CN104962549B - A kind of Rhodopseudomonas palustris synchronizes method and the application of concentration - Google Patents
A kind of Rhodopseudomonas palustris synchronizes method and the application of concentration Download PDFInfo
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- CN104962549B CN104962549B CN201510443585.9A CN201510443585A CN104962549B CN 104962549 B CN104962549 B CN 104962549B CN 201510443585 A CN201510443585 A CN 201510443585A CN 104962549 B CN104962549 B CN 104962549B
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Abstract
The invention discloses a kind of methods that Rhodopseudomonas palustris synchronizes concentration, include the following steps：Step 1: culture medium is prepared and sterilization；Step 2: inoculated and cultured；Step 3: thalline synchronization process；Step 4: physical concentration is handled.The Rhodopseudomonas palustris handled through this method is up to 7.3 × 1012A/g, thalli morphology is uniform, and growth is in contemporaneity, efficiently brings back to life；It is good by high concentration Rhodopseudomonas palustris palatability made from the above method the present invention discloses the application that a kind of Rhodopseudomonas palustris synchronizes concentration, it can be used as biological feed early period in cultivation, albumen and B family vitamin be provided；In the cultivation middle and later periods, it can be used for regulating and controlling water quality, Rhodopseudomonas palustris of periodically splashing is conducive to the lower water column in cultivation and forms dominant bacteria phase, effectively inhibits pathogenic bacteria.
Description
Technical field
The present invention relates to the technical fields of probiotics more particularly to a kind of Rhodopseudomonas palustris to synchronize concentration
Method and application.
Background technology
One kind that Rhodopseudomonas palustris (Rhodop seudanonas palustris) belongs to photosynthetic bacteria, belongs to red
Spirillaceae, Rhodopseudomonas, cell is short or elongated rod shape, arc or spiral shape；It is bred in a manner of binary fission or budding.This bacterium contains
There are the carotenoid of antibiotics sensitivity test or b and monoid 1,2,4, and some carotenoids being not yet included in above-mentioned monoid
Element.A kind of water quality cleansing agent is can be used as, it is facultative good foster due to Rhodopseudomonas palustris photoheterotrophy, in illumination anaerobism or dark
It is micro- it is aerobic under conditions of can grow, for reduce ammonia nitrogen, nitrite, COD contents, and improve dissolved oxygen have notable effect
Fruit；As a kind of feed addictive, thalline is rich in mycoprotein and B family vitamin, and palatability is good, full of nutrition, is supported in aquatic products
It grows in industry and has broad application prospects.
Currently, commercially available photosynthetic bacteria type is various, character is liquid mostly.The Rhodopseudomonas palustris that routine culture goes out has
Following disadvantage：1, thalline different stages of growth mix, and the speed of growth is difficult to control；2, thalline free settling is layered, and influences product
Shape；3, resurrection rate is low, after incubation the phase, and culture medium exhaustion leads to thalline mortality, secondary metabolite（Such as sulphur）Accumulation, poison
Evil somatic cells.In view of this, the present inventor's research and devise a kind of Rhodopseudomonas palustris synchronize concentration method and
Using thus this case generates.
Invention content
The purpose of the present invention is to provide method and application that a kind of Rhodopseudomonas palustris synchronizes concentration, the present invention is logical
It crosses and red-blue illumination adjustment is carried out to the growth cycle of Rhodopseudomonas palustris, slow down bacterial metabolism activity, promote all thalline thin
Born of the same parents' form is consistent, promotes bacterium-liquid to detach by adding macromolecules adsorption material, to obtain highly enriched bacterium mud, add carrier and
Drier is allowed to fully dispersed, is finally made the powder forms of high quality.
