pit bottom water, and the dilution ratio of the pit bottom water is 3-5 times. The culture medium of pot bottom water is a culture medium directly using pot bottom water, and the COD content of the pit bottom water is 150,000-160,000 mg / L, and the COD content of the pot bottom water is 5,000-6,000 mg / L. S2. Extraction: After sterilization of the bioflocculating bacterial solution, extraction is performed by a combination of freeze-thaw and acidification to obtain the bioflocculating agent; the freeze-thaw process includes freezing with liquid nitrogen and then thawing with a constant temperature water bath; the volume ratio of liquid nitrogen to the bioflocculating bacterial solution is (45-55):(8-12); the temperature of the constant temperature water bath is 28-35℃.

2. The method for preparing a bioflocculant according to claim 1, wherein In step S2, the acidification includes acidification extraction using hydrochloric acid; the mass fraction of the hydrochloric acid is 20%; the volume ratio of the hydrochloric acid to the bioflocculating bacteria liquid is (4-6):(8-12); and the hydrochloric acid acidification treatment time is 4-6 min.

3. The method for preparing the bioflocculant as described in claim 1, characterized in that, The activated sludge is fresh precipitate or suspension separated from the secondary sedimentation tank of a brewery wastewater treatment plant; the aerobic sludge is the sludge obtained from the secondary aerobic tank of the biological contact oxidation-oxidation process A 2 O 2 The sludge-water mixture obtained in the secondary aerobic tank of the process section.

4. The method for preparing the bioflocculant as described in claim 1, characterized in that, The conditions for the water at the bottom of the pit include: pH 4-5; BOD5 content 37000-38000 mg / L; NH3-N content 3000-4000 mg / L; TP content 2000-2500 mg / L; TN content 7000-8000 mg / L; and the conditions for the water at the bottom of the pot include: pH 3-4; BOD5 content 1500-1600 mg / L.

5. The method for preparing the bioflocculant as described in claim 4, characterized in that, The exogenous culture medium diluted with the pit bottom water is obtained by adding additional nutrients to the culture medium diluted with the pit bottom water. The exogenous culture medium for the bottom water is obtained by adding nutrients to the culture medium for the bottom water; the nutrients include at least one of carbon source, nitrogen source and inorganic element.

6. The method for preparing the bioflocculant as described in claim 5, characterized in that, The nutrients include at least one of glucose, KH2PO4, K2HPO4, MgSO4, NaCl, and urea; the content of the nutrients is as follows: glucose 8-12 g / L, KH2PO4 1-5 g / L, K2HPO4 2-7 g / L, MgSO4 0.1-0.5 g / L, NaCl 0.5-3 g / L, and urea 0.1-2 g / L.

7. The method for preparing the bioflocculant as described in claim 1, characterized in that, In step S1, the expanded culture conditions are: temperature 28-32℃, rotation speed 125-135r / min, and time 30-40h.

8. The method for preparing the bioflocculant as described in claim 1, characterized in that, In step S1, the volume ratio of the seed liquid of the floc-producing microorganism to the culture medium for expanded culture is (3-8):

100.

9. A bioflocculant prepared by the method of any one of claims 1-8.

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