Summary of the invention
The purpose of this invention is to provide a kind of utilization and the red bacterium of class ball (Rhodobacter sphaeroides) is carried out the fermentation process of the industrially scalable of deep layer, the fermented liquid that obtains is healthy and helpful to human body, can regulate the immunity system of human body, increase resistance of human body.
In the inventor's series of patents (Chinese CN1184307C), related to similar fermentation process, but, optimized the composition of fermentation culture through the continuation in several years of contriver test, and optimized fermentation condition, obtained better effects if, fermented liquid that cost is lower.
The present invention utilizes the method for the red bacterium industrially scalable fermentation of class ball, this method is that the red bacterium of class ball (Rhodobactersphaeroides) is passed through the first class seed pot of 500ml Erlenmeyer flask, 50-200L, secondary seed jar and the submerged fermentation of 5000-20000L large fermentation tank of 500-2000L successively
Fermentation condition is respectively:
The 500ml Erlenmeyer flask: constant temperature 25-35 ℃, with 300LX incandescent light photograph, shaking speed per minute 50-60 carried out 24 hours under changeing;
200L seeding tank: pH6.4-7.5, temperature is 25-35 ℃, with of the visor illumination of 300LX incandescent light by two different angles, constant temperature 25-35 ℃ of fermentation 24 hours carried out enlarged culturing 24 hours by first class seed pot (50-200L) and secondary seed jar (500-2000L), and culture temperature wherein, pH value, illumination and venting pressure are identical with first class seed pot;
The pressure of firsts and seconds seeding tank all is 0.04Mpa;
The fermentation condition of large fermentation tank is as follows: by the illumination that the visor of three different angles is given the vehement light of 300-500LX, and material liquid pH 6.4-7.5, intake pressure 0.1-0.3Mpa adjusts a jar internal pressure 0.05-0.08Mpa, controlled temperature 25-35 ℃, ferments 24 hours;
Promptly obtain fermented liquid, it is characterized in that: the red bacterium of described class ball is for being preserved in China Committee for Culture Collection of Microorganisms common micro-organisms center, numbering CGMCC No.1562, and the compound method of described nutrient solution is:
Inorganic salt solution proportioning in the 1000ml distilled water;
MgSO
47H
2O 0.2-3.0 gram
CaCl2H
2O 0.1-1.0 gram
FeSO
47H
2O 0.1-1.0 gram
NaEDTA 0.1-1.0 gram
Trace element solution proportioning in the 250ml distilled water;
H
3BO
40.5-1.5 gram
MnSO
4The 50-200 milligram
NaMoO
4The 20-100 milligram
ZnSO
4The 10-20 milligram
Cu (NO
3)
2The 2-3 milligram
Sodium orthomolybdate 6-15 milligram
Sodium Selenite 6-15 milligram;
Growth factor solution proportioning in the 100ml distilled water
Vitamin H 0.2-0.5 milligram
Vitamins B
1The 10-30 milligram
Nicotinic acid 10-30 milligram
Para-amino benzoic acid 10-20 milligram;
The lactic acid solution proportioning
900 milliliters of distilled water
Lactic acid 30-100 milliliter is transferred pH6.4 with the NaOH solution of 12N;
The ammoniumsulphate soln proportioning
800 milliliters of distilled water
Ammonium sulfate 3-8 gram
Water adds to 1000 milliliters;
Add to 1000 milliliters with distilled water the dissolving back, transfers the pH6.4 phosphoric acid buffer with the NaOH solution of 12N then
K
2HPO
4The 15-50 gram
KH
2PO
4The 10-15 gram
Water adds to 1000 milliliters;
Get respectively:
A, inorganic salt solution 10-30%,
B, trace element solution 0.1-1.0%
C, growth factor solution 0.1%
D, lactic acid solution 2-10%
E, ammoniumsulphate soln 0.5-2%
F, phosphoric acid buffer 1-5%
Proportioning a-f is mixed according to above-mentioned per-cent, and all the other are water, promptly obtain nutrient solution.Add in addition when doing slant tube or culture dish substratum: peptone 1%, yeast extract paste 0.2%.