To achieve the above object, the present invention the technical solution to solve the technical problem is that：
A kind of method that Rhodopseudomonas palustris synchronizes concentration, includes the following steps：
Step 1: Rhodopseudomonas palustris culture
Triangular flask culture weighs sodium acetate 8g/l, ammonium chloride 2g/l, yeast extract 3g/l, bittern 5ml/l, calcium chloride 0.5g/
L, potassium fulvate 0.8g/l, dipotassium hydrogen phosphate 1g/l, with a concentration of 3% dioxygen water sterilization 30 minutes after the above reagent dissolving；Sterilizing
It finishes and sodium bicarbonate 2g/l dissolvings is added, be fitted into 1L triangular flasks；Alternatively,
Large-scale culture weighs sodium acetate 8kg/t, 2 kg/t of ammonium chloride, 3 kg/t of yeast extract, 5 L/t of bittern, calcium chloride
0.5 kg/t, 0.8 kg/t of potassium fulvate, 1 kg/t of dipotassium hydrogen phosphate, with a concentration of 3% dioxygen water sterilization after the above reagent dissolving
30 minutes；Sterilizing, which finishes, is added sodium bicarbonate 2 kg/t dissolvings, is added in polybag；
Step 2: inoculated and cultured
It is inoculated with Rhodopseudomonas palustris bacterium solution by 20% inoculum concentration, 32 DEG C, 3000-5300lx incandescent lights are trained under the conditions of shining
72h-96h is supported, culture solution is diluted 100 times, OD values are measured at 660nm and reach 1.8 or more；Utensil goes out used in inoculation
Bacterium is handled, seeded process sterile working；
Step 3: thalline synchronization process
Culture solution is placed under dark condition, not contact nature light, by feux rouges:Blue light ratio is 4：1~1：1 arranges in pairs or groups
3~5h is irradiated in irradiation；Purpose is that Rhodopseudomonas palustris metabolic activity is made gradually to slow down, and form gradually reaches unanimity；
Step 4: physical concentration is handled
Polyacrylamide amine cation, sodium carboxymethylcellulose, Guar are added into the culture solution after above-mentioned synchronization process
The one or more of bean gum, xanthans, chitosan, ratio are 0.5%~1.5%, and the place of being protected from light preserves 3.5-5 hours, removes supernatant
Liquid, you can obtain hyperconcetration Rhodopseudomonas palustris, cell number is up to 7.3 × 1012A/g.
A kind of Rhodopseudomonas palustris synchronizes the application of concentration, takes the above-mentioned Rhodopseudomonas palustris after physical concentration
Protective agent polyethylene glycol 400, the one or more of trehalose, glycerine, sodium glutamate, Tween 80, addition is added in portion thereto
Amount is 8%~35%, is uniformly mixed to get Rhodopseudomonas palustris hyperconcetration active bacteria formulation；This product character is liquid, effective marsh
Red pseudomonas number is up to 7 × 1011 A/ml.
A kind of Rhodopseudomonas palustris synchronizes the application of concentration, takes the Rhodopseudomonas palustris one after above-mentioned physical concentration
Part, a or several part in silica, sodium sulphate, calcium oxide, maize cob meal and magnesium sulfate is added thereto, cold nebulization is dry
Yeast powder portion is added to three parts in dry processing, pulvis after drying；Rhodopseudomonas palustris is made and concentrates bacterium powder；According to said method make
Powder product in Rhodopseudomonas palustris cell number up to 5.0 × 109 A/g.
The difference of red blue light spectrum wavelength is absorbed and utilized using Rhodopseudomonas palustris by the present invention（Feux rouges dominant wavelength ranges
The nm of 610nm~720, blue light dominant wavelength ranges 400nm~520 nm）, by the growth cycle to Rhodopseudomonas palustris into
Row red-blue illumination adjusts, and slows down bacterial metabolism activity, promotes all somatic cells forms consistent；By adding macromolecules adsorption
Material promotes bacterium-liquid separation, to obtain highly enriched bacterium mud.Carrier is added into the Rhodopseudomonas palustris bacterium solution after above-mentioned concentration
It is allowed to fully dispersed with appropriate drier, makes powder forms.
