CN103435167A - Biological decolorizing method of melanoidin pigment - Google Patents
Biological decolorizing method of melanoidin pigment Download PDFInfo
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- CN103435167A CN103435167A CN2013104235158A CN201310423515A CN103435167A CN 103435167 A CN103435167 A CN 103435167A CN 2013104235158 A CN2013104235158 A CN 2013104235158A CN 201310423515 A CN201310423515 A CN 201310423515A CN 103435167 A CN103435167 A CN 103435167A
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- melanoidin
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Abstract
The invention discloses a biological decolorizing method of melanoidin pigment. According to the technical scheme, aspergillus tubingensis is used for decolorizing the melanoidin pigment. The biological decolorizing method comprises the following steps of: culturing a strain with preservation number of CGMCC No. 7174 for 3 to 5 days at the temperature of 29-31 DEG C so as to prepare the aspergillus tubingensis, then inoculating aspergillus tubingensis spore suspension into a melanoidin pigment liquid medium, and carrying out shake cultivation for 4 to 10 days under the condition of controlling the temperature at 30-50 DEG C, the pH at 3.0-8.0 and the rotating speed at 50-350rpm for decolorizing reaction. The biological decolorizing method can be used for directly decolorizing the melanoidin pigment and has the advantages of safety, effectiveness, mild conditions, little secondary pollution, environmental protection, economy and the like; the decolorizing rate of the melanoidin pigment can reach up to 75%; a good strain and biological treatment method is provided for practical industrial wastewater treatment.
Description
Technical field
The invention belongs to the biological wastewater treatment field, particularly a kind of melanoidin pigment biodecolouring process.
Background technology
The melanoidin pigment is sugar and amino acid generation Maillard reaction and the class brown polymkeric substance that forms, belong to that to take small peptide mixture or protein be the polymer mixture that compound that basic skeleton structure, the polymerization degree are not high forms, excessive, there is strong anti-oxidation and cytotoxicity when edible.The main component of its waste water high chroma that to be sugar-refining industry and extension industry thereof produce as the molasses-spirit industry, its heatproof, fast light photograph, extend its colour storage period and do not subtract.It is higher that physico-chemical process removes the pigment cost, subdue as means, pollutent and there are the advantages such as safe and effective, mild condition, secondary pollution are few, environmental protection and economy as the decolouring of the microorganism of purpose and take microbial metabolism, can only remove organism wherein but traditional activated sludge process is disposed this type of waste water, can't effectively remove pigment.Therefore finding special-purpose decolouring microorganism is one of possible approaches improved the microorganism decolorizing efficiency.
Tabin aspergillus (Aspergillus Tubingensis) is a kind of fungi that belongs to Eurotiale Trichocomaceae Aspergillus, can be grown on the base things such as soil, hami melon, agaric, cow dung.Bacterium colony ramp on Cha Shi agar, cultivate 7 days diameter 50～75mm for 25 ℃, and this bacterium colony is grown the fastest on wort agar.Tabin aspergillus quality velvet shape, have a small amount of sterility growth, and the conidium structure is a large amount of, is filbert or black, and without transudate, the bacterium colony reverse side is tawny.
The report of external existing many related microorganisms.Aspect bacterium, as having studied plant lactobacillus, Tondee etc. can reach 68% to initial pH6.0 the highest percent of decolourization that adds the melanoidin pigment solution of nutritive substance; The one thread non-heterocyst Bollinger body in the strain ocean cyanobacteria that quivers is being cultivated 75% the melanoidin pigment solution of can decolouring in 30 days; Kumar etc. have studied bacillus thuringiensis, and bacillus brevis and genus bacillus are to four kinds of different decolorizing effects of synthetic melanoidin.The decolorizing effect of fungi is generally good than bacterium, and at present bibliographical information has Coriolous Dersicolor (Fr.) Quel fungus, Phanerochaete chrysosporium, aspergillus niger, Aspergillus fumigatus, flavus, a streptomycete etc.Percent of decolourization is higher the Coriolous Dersicolor (Fr.) Quel fungus of the report such as Watanabe, under top condition, to synthetic melanoidin percent of decolourization, is 80%.Some aspergillus niger percent of decolourization when glucose exists can reach 65%, if can not decolour without glucose.Murata etc. separate and obtain a streptomycete TT14 from soil, and under optimal culture condition, percent of decolourization is 64%.And at present, the domestic research in this field decolouring as molasses alcohol waste water in actual waste water that focuses mostly on.