Preferably:
Inorganic salt solution proportioning in the 1000ml distilled water;
MgSO
47H
2O 0.3 gram
CaCl2H
2O 0.05 gram
FeSO
47H
2O 0.5 gram
NaEDTA 0.5 gram
Trace element solution proportioning in the 250ml distilled water;
H
3BO
41.0 gram
MnSO
4120 milligrams
NaMoO
460 milligrams
15 milligrams of ZnSO4
Cu (NO
3)
22 milligrams
12 milligrams of Sodium orthomolybdates
12 milligrams of Sodium Selenites;
Growth factor solution proportioning in the 100ml distilled water
0.5 milligram of vitamin H
Vitamins B
110 milligrams
10 milligrams in nicotinic acid
10 milligrams of para-amino benzoic acid;
The lactic acid solution proportioning
900 milliliters of distilled water
40 milliliters of lactic acid are transferred pH6.4 with the NaOH solution of 12N;
The ammoniumsulphate soln proportioning
800 milliliters of distilled water
Ammonium sulfate 5 grams
Water adds to 1000 milliliters;
Add to 1000 milliliters with distilled water the dissolving back, transfers the pH6.4 phosphoric acid buffer with the NaOH solution of 12N then
K
2HPO
440 grams
KH
2PO
410 grams
Water adds to 1000 milliliters;
Get respectively:
A, inorganic salt solution 20%,
B, trace element solution 0.2%
C, growth factor solution 0.1%
D, lactic acid solution 5%
E, ammoniumsulphate soln 1.5%
F, phosphoric acid buffer 3%
Proportioning a-f is mixed according to above-mentioned per-cent, and all the other are water, promptly obtain nutrient solution.Add in addition when doing slant tube or culture dish substratum: peptone 1%, yeast extract paste 0.2%.
The fermented liquid that obtains with aforesaid method has following purposes:
Prepare biologic beverage with it, wherein with fermented liquid with after pure water mixes, adjust taste with foodstuff additive.
Irrigate butt rot place with 5-10 times of fermented liquid and can treat root rot.
With the fermented liquid spray water fruits described in the 50-100 claim 3 doubly with can eat vegetables raw, the SOD of fruit is improved.
With the fermented liquid described in the 100-150 claim 3 doubly as the viable bacteria biomaterial.
Give breeded fish, the 3-5ppm by the pond water yield in the pond of shrimp or crab adds, can purify water, reduce disease, improve the speed of growth.
Fermented liquid and water are according to 1: 5-10 adds doubly in the feed that is added to milk cow, pig, chicken or the tap water and feeds.
Frequency and dosage that above fermented liquid uses is, 1-3 every day, time limit of service more than 1 month effect obvious.
Beneficial effect:
Shortened fermentation time, production cost is low, and result of use is good, has obtained extraordinary effect in the application of reality.
Concrete application and detected result are as follows:
Detect unit: Institute of Microorganism, Academia Sinica;
(2004) little searching detects for No. 095 and identifies responsible official: Zhou Yuguang
Find the needed multiple nutritional components of plant, comprised the various plants necessary composition of growing, also detected the sensitivity tests of product of the present invention and 42 kinds of medicines commonly used.
Detect unit: the Agricultural Science Institute, Yantai of Shandong Province;
People: Jiang Xueling is finished in test
Title: plant-growth microbial inoculum test summary report on cucumber
The result: test obtains through control group:
First group: the per mu yield net return increases by 766.5 yuan to 995.25 yuan
Second group: the per mu yield net return increases by 566.5 yuan to 916.8 yuan (obtaining through per mu yield and standard market price then)
Input-output ratio is respectively: 1: 2.83, and 1: 8.48,1: 61.12.