Specific implementation mode
Embodiment 1
A kind of method that Rhodopseudomonas palustris synchronizes concentration, includes the following steps：
Step 1: Rhodopseudomonas palustris culture
Triangular flask culture weighs sodium acetate 8g/l, ammonium chloride 2g/l, yeast extract 3g/l, bittern 5ml/l, calcium chloride 0.5g/
L, potassium fulvate 0.8g/l, dipotassium hydrogen phosphate 1g/l, with a concentration of 3% dioxygen water sterilization 30 minutes after the above reagent dissolving；Sterilizing
It finishes and sodium bicarbonate 2g/l dissolvings is added, be fitted into 1L triangular flasks；Alternatively,
Large-scale culture weighs sodium acetate 8kg/t, 2 kg/t of ammonium chloride, 3 kg/t of yeast extract, 5 L/t of bittern, calcium chloride
0.5 kg/t, 0.8 kg/t of potassium fulvate, 1 kg/t of dipotassium hydrogen phosphate, with a concentration of 3% dioxygen water sterilization after the above reagent dissolving
30 minutes；Sterilizing, which finishes, is added sodium bicarbonate 2 kg/t dissolvings, is added in polybag；
Step 2: inoculated and cultured
It is inoculated with Rhodopseudomonas palustris bacterium solution by 20% inoculum concentration, 32 DEG C, 3000-5300lx incandescent lights are trained under the conditions of shining
72h-96h is supported, culture solution is diluted 100 times, OD values are measured at 660nm and reach 1.8 or more；Utensil goes out used in inoculation
Bacterium is handled, seeded process sterile working；
Step 3: thalline synchronization process
Culture solution is placed under dark condition, not contact nature light, by feux rouges:Blue light ratio is 4：1~1：1 arranges in pairs or groups
3~5h is irradiated in irradiation；Purpose is that Rhodopseudomonas palustris metabolic activity is made gradually to slow down, and form gradually reaches unanimity；
Step 4: physical concentration is handled
Polyacrylamide amine cation, sodium carboxymethylcellulose, Guar are added into the culture solution after above-mentioned synchronization process
The one or more of bean gum, xanthans, chitosan, ratio are 0.5%~1.5%, and the place of being protected from light preserves 3.5-5 hours, removes supernatant
Liquid, you can obtain hyperconcetration Rhodopseudomonas palustris, cell number is up to 7.3 × 1012A/g.
Embodiment 2
A kind of Rhodopseudomonas palustris synchronizes the application of concentration, takes the above-mentioned Rhodopseudomonas palustris after physical concentration
Protective agent polyethylene glycol 400, the one or more of trehalose, glycerine, sodium glutamate, Tween 80, addition is added in portion thereto
Amount is 8%~35%, is uniformly mixed to get Rhodopseudomonas palustris hyperconcetration active bacteria formulation；This product character is liquid, effective marsh
Red pseudomonas number is up to 7 × 1011 A/ml.
Embodiment 3
A kind of Rhodopseudomonas palustris synchronizes the application of concentration, takes the Rhodopseudomonas palustris one after above-mentioned physical concentration
Part, a or several part in silica, sodium sulphate, calcium oxide, maize cob meal and magnesium sulfate is added thereto, cold nebulization is dry
Yeast powder portion is added to three parts in dry processing, pulvis after drying；Rhodopseudomonas palustris is made and concentrates bacterium powder；According to said method make
Powder product in Rhodopseudomonas palustris cell number up to 5.0 × 109 A/g.
Embodiment 4
6 parts of culture of Penaeus vannamei later stage pond sludge, every part of 100g is taken to mix with weight such as pure water, surveyed after mixing
Fixed initial COD, ammonia nitrogen, dissolved oxygen, nitrite index（COD and dissolved oxygen index are measured using national standard method, ammonia nitrogen and Asia
Nitrate index is measured using assay kit）.Experiment divides experimental group and control group, every group of two parallel tests；Example 1
It concentrates Rhodopseudomonas palustris 1.0ml and dilutes 1000 times, solution is uniformly mixed after 1.0ml dilutions are added in experimental group one；Experimental group
Two, which are added 3 powder product of 1.0g embodiments, is uniformly mixed；The weight waters such as control group addition are uniformly mixed；It is above-mentioned to place 48h measurement
Indices, the experimental results are shown inthe following table：
From upper table data：Product has larger improvement using the measurement index numerical value of front and back selection：Compared to before using and
Control group data, the poisonous and harmful index of two experimental groups（COD, ammonia nitrogen, nitrite index）It significantly reduces, especially ammonia nitrogen
It is brought down below detection limit range by content is exceeded with nitrite index, the dissolved oxygen index of experimental group is increased significantly, reaches
Within normal range (NR).
Experiment shows：With obvious effects for degradation of ammonia nitrogen, COD, nitrite index using above-described embodiment product, this is right
There is remarkable result in the substrate and water quality for improving cultivation middle and later periods aging water body.High concentration Rhodopseudomonas palustris in this product
Palatability is good, can be used as biological feed early period in cultivation, provides albumen and B family vitamin；In the cultivation middle and later periods, can be used for regulating and controlling
Water quality, Rhodopseudomonas palustris of periodically splashing are conducive to the lower water column in cultivation and form dominant bacteria phase, effectively inhibit pathogenic bacteria.