Summary of the invention
The present invention is directed to above-mentioned technical problem, disclose a kind of method of utilizing Tabin aspergillus to be decoloured to the melanoidin pigment, to reach the purpose to melanoidin pigment direct bleaching, make melanoidin pigment decolouring more direct, easy handling; The present invention further makes the diversification more of melanoidin pigment decoloring method.
The invention provides a kind of melanoidin pigment biodecolouring process, technical scheme is as follows:
A kind of melanoidin pigment biodecolouring process, used Tabin aspergillus to be decoloured to the melanoidin pigment.
This technical scheme comprises the steps:
(1) Tabin aspergillus is that preserving number is that the bacterial classification of CGMCC.NO.7174 is to cultivate cultivation in 3～5 days under 29～31 ℃ in temperature, prepares spore suspension with Tween80 eluant solution spore and makes.
(2) the melanoidin pigment is for making and be diluted to A by oneself
475=3.5 make, and add in addition nutrition source, spore suspension is accessed to shaking culture in melanoidin pigment liquid nutrient medium and within 4～10 days, carry out decoloring reaction, after reaction finishes, thalline are carried out to the suction filtration separation.
Preferably, in step (1), melanoidin pigment substratum miospore suspension inoculation amount is 10
3～10
6individual/mL.
Preferably, in step (2), the decoloring reaction temperature is 30～50 ℃.
Preferably, in step (2), decoloring reaction pH is 3.0～8.0.
Preferably, in step (2), the decoloring reaction rotating speed is 50～350rpm.
Preferably, Ensure Liquid source, step (2) China and foreign countries is a kind of, peptone and the NaNO in glucose, fructose and sucrose
3in a kind of, KH
2pO
4, MgSO
47H
2o.
Preferably, additional nutrition source consumption is respectively: a kind of 5～30g/L in glucose, fructose and sucrose, peptone and NaNO
3in a kind of 1.0～5.0g/L, KH
2pO
40.5～1.5g/L, MgSO
47H
2o0.05～0.5g/L.
Wherein, after Tabin aspergillus prepares spore suspension with Tween80 eluant solution spore, use the blood counting chamber counting; Reaction is tested the variation of 475nm place absorbancy and is calculated percent of decolourization after finishing by visible spectrophotometer.
Beneficial effect of the present invention:
The advantages such as that the present invention has is safe and effective, mild condition, secondary pollution are few, environmental protection and economy.And, but the invention provides a kind of bacterial classification of efficient decolorizing melanoidin pigment, the highest percent of decolourization that this bacterial classification is cultivated the melanoidin pigment in 4 days can reach 75%; On the other hand, the subject range of this bacterial strain is wider, under the slow speed of revolution or low pH, still has certain decoloring ability.Bacterial classification provided by the invention and decoloration process have good biological decolouring efficiency and potential economic benefit, can be the actual industrial wastewater treatment a kind of good bacterial classification and bioremediation are provided.Further make the diversification more of melanoidin pigment decoloring method.
Embodiment
Further describe the present invention referring to embodiment, can implement according to this with reference to the specification sheets word to make those skilled in the art, protection domain of the present invention is not limited by embodiments of the present invention.The spawn culture that the Tabin aspergillus used is CGMCC.NO.7174 for preserving number forms.
Embodiment 1
(1) the melanoidin pigment being put into to liquid amount is that 60ml/250ml Oscillating bottle is diluted to A
475=3.5, additional sucrose 30g/L, NaNO
31.5g/L, MgSO
4.7H
2o0.05g/L, KH
2pO
41.0g/L, and pH is adjusted to 4.0.
(2) take out the Tabin aspergillus bacterial strain switching PDA test tube slant substratum of preservation, putting into biochemical cultivation case maintenance temperature is 30 ℃ of cultivations 5 days.Repeatedly rinse spore with sterilized 0.5% tween 80 solution, add the granulated glass sphere vibrating dispersion, filter and obtain spore suspension, blood counting chamber is measured spore suspension concentration.