The different administration method cucumber production promoting width of cloth reaches more than 10%.
Application process is: underground dashing executed in conjunction with foliage-spray or independent foliage-spray.100 times of dilutions of fermented liquid.
Detect unit: the Agricultural Science Institute, Yantai of Shandong Province;
People: Jiang Xueling is finished in test
Title: plant-growth microbial inoculum test summary report on capsicum
The result: test obtains through control group:
Stimulation ratio reaches: 5-10.7%
Input-output ratio is respectively: 1: 3.1, and 1: 2.9,1: 29.5.
Drop into 1 yuan fermented liquid, can obtain 2.9 yuan of repayment, foliage-spray can obtain 29.5 yuan of repayment.
Application process is: underground dashing executed in conjunction with foliage-spray or independent foliage-spray.Foliage-spray is carried out in 100 times of dilutions of fermented liquid, guarantees continuous more than 3 times.
Detect unit: Accessories during Binzhou soil and fertilizer workstation;
People: Sun Fu is finished in test
Title: plant-growth microbial inoculum test summary report on cucumber
Time 2005.4.16 to 2005.7.25
The result: test obtains through control group:
The per mu yield net return increases by 436.24 yuan to 754.565 yuan (obtaining through per mu yield and standard market price then)
Input-output ratio is respectively: 1: 2.18, and 1: 6.76,1: 50.3.
The different administration method cucumber production promoting width of cloth reaches more than 10%.
Application process is: underground dashing executed in conjunction with foliage-spray or independent foliage-spray.100 times of dilutions of fermented liquid.
Detect unit: Accessories during Binzhou soil and fertilizer workstation;
People: Sun Fu is finished in test
Title: plant-growth microbial inoculum test summary report on capsicum
Time 2005.4.16 to 2005.7.31
The result: test obtains through control group:
Input-output ratio is respectively: 1: 2.9, and 1: 2.6,1: 25.8.
The different administration method capsicum volume increase width of cloth reaches more than 5%.
Detection is nuisanceless to people, animal.
Application process is: underground dashing executed in conjunction with foliage-spray or independent foliage-spray.100 times of dilutions of fermented liquid.
Survey report (Centra Lab, Shandong-Prov Academy of Agricutural Sciences)
The effect that following three kinds of agricultural chemicals is had degraded: SD-1750, effective cypermethrin, acephatemet missible oil, Chlorpyrifos 94, effect is very obvious.
The bacterial classification source:
In Penglai City, Shandong Province village in the epimere mud of market town Song Jia river.
Culture presevation number:
The common micro-organisms center C GMCC No.1562 of China Committee for Culture Collection of Microorganisms.
The preservation time: on December 12nd, 2005.
Separation method:
In getting 1 liter in korneforos mud the 7-8 month, be respectively charged in 22 500ml Erlenmeyer flasks for 10 liters with prepared culture according to the method described above, in the good bottle of bundle tampon plug, the external application old newspaper is wrapped, tighten with paper string, after placing high-temperature sterilization pot internal heating to 121 ℃ to carry out sterilization in 30 minutes, reduce to 35 ℃.Divide to go into 22 flask culture liquid with mud at leisure respectively fifty-fifty in the stirring of sterilisable chamber inner edge.Regulate pH6.4-7.5 with 10% lactic acid solution then, good with the tampon plug on the former bottle, wrap newspaper and tighten with paper string, place on the 25-35 ℃ of constant temperature shaking table, shaking speed per minute 50-60 commentaries on classics is shaken, and simultaneously to the incandescent light photograph, illumination requires every square metre of 100LX to get final product.Shaking table was cultivated after 40 hours, get the nutrient solution (getting upper strata liquid) of red reaction, move into culture dish (according to above-mentioned culture medium prescription preparation) at the indoor transfer pipet of inoculation, place under the above-mentioned culture condition and cultivate a week, and then from culture dish, in sterilisable chamber, get the bigger bacterium colony of red point and insert in the above-mentioned identical nutrient solution, enter and hold up strong the cultivation, carry out tens of times separation and Culture so repeatedly, hold up strong, the sampling microscopy, reaching does not have the purebred of assorted bacterium and can reach more than 5,000,000,000/milliliter at 24 hours fermentation index, being set in wavelength 525-530nm place mensuration light absorption value with spectrophotometer is 1.50, can be and produces with planting.
