CN105039417A - Method for preparing microbial flocculant through enteromorpha hydrolysate fermentation - Google Patents

Abstract

The invention discloses a method for preparing a microbial flocculant through enteromorpha hydrolysate fermentation. The method comprises the steps of preparation of liquid strain, fermentation and product aftertreatment. The fermentation process comprises the steps that 1, fresh enteromorpha is cleaned, salt, sediment and unwanted algae are removed, then the fresh enteromorpha is smashed and added to a hydrolysis tank, water is added to regulate acidity, alkali is added for neutralization, filter pressing and clarification are conducted, and then a hydrolysate is obtained; 2, the hydrolysate is diluted till the sugar concentration of reducing sugar is 15-30 g/L, fermentation liquor is prepared and added to a fermentation tank, and high-temperature sterilization and cooling are conducted; 3, a seed solution is inoculated into the fermentation tank for fermentation, and fermentation is ended after the content of residual sugar in the fermentation liquor is reduced to the level below 3 g/L. According to the method, enteromorpha is used as the raw material, microbial flocculant fermentation is directly conducted after acid hydrolysis filtration, the production method is simple, production cost is low, grain is saved, and waste is turned into wealth.

Description

A kind of Enteromorpha hydrolyzate fermentation is for the method for microbial flocculant

Technical field

The present invention relates to microbial flocculant preparation field, particularly a kind of Enteromorpha hydrolyzate fermentation is for the method for microbial flocculant.

Background technology

Enteromorpha is a kind of large-scale green alga, belongs to Ulvaceae kelp, and common kind has bar Enteromorpha, all over Enteromorpha etc.Enteromorpha all has distribution in the southern and northern sea area of China, and Enteromorpha growth is rapidly, day rate of propagation can reach 12.7%, can form considerable biomass in the short period of time, be one urgently biological resources leaved for development.Enteromorpha contains a large amount of nutritive ingredients, and containing the polysaccharide of about 50% in Enteromorpha, also contain the albumen of about 12%, Enteromorpha can produce the carbohydrates such as glucose, wood sugar, rhamnosyl after acid hydrolysis simultaneously.These materials can well be utilized by microorganism.Due to pollution and the eutrophication of seawater in recent years, China coast creates a large amount of Enteromorphas accrued, because do not have effective disposal of resources approach, causes great impact to the ecotope of bank.

At present, along with socioeconomic fast development, the demand of tap water and the discharge of sewage are all in sharp increase.In present stage water treatment method, the medicament the most often used is chemical floc, as polymerize aluminum chloride and polyacrylamide etc., chemical floc easily causes the secondary pollution to environment, chemical floc itself has certain biohazardous, and chemical floc is not easily degraded, the mud after water treatment is more difficult carries out harmless treatment.Microbial polysaccharide flocculant did not both have toxicity, had easy degraded, and be the good substitute of existing chemical floc, prospect is very bright and clear.

Publication number is that the Chinese invention patent " a kind of method of preparing bioflocculant through high-concentration fermentation " of 102586331B discloses the method preparing microbial flocculant with micrococci DS016 liquid state fermentation.It is that the glucose of raw material and relevant carbohydrate do carbon source with corn that existing fermentation adopts for the method great majority of microbial flocculant, and raw materials cost is higher.

Summary of the invention

The object of the invention is for the deficiencies in the prior art, solve microorganism and prepare the high technical problem of flocculation agent process Raw cost.

Object of the present invention is achieved by following technical scheme:

Enteromorpha hydrolyzate fermentation, for a method for microbial flocculant, comprises the steps:

One, the preparation of liquid spawn

(1) slant strains cultivate: by micrococci DS16 strain inoculation on slant medium, and at 28-30 DEG C heat insulating culture 36h;

Described slant medium consists of: extractum carnis 3-6g/L, peptone 8-12g/L, liquid sugar 7-12g/L, NaCl3-6g/L, pH=7.0-7.2;

Described micrococci DS16 bacterial classification is separated by Shandong Provincial Pharmaceutical Biological Tech. Research Center and a kind of bacterial strain producing high-effective microorganism polysaccharide flocculant preserved.

(2) strain cultivation: add Shake flask medium 50mL in 250ml triangle is bottled, inoculates a transfering loop slant strains, and 100-120r/min, 28-30 DEG C of heat insulating culture 12h, obtains liquid spawn.