(3) fixedly inoculating spores quantity is 4.3 * 10
4individual/ml, calculate the required spore suspension volume that adds with this.Under aseptic condition, the spore suspension of getting respective volume is inoculated in previously prepared good melanoidin pigment liquid nutrient medium, temperature is 39 ℃, 150rpm shaking culture 6d, take out shaking flask, the suction filtration separating thallus, get 1mL filtrate and use 0.1M, the acetic acid that pH is 5.0-10 times of sodium acetate buffer dilutions, visible spectrophotometer is surveyed the variation of 475nm place, decolouring front and back absorbancy.Thalline is put into 105 ℃, baking oven and is dried 24 hours, then with electronic analytical balance, weighs.Recording percent of decolourization is 53%, and dry cell weight is 3.01g/L.
Embodiment 2
(1) the melanoidin pigment being put into to liquid amount is that 50ml/250ml Oscillating bottle is diluted to A
475=3.5, the fructose of additional 5g/L, NH
4cl2.0g/L, MgSO
4.7H
2o0.5g/L, KH
2pO
40.5g/L, and pH is adjusted to 7.0.
(2) take out the Tabin aspergillus bacterial strain switching PDA test tube slant substratum of preservation, putting into biochemical cultivation case maintenance temperature is 29 ℃ of cultivations 4 days.Repeatedly rinse spore with sterilized 0.5%Tween80 solution, add the granulated glass sphere vibrating dispersion, filter and obtain spore suspension, blood counting chamber is measured spore suspension concentration.
(3) fixedly inoculating spores quantity is 4.3 * 10
5individual/ml, calculate the required spore suspension volume that adds with this.Under aseptic condition, the spore suspension of getting respective volume is inoculated in previously prepared good melanoidin pigment liquid nutrient medium, temperature is 30 ℃, 350rpm shaking culture 4d, take out shaking flask, the suction filtration separating thallus, get 1mL filtrate and use 0.1M, the acetic acid of pH5.0-10 times of sodium acetate buffer dilutions, visible spectrophotometer is surveyed the variation of 475nm place, decolouring front and back absorbancy.Thalline is put into 105 ℃, baking oven and is dried 24 hours, then with electronic analytical balance, weighs.Recording percent of decolourization is 30%, and dry cell weight is 2.5g/L.
Embodiment 3
(1) the melanoidin pigment being put into to liquid amount is that 50ml/250ml Oscillating bottle is diluted to A
475=3.5, additional fructose 22g/L, peptone 1.5g/L, MgSO
4.7H
2o0.25g/L, KH
2pO
41.5g/L pH is adjusted to 8.0.
(2) take out the Tabin aspergillus bacterial strain switching PDA test tube slant substratum of preservation, putting into biochemical cultivation case maintenance temperature is 31 ℃ of cultivations 3 days.Repeatedly rinse spore with sterilized 0.5%Tween80 solution, add the granulated glass sphere vibrating dispersion, filter and obtain spore suspension, blood counting chamber is measured spore suspension concentration.
(3) fixedly inoculating spores quantity is 4.3 * 10
4individual/ml, calculate the required spore suspension volume that adds with this.Under aseptic condition, the spore suspension of getting respective volume is inoculated in previously prepared good melanoidin pigment liquid nutrient medium, temperature is 40 ℃, 150rpm shaking culture 6d, take out shaking flask, the suction filtration separating thallus, get 1mL filtrate and use 0.1M, pH5.0 acetic acid-10 times of sodium acetate buffer dilutions, visible spectrophotometer is surveyed the variation of 475nm place, decolouring front and back absorbancy.Thalline is put into 105 ℃, baking oven and is dried 24 hours, then with electronic analytical balance, weighs.Recording percent of decolourization is 65%, and dry cell weight is 3.26g/L.
Embodiment 4
(1) the melanoidin pigment being put into to liquid amount is that 50ml/250ml Oscillating bottle is diluted to A
475=3.5, additional glucose 30g/L, NaNO
33.5g/L, MgSO
4.7H
2o0.5g/L, KH
2pO
41.0g/L, and pH is adjusted to 6.0.