Production method:
Test tube or culture dish bacterial classification are inserted in sterilisable chamber (culture medium prescription in the Erlenmeyer flask is the same in the nutrient solution of 500ml Erlenmeyer flask, sterilization and temperature and pH value are the same), place on the shaking table constant temperature 25-35 ℃, shine with the 300LX incandescent light, shaking speed per minute 50-60 carried out 24 hours under changeing, sampling Detection inserts the 50-200L seeding tank with the inoculator after the sterilization more than 5,000,000,000/milliliter the time, inoculum size should be greater than 10%, the nutrient solution prescription of seeding tank is the same, surveying pH6.4-7.5 after the nutrient solution preparation is finished is advisable, transfer the NaOH solution of pH with lactic acid solution or 12N, sterilisation temp carried out 30 minutes for 121 ℃.Inoculation when being cooled to 35 ℃ toward the logical cold water of tank body chuck then, inoculation is led to pressurized air after the into filtration sterilization after finishing and is entered from the bottom of jar and stir, temperature in jar is 25-35 ℃, and intake pressure 0.1-0.3Mpa is advisable about jar internal pressure 0.03-0.08Mpa.Equally with the visor illumination 300LX of incandescent light by two different angles.The adjustment of jar internal pressure is from the purging valve control of tank top.Constant temperature 25-35 ℃ of fermentation is after 24 hours; be forwarded to secondary seed jar (nutrient solution and the first class seed pot of secondary seed jar are identical) by first class seed pot (200L) and secondary kind in the connecting tube of jar (2000L) as stated above again and carried out enlarged culturing 24 hours; culture temperature wherein, pH value, illumination and venting pressure are identical with first class seed pot; after secondary seed jar fermentation culture is finished, be forwarded in the same way and carry out the mass-producing fermentation in the 10000L large fermentation tank.Base material formulation in the large fermentation tank as hereinbefore, just used distilled water is made into the membrane filtration water purification, the condition that changes large fermentation tank over to is as follows: by the illumination that the visor of three different angles is given incandescent light 1000LX, material liquid pH 6.4-7.5, intake pressure 0.3Mpa, adjust a jar internal pressure 0.05-0.08Mpa, 35 ℃ of controlled temperature fermented 24 hours, and sampling in per 2 hours detects, stop fermentation immediately when finding microbiological contamination in the fermentation, the finished product bacterium liquid of fermentation is hundred million/milliliter of 40-50.The fermented liquid that obtains is agricultural and uses and aquaculture and domestic birds and animals usefulness, functional oral liquid of the present invention.