Described Shake flask medium consists of: glucose 8-12g/L, peptone 6-12g/L, yeast extract 0.3-0.8g/L, K ₂hPO ₄3-8g/L, KH ₂pO ₄2-7g/L, MgSO ₄7H ₂o0.3-0.9g/L, pH=7.

Two, ferment

(1) fresh Enteromorpha cleaning is removed freshen, silt and assorted algae, smash with pulverizer after adding suitable quantity of water.Then join in hydrolytic decomposition pot, add water and make material-water ratio be 1:30-1:20, add sulfuric acid and regulate acidity to be 0.5%-5%(w/w), pass into high-temperature steam, be warming up to 110-125 DEG C, insulation 0.5-2h is hydrolyzed; Add white lime and be neutralized to pH value about 7.0, press filtration is clarified, and obtains hydrolyzed solution, calcium sulphate solid after recovery press filtration;

(2) hydrolyzed solution in above-mentioned steps (1) being diluted to reducing sugar sugar concentration is 15-30g/L, and adds urea 0.5-1.0g/L according to following concentration, (NH4) ₂sO ₄1-5g/L, peptone 0.1-0.5g/L, K ₂hPO43-8g/L, NaH ₂pO ₄2-7g/L, MgSO ₄7H ₂o0.1-0.5g/L, MnCl ₂5H ₂o0.03g/L, regulates pH=7, and preparation fermented liquid adds in fermentor tank, passes into 121 DEG C of high-temperature steams, and insulation 30min, is cooled to 20-40 DEG C.

(3) by after in aforesaid liquid strain inoculation to fermentor tank, be 0.01-0.03Mpa, sterile air ventilating ratio 1: 0.25-0.65 (V/V) at tank pressure, mixing speed 90-170r/min, temperature is under the condition of 20-30 DEG C, cultivate 18-24h, after fermented liquid residual sugar content is down to below 3g/L, fermentation ends.

Three, product aftertreatment:

(1) fermentation liquor pretreatment, regulates fermented liquid pH to be 6.0-7.5 with sodium hydroxide solution (it is 5-15% that its concentration is preferably weight percent), after the sterilizing filter of 0.1-1um is degerming, obtain bacteria-removing liquid.

(2) by bacteria-removing liquid in above-mentioned steps (1) in inlet temperature: 140-145 DEG C; Be dried to moisture through centrifugal spray drying tower at air outlet temperature 75-80 DEG C and be less than 5%;

(3) load in aluminium plastic bag after the product after spraying dry being crossed 80 mesh sieves, vacuumize with Vacuum Packaging Machine and pack, obtain the finished product.

The present invention utilizes the Enteromorpha of the low cost that can breed in a large number to be raw material, after acid hydrolysis is filtered, directly carries out microbial flocculant fermentation, the mode of production is easy, production cost is low, saves food, turns waste into wealth.

Product prepared by the present invention is a kind of microbial polysaccharide flocculant efficiently, and nontoxic degradable, does not have secondary pollution, effectively can substitute the existing inorganic flocculating agent and the organic floculant that have open defect.Product flocculation activity is high, effectively can remove the objectionable impuritiess such as COD, the SS in the water bodys such as sanitary sewage, industrial sewage and surface water.Microbial polysaccharide flocculant can reach 98% to the kaolin suspension 5min settling efficiency of 0.4%; This product of single utilization carries out flocculation treatment COD removal efficiency to sewage work Yuan Shui and reaches 85%, can reach 91% to SS removal efficiency.

Embodiment

Below in conjunction with embodiment, the invention will be further described,

embodiment 1:

One, liquid spawn preparation

(1) cultivation of slant strains: by DS16 strain inoculation in slant medium, and at 28 DEG C heat insulating culture 36h.

Described slant medium consists of extractum carnis 3g/L, peptone 8g/L, liquid sugar 7g/L, NaCl3g/L, pH=7.0.

(2) strain cultivation: fill Shake flask medium 50mL in 250mL triangular flask, inoculates a transfering loop slant strains, and at 100r/min, under 28 DEG C of conditions, heat insulating culture 12h, obtains liquid spawn.

Described Shake flask medium consists of glucose 8g/L, peptone 6g/L, yeast extract 0.3g/L, K ₂hPO ₄3g/L, KH ₂pO ₄2g/L, MgSO ₄7H ₂o0.3g/L, pH=7.