(2) take out the Tabin aspergillus bacterial strain switching PDA test tube slant substratum of preservation, putting into biochemical cultivation case maintenance temperature is 30 ℃ of cultivations 5 days.Repeatedly rinse spore with sterilized 0.5%Tween80 solution, add the granulated glass sphere vibrating dispersion, filter and obtain spore suspension, blood counting chamber is measured spore suspension concentration.
(3) fixedly inoculating spores quantity is 4.3 * 10
4individual/ml, calculate the required spore suspension volume that adds with this.Under aseptic condition, the spore suspension of getting respective volume is inoculated in previously prepared good melanoidin pigment liquid nutrient medium, temperature is 50 ℃, 50rpm shaking culture 8d, take out shaking flask, the suction filtration separating thallus, get 1mL filtrate and use 0.1M, pH5.0 acetic acid-10 times of sodium acetate buffer dilutions, visible spectrophotometer is surveyed the variation of 475nm place, decolouring front and back absorbancy.Thalline is put into 105 ℃, baking oven and is dried 24 hours, then with electronic analytical balance, weighs.Recording percent of decolourization is 40%, and dry cell weight is 2.2g/L.
Embodiment 5
(1) the melanoidin pigment being put into to liquid amount is that 50ml/250ml Oscillating bottle is diluted to A
475=3.5, additional glucose 30g/L, NaNO
31.5g/L, MgSO
4.7H
2o0.5g/L, KH
2pO
41.0g/L pH is adjusted to 6.0.
(2) take out the Tabin aspergillus bacterial strain switching PDA test tube slant substratum of preservation, putting into biochemical cultivation case maintenance temperature is 30 ℃ of cultivations 5 days.Repeatedly rinse spore with sterilized 0.5%Tween80 solution, add the granulated glass sphere vibrating dispersion, filter and obtain spore suspension, blood counting chamber is measured spore suspension concentration.
(3) fixedly inoculating spores quantity is 4.3 * 10
4individual/ml, calculate the required spore suspension volume that adds with this.Under aseptic condition, the spore suspension of getting respective volume is inoculated in previously prepared good melanoidin pigment liquid nutrient medium, adds CaCO
3particle, temperature is 39 ℃, and 150rpm shaking culture 8d takes out shaking flask, and the suction filtration separating thallus, get 1mL filtrate and use 0.1M, pH5.0 acetic acid-10 times of sodium acetate buffer dilutions, visible spectrophotometer is surveyed the variation of 475nm place, decolouring front and back absorbancy.Thalline is put into 105 ℃, baking oven and is dried 24 hours, then with electronic analytical balance, weighs.Recording percent of decolourization is 75%, and dry cell weight is 4.2g/L.
Embodiment 6
(1) the melanoidin pigment being put into to liquid amount is that 50ml/250ml Oscillating bottle is diluted to A
475=3.5, additional glucose 30g/L, NaNO
35.0g/L, MgSO
4.7H
2o0.5g/L, KH
2pO
41.0g/L, and pH is adjusted to 6.0.
(2) take out the Tabin aspergillus bacterial strain switching PDA test tube slant substratum of preservation, putting into biochemical cultivation case maintenance temperature is 29 ℃ of cultivations 5 days.Repeatedly rinse spore with sterilized 0.5%Tween80 solution, add the granulated glass sphere vibrating dispersion, filter and obtain spore suspension, blood counting chamber is measured spore suspension concentration.
(3) fixedly inoculating spores quantity is 4.3 * 10
4individual/ml, calculate the required spore suspension volume that adds with this.Under aseptic condition, the spore suspension of getting respective volume is inoculated in previously prepared good melanoidin pigment liquid nutrient medium, 30 ℃, add glucose 20g/L after 50rpm shaking culture 3d, within the 8th day, take out shaking flask, the suction filtration separating thallus, get 1mL filtrate and use 0.1M, pH5.0 acetic acid-10 times of sodium acetate buffer dilutions, visible spectrophotometer is surveyed the variation of 475nm place, decolouring front and back absorbancy.Thalline is put into 105 ℃, baking oven and is dried 24 hours, then with electronic analytical balance, weighs.Recording percent of decolourization is 42%, and dry cell weight is 2.8g/L.