Embodiment 1
The compound method of fermented liquid nutrient solution:
Inorganic salt solution proportioning in the 1000ml distilled water;
MgSO
47H
2O 0.3 gram
CaCl2H
2O 0.05 gram
FeSO
47H
2O 0.5 gram
NaEDTA 0.5 gram
Trace element solution proportioning in the 250ml distilled water;
H
3BO
41.0 gram
MnSO
4120 milligrams
NaMoO
460 milligrams
ZnSO
415 milligrams
Cu (NO
3)
22 milligrams
12 milligrams of Sodium orthomolybdates
12 milligrams of Sodium Selenites;
Growth factor solution proportioning in the 100ml distilled water
0.5 milligram of vitamin H
Vitamins B
110 milligrams
10 milligrams in nicotinic acid
10 milligrams of para-amino benzoic acid;
The lactic acid solution proportioning
900 milliliters of distilled water
40 milliliters of lactic acid are transferred pH6.4 with the NaOH solution of 12N;
The ammoniumsulphate soln proportioning
800 milliliters of distilled water
Ammonium sulfate 5 grams
Water adds to 1000 milliliters;
Add to 1000 milliliters with distilled water the dissolving back, transfers the pH6.4 phosphoric acid buffer with the NaOH solution of 12N then
K
2HPO
440 grams
KH
2PO
410 grams
Water adds to 1000 milliliters;
Get respectively:
A, inorganic salt solution 20%,
B, trace element solution 0.2%
C, growth factor solution 0.1%
D, lactic acid solution 5%
E, ammoniumsulphate soln 1.5%
F, phosphoric acid buffer 3%
Proportioning a-f is mixed according to above-mentioned per-cent, and all the other are water, promptly obtain nutrient solution.
Add in addition when doing slant tube or culture dish substratum: peptone 1%, yeast extract paste 0.2%.
Embodiment 2
Inorganic salt solution proportioning in the 1000ml distilled water;
MgSO
47H
2O 3.0 grams
CaCl2H
2O 1.0 grams
FeSO
47H
2O 1.0 grams
NaEDTA 1.0 grams
Trace element solution proportioning in the 250ml distilled water;
H
3BO
41.5 gram
MnSO
4200 milligrams
NaMoO
4100 milligrams
ZnSO
420 milligrams
Cu (NO
3)
23 milligrams
15 milligrams of Sodium orthomolybdates
15 milligrams of Sodium Selenites;
Growth factor solution proportioning in the 100ml distilled water
0.5 milligram of vitamin H
Vitamins B
130 milligrams
30 milligrams in nicotinic acid
20 milligrams of para-amino benzoic acid;
The lactic acid solution proportioning
900 milliliters of distilled water
100 milliliters of lactic acid are transferred pH6.4 with the NaOH solution of 12N;
The ammoniumsulphate soln proportioning
800 milliliters of distilled water
Ammonium sulfate 8 grams
Water adds to 1000 milliliters;
Add to 1000 milliliters with distilled water the dissolving back, transfers the pH6.4 phosphoric acid buffer with the NaOH solution of 12N then
K
2HPO
450 grams
KH
2PO
415 grams
Water adds to 1000 milliliters;
Get respectively:
A, inorganic salt solution 30%,
B, trace element solution 1.0%
C, growth factor solution 0.1%
D, lactic acid solution 10%
E, ammoniumsulphate soln 2%
F, phosphoric acid buffer 5%
Proportioning a-f is mixed according to above-mentioned per-cent, and all the other are water, promptly obtain nutrient solution.
Add in addition when doing slant tube or culture dish substratum: peptone 1%, yeast extract paste 0.2%.
Embodiment 3
Inorganic salt solution proportioning in the 1000ml distilled water;
MgSO
47H
2O 0.2 gram
CaCl2H
2O 0.1 gram
FeSO
47H
2O 0.1 gram
NaEDTA 0.1 gram
Trace element solution proportioning in the 250ml distilled water;
H
3BO
40.5 gram
MnSO
450 milligrams
NaMoO
420 milligrams
ZnSO
410 milligrams
Cu (NO
3)
22 milligrams
6 milligrams of Sodium orthomolybdates
6 milligrams of Sodium Selenites;
Growth factor solution proportioning in the 100ml distilled water
0.2 milligram of vitamin H
Vitamins B
110 milligrams
10 milligrams in nicotinic acid
10 milligrams of para-amino benzoic acid;
The lactic acid solution proportioning
900 milliliters of distilled water
30 milliliters of lactic acid are transferred pH6.4 with the NaOH solution of 12N;
The ammoniumsulphate soln proportioning
800 milliliters of distilled water
Ammonium sulfate 3 grams
Water adds to 1000 milliliters;
Add to 1000 milliliters with distilled water the dissolving back, transfers the pH6.4 phosphoric acid buffer with the NaOH solution of 12N then
K
2HPO
415 grams
KH
2PO
410 grams
Water adds to 1000 milliliters;
Get respectively:
A, inorganic salt solution 10%,
B, trace element solution 0.1%
C, growth factor solution 0.1%
D, lactic acid solution 2%
E, ammoniumsulphate soln 0.5%
F, phosphoric acid buffer 1%
Proportioning a-f is mixed according to above-mentioned per-cent, and all the other are water, promptly obtain nutrient solution.