Two, ferment

(1) fresh Enteromorpha cleaning is removed freshen, silt and assorted algae, smash with pulverizer after adding suitable quantity of water, join in hydrolytic decomposition pot, add water and make material-water ratio be 1:20, add sulfuric acid and make acidity be 0.5%(w/w).Pass into high-temperature steam, be warming up to 110 DEG C, insulation 0.5h is hydrolyzed.Add white lime and be neutralized to pH value about 7.0, press filtration is clarified, and obtains hydrolyzed solution, calcium sulphate solid after recovery press filtration.

(2) hydrolyzed solution in above-mentioned (1) is diluted to reducing sugar sugar concentration 15g/L, and adds urea 0.5g/L, (NH ₄) ₂sO ₄1g/L, peptone 0.1g/L, K ₂hPO ₄3g/L, NaH ₂pO ₄2g/L, MgSO ₄7H ₂o0.1g/L, MnCl ₂5H ₂o0.03g/L, regulate pH=7, preparation fermented liquid add in fermentor tank, pass into high-temperature steam 121 DEG C, insulation 30min, be cooled to 20 DEG C for subsequent use.

(3) by aforesaid liquid bacterial classification access fermentor tank, the culture condition of control: tank pressure is 0.01Mpa, sterile air ventilating ratio 1: 0.25 (V/V), and mixing speed 90r/min, temperature is 30 DEG C, incubation time 18h.After fermented liquid residual sugar content is down to below 3g/L, fermentation ends.

Three, product aftertreatment

(1) fermentation liquor pretreatment, regulates pretreated fermentation liquid pH=7.5 by fermented liquid with sodium hydroxide solution (its concentration is preferably weight percent for 5%).Bacteria-removing liquid is obtained after the sterilizing filter of 0.1-1um is degerming.

(2) by bacteria-removing liquid in above-mentioned steps (1) in inlet temperature: 140 DEG C; Be dried to moisture through centrifugal spray drying tower at air outlet temperature 75 DEG C and be less than 5%;

(3) load in aluminium plastic bag after the product after spraying dry being crossed 80 mesh sieves, vacuumize with Vacuum Packaging Machine and pack, obtain the finished product.

embodiment 2

One, liquid spawn preparation

The cultivation of slant strains: by DS16 strain inoculation in inclined-plane, and at 30 DEG C heat insulating culture 36h;

Described slant medium consists of extractum carnis 6g/L, peptone 12g/L, liquid sugar 12g/L, NaCl6g/L, pH=7.2;

The cultivation of liquid spawn: fill Shake flask medium 50mL in 250mL triangular flask, inoculates a transfering loop slant strains, in 120r/min, and heat insulating culture 12h at 30 DEG C;

Described Shake flask medium consists of glucose 1210g/L, peptone 12g/L, yeast extract 0.8g/L, K ₂hPO ₄8g/L, KH ₂pO ₄7g/L, MgSO ₄7H ₂o0.9g/L, pH=7;

Two, ferment

(1) fresh Enteromorpha cleaning is removed freshen, silt and assorted algae, smash with pulverizer after adding suitable quantity of water, then join in hydrolytic decomposition pot, add water and make material-water ratio be 1:30, add sulfuric acid and make acidity be 5%(w/w).Pass into high-temperature steam, be warming up to 125 DEG C, insulation 2h is hydrolyzed.Add white lime and be neutralized to pH value about 7.0, press filtration is clarified, calcium sulphate solid after recovery press filtration.

(2) hydrolyzed solution in above-mentioned (1) is diluted to reducing sugar sugar concentration 30g/L, and adds urea 1.0g/L, (NH ₄) ₂sO ₄5g/L, peptone 0.5g/L, K ₂hPO ₄8g/L, NaH ₂pO ₄7g/L, MgSO ₄7H ₂o0.5g/L, MnCl ₂5H ₂o0.03g/L, regulates pH=7, prepares fermented liquid and add in fermentor tank, pass into high-temperature steam 121 DEG C, insulation 30min.Be cooled to 40 DEG C for subsequent use.

(3) accessed after in fermentor tank by aforesaid liquid bacterial classification, the culture condition of control: tank pressure 0.03Mpa, sterile air ventilating ratio 1: 0.65 (V/V), mixing speed 170r/min, temperature is 30 DEG C, incubation time 18-24h.After fermented liquid residual sugar content is down to below 3g/L, fermentation ends.