In above-described embodiment, find out, the method for utilizing Tabin aspergillus to be decoloured to the melanoidin pigment, reached the purpose to melanoidin pigment direct bleaching, make melanoidin pigment decolouring more direct, easy handling; Further in embodiment, can find out that reaction conditions of the present invention is gentle, secondary pollution is few, environmental protection and economy, and efficient to the decolouring of melanoidin pigment.
Claims (8)
1. a melanoidin pigment biodecolouring process, is characterized in that: use Tabin aspergillus to be decoloured to the melanoidin pigment.
2. melanoidin pigment biodecolouring process according to claim 1, is characterized in that: comprise the steps:
(1) Tabin aspergillus is that preserving number is that the bacterial classification of CGMCC.NO.7174 is to cultivate cultivation in 3～5 days under 29～31 ℃ in temperature, prepares spore suspension with Tween80 eluant solution spore and makes.
(2) the melanoidin pigment is for making and be diluted to A by oneself
475=3.5 make, and add in addition nutrition source, spore suspension is accessed to shaking culture in melanoidin pigment liquid nutrient medium and within 4～10 days, carry out decoloring reaction, after reaction finishes, thalline are carried out to the suction filtration separation.
3. melanoidin pigment biodecolouring process according to claim 2, it is characterized in that: in step (1), melanoidin pigment substratum miospore suspension inoculation amount is 10
3～10
6individual/mL.
4. melanoidin pigment biodecolouring process according to claim 2, it is characterized in that: described in step (2), the decoloring reaction temperature is 30～50 ℃.
5. melanoidin pigment biodecolouring process according to claim 2, it is characterized in that: described in step (2), decoloring reaction pH is 3.0～8.0.
6. melanoidin pigment biodecolouring process according to claim 2, it is characterized in that: described in step (2), the decoloring reaction rotating speed is 50～350rpm.
7. melanoidin pigment biodecolouring process according to claim 2 is characterized in that: additional nutrition source is a kind of, peptone and the NaNO in glucose, fructose and sucrose
3in a kind of, KH
2pO
4, MgSO
47H
2o.
8. melanoidin pigment biodecolouring process according to claim 7, it is characterized in that: additional nutrition source consumption is respectively: a kind of 5～30g/L in glucose, fructose and sucrose, peptone and NaNO
3in a kind of 1.0～5.0g/L, KH
2pO
40.5～1.5g/L, MgSO
47H
2o0.05～0.5g/L.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106644807A (en) * | 2016-09-19 | 2017-05-10 | 四川大学 | Model for testing black, white and yellow yeast ratio in yeast and method thereof |
| CN109626598A (en) * | 2019-01-23 | 2019-04-16 | 南华大学 | A method of utilizing Tabin aspergillus and phytate in-situ immobilization hexavalent uranium polluted surface water |
| CN114107416A (en) * | 2021-11-19 | 2022-03-01 | 广东容大生物股份有限公司 | Development and industrialization research of bioactive peptide |
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| CN102839128A (en) * | 2011-06-23 | 2012-12-26 | 广西大学 | Microorganism used for decolouring molasses alcohol waste water and decolouring method |
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2013
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Patent Citations (4)
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| WO2000005349A1 (en) * | 1998-07-21 | 2000-02-03 | Unilever N.V. | Phenol oxidizing enzymes |
| JP2003304890A (en) * | 2002-04-15 | 2003-10-28 | Ajinomoto Co Inc | Method for producing erasable dye with microorganism, erasable dye produced thereby and erasable ink containing erasable dye |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106644807A (en) * | 2016-09-19 | 2017-05-10 | 四川大学 | Model for testing black, white and yellow yeast ratio in yeast and method thereof |
| CN109626598A (en) * | 2019-01-23 | 2019-04-16 | 南华大学 | A method of utilizing Tabin aspergillus and phytate in-situ immobilization hexavalent uranium polluted surface water |
| CN114107416A (en) * | 2021-11-19 | 2022-03-01 | 广东容大生物股份有限公司 | Development and industrialization research of bioactive peptide |
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| CN103435167B (en) | 2015-11-18 |
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Inventor after: Liu Youyan Inventor after: Lan Lina Inventor after: Cheng Ning Inventor after: Tang Aixing Inventor before: Liu Youyan |
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