Add in addition when doing slant tube or culture dish substratum: peptone 1%, yeast extract paste 0.2%.
Embodiment 4
Inorganic salt solution proportioning in the 1000ml distilled water;
MgSO
47H
2O 3.0 grams
CaCl2H
2O 0.1 gram
FeSO
47H
2O 1.0 grams
NaEDTA 1.0 grams
Trace element solution proportioning in the 250ml distilled water;
H
3BO
40.5 gram
MnSO
4200 milligrams
NaMoO
420 milligrams
ZnSO
410 milligrams
Cu (NO
3)
23 milligrams
15 milligrams of Sodium orthomolybdates
6 milligrams of Sodium Selenites;
Growth factor solution proportioning in the 100ml distilled water
0.2 milligram of vitamin H
Vitamins B
130 milligrams
30 milligrams in nicotinic acid
10 milligrams of para-amino benzoic acid;
The lactic acid solution proportioning
900 milliliters of distilled water
500 milliliters of lactic acid are transferred pH6.4 with the NaOH solution of 12N;
The ammoniumsulphate soln proportioning
800 milliliters of distilled water
Ammonium sulfate 5 grams
Water adds to 1000 milliliters;
Add to 1000 milliliters with distilled water the dissolving back, transfers the pH6.4 phosphoric acid buffer with the NaOH solution of 12N then
K
2HPO
430 grams
KH
2PO
412 grams
Water adds to 1000 milliliters;
Get respectively:
A, inorganic salt solution 15%,
B, trace element solution 0.5%
C, growth factor solution 0.1%
D, lactic acid solution 5%
E, ammoniumsulphate soln 1.5%
F, phosphoric acid buffer 3%
Proportioning a-f is mixed according to above-mentioned per-cent, and all the other are water, promptly obtain nutrient solution.
Add in addition when doing slant tube or culture dish substratum: peptone 1%, yeast extract paste 0.2%.
The application of fermented liquid:
1, is equipped with 3% Flos Sophorae honey with fermented liquid and can makes immunomodulatory, the oral liquid of maintenance intestinal function.
2, the Alternanthera sessilis (L.) DC. extracting solution connects the blood sugar reducing liquid that fermented liquid that secondary seed fermentation finishes is blood sugar regulation.
3, adjust taste with fermented liquid and pure water with foodstuff additive and be biologic beverage.
4, be directly used in plant nutrition liquids such as pollution-free vegetable, fruit with fermented liquid, can produce SOD vegetables and fruit.
5, the root rot of fruit trees such as apple tree, cherry tree, apricot, pear tree is irrigated butt rot place with 5-10 times of fermented liquid and can be treated root rot.
6, application of fermentation liquid and water are according to 1: 5-10 adds doubly in the feed that is added to milk cow, pig, chicken or the tap water and feeds, and can make milk cow improve the rate 10-20% of giving milk, and makes domestic birds and animals improve immunizing power, reduces disease, accelerates growth.
7, give breeded fish, the 3-5ppm by the pond water yield in the pond of fishery products such as shrimp, crab adds, can purify water, reduce disease, improve the speed of growth.
The product of producing with fermented liquid of the present invention has human body is had no side effect, and can play the effect of adjusting to Immune System, and is very favourable to HUMAN HEALTH.