Three, product aftertreatment:

(1) regulate fermented liquid pH to be 7.0 with sodium hydroxide solution (it is 15% that its concentration is preferably weight percent), then after the sterilizing filter of 0.1-1um is degerming, obtain bacteria-removing liquid.

(2) by bacteria-removing liquid in above-mentioned steps (1) in inlet temperature: 145 DEG C; Be dried to moisture through centrifugal spray drying tower at air outlet temperature 80 DEG C and be less than 5%;

(3) load in aluminium plastic bag after the product after spraying dry being crossed 80 mesh sieves, vacuumize with Vacuum Packaging Machine and pack, obtain the finished product.

Measure microbial polysaccharide flocculant flocculating effect prepared by embodiment 1-2, result shows: the method produce flocculation agent to 0.4% kaolin suspension 5min settling efficiency can reach 98%; This product of single utilization carries out flocculation treatment COD removal efficiency to sewage work Yuan Shui and reaches 85%, can reach 91% to SS removal efficiency.

Claims (4)

1. Enteromorpha hydrolyzate fermentation is for a method for microbial flocculant, it is characterized in that, comprise liquid spawn preparation, fermentation and product aftertreatment, described fermentation comprises the steps:

(1) fresh Enteromorpha cleaning is removed freshen, silt and assorted algae, pulverize, join in hydrolytic decomposition pot, add water and make material-water ratio be 1:30-1:20, regulate acidity and temperature to be hydrolyzed, then add alkali and be neutralized to pH value about 7.0, press filtration is clarified, and obtains Enteromorpha hydrolyzed solution;

(2) Enteromorpha hydrolyzed solution in above-mentioned (1) being diluted to reducing sugar sugar concentration is 15-30g/L, and adds urea 0.5-1.0g/L, (NH ₄) ₂sO ₄1-5g/L, peptone 0.1-0.5g/L, K ₂hPO ₄3-8g/L, NaH ₂pO ₄2-7g/L, MgSO ₄7H ₂o0.1-0.5g/L, MnCl ₂5H ₂o0.03g/L, regulates pH=7, prepares fermented liquid and add in fermentor tank, pass into high-temperature steam 121 DEG C, and insulation 30min, is cooled to 20-40 DEG C;

(3) seed liquor is inoculated in fermentor tank, be 0.01-0.03Mpa, sterile air ventilating ratio 1: 0.25-0.65 (V/V) at tank pressure, mixing speed 90-170r/min, temperature is under the condition of 20-30 DEG C, cultivate 18-24h, after fermented liquid residual sugar content is down to below 3g/L, fermentation ends.

2. Enteromorpha hydrolyzate fermentation according to claim 1 is for the method for microbial flocculant, it is characterized in that, the described acid of described fermenting process step (1) is sulfuric acid, and alkali is white lime.

3. Enteromorpha hydrolyzate fermentation according to claim 2 is for the method for microbial flocculant, it is characterized in that, described product aftertreatment comprises the steps:

(1) adding sodium hydroxide solution regulates fermented liquid pH to be 6.0-7.5; Bacteria-removing liquid is obtained after the sterilizing filter of 0.1-1um is degerming;

(2) by bacteria-removing liquid in above-mentioned steps (1) in inlet temperature: 140-145 DEG C; Be dried to moisture through centrifugal spray drying tower at air outlet temperature 75-80 DEG C and be less than 5%;

(3) product after spraying dry is loaded in aluminium plastic bag, vacuumize with Vacuum Packaging Machine and pack, obtain the finished product.

4. Enteromorpha hydrolyzate fermentation according to claim 3 is for the method for microbial flocculant, it is characterized in that, described liquid spawn preparation comprises the steps:

(1) slant strains cultivate: by micrococci DS16 strain inoculation on slant medium, and at 28-30 DEG C heat insulating culture 36h; Described slant medium consists of: extractum carnis 3-6g/L, peptone 8-12g/L, liquid sugar 7-12g/L, NaCl3-6g/L, pH=7.0-7.2;

(2) strain cultivation: add Shake flask medium 50mL in 250ml triangle is bottled, inoculates a transfering loop slant strains, and 100-120r/min, 28-30 DEG C of heat insulating culture 12h, obtains liquid spawn; Described Shake flask medium consists of: glucose 8-12g/L, peptone 6-12g/L, yeast extract 0.3-0.8g/L, K ₂hPO ₄3-8g/L, KH ₂pO ₄2-7g/L, MgSO ₄7H ₂o0.3-0.9g/L, pH=7.