Rhodopseudomonas palustris is synchronized can effectively to extend the shelf life with after concentration, reduce layering and thalline
Phenomena such as dead.The effective carrier added in powder product can accelerate the recovery of Rhodopseudomonas palustris, fast onset effect.
Claims (3)
1. a kind of method that Rhodopseudomonas palustris synchronizes concentration, it is characterised in that：Include the following steps：
Step 1: Rhodopseudomonas palustris culture
Triangular flask culture weighs sodium acetate 8g/l, ammonium chloride 2g/l, yeast extract 3g/l, bittern 5ml/l, calcium chloride 0.5g/l, yellow
Rotten acid potassium 0.8g/l, dipotassium hydrogen phosphate 1g/l, with a concentration of 3% dioxygen water sterilization 30 minutes after the above reagent dissolving；It has sterilized
Finish and sodium bicarbonate 2g/l dissolvings are added, is fitted into 1L triangular flasks；Alternatively,
Large-scale culture weighs sodium acetate 8kg/t, ammonium chloride 2kg/t, yeast extract 3kg/t, bittern 5L/t, calcium chloride 0.5kg/
T, potassium fulvate 0.8kg/t, dipotassium hydrogen phosphate 1kg/t, with a concentration of 3% dioxygen water sterilization 30 minutes after the above reagent dissolving；
Sterilizing, which finishes, is added sodium bicarbonate 2kg/t dissolvings, is added in polybag；
Step 2: inoculated and cultured
It is inoculated with Rhodopseudomonas palustris bacterium solution by 20% inoculum concentration, 32 DEG C, 3000-5300lx incandescent lights are cultivated under the conditions of shining
72h-96h, dilutes 100 times by culture solution, and OD values are measured at 660nm and reach 1.8 or more；Utensil sterilizes used in inoculation
Processing, seeded process sterile working；
Step 3: thalline synchronization process
Culture solution is placed under dark condition, not contact nature light, by feux rouges:Blue light ratio is 4：1~1：1 carries out collocation photograph
It penetrates, irradiates 3~5h；Purpose is that Rhodopseudomonas palustris metabolic activity is made gradually to slow down, and form gradually reaches unanimity；
Step 4: physical concentration is handled
Be added into the culture solution after above-mentioned synchronization process polyacrylamide amine cation, sodium carboxymethylcellulose, guar gum,
The one or more of xanthans, chitosan, ratio are 0.5%~1.5%, and the place of being protected from light preserves 3.5-5 hours, removes supernatant,
Hyperconcetration Rhodopseudomonas palustris can be obtained, cell number is up to 7.3 × 1012A/g.
2. a kind of Rhodopseudomonas palustris as described in claim 1 synchronizes the application of the method for concentration, it is characterised in that：It takes
The Rhodopseudomonas palustris after physical concentration described in claim 1 is a, and protective agent polyethylene glycol is added thereto
400, the one or more of trehalose, glycerine, sodium glutamate, Tween 80, additive amount are 8%~35%, are uniformly mixed to get natural pond
Damp red pseudomonas hyperconcetration active bacteria formulation；This product character is liquid, and effective Rhodopseudomonas palustris number is up to 7 × 1011A/
ml。
3. a kind of Rhodopseudomonas palustris as described in claim 1 synchronizes the application of the method for concentration, it is characterised in that：It takes
Rhodopseudomonas palustris after physical concentration described in claim 1 is a, be added thereto silica, sodium sulphate,
A or several part in calcium oxide, maize cob meal and magnesium sulfate, yeast powder one is added after drying in low temperature spray drying processing, pulvis
Part is to three parts；Rhodopseudomonas palustris is made and concentrates bacterium powder；Rhodopseudomonas palustris is thin in the powder product according to said method made
Born of the same parents' number is up to 5.0 × 109A/g.
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| CN109929783A (en) * | 2019-04-22 | 2019-06-25 | 广东宏隆生物科技有限公司 | The culture medium and cultural method of Rhodopseudomonas palustris |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101407774A (en) * | 2008-11-20 | 2009-04-15 | 中国科学院广州地球化学研究所 | Preparation technique of photosynthetic bacteria preparation |
| CN102212491A (en) * | 2011-03-29 | 2011-10-12 | 天津市农业生物技术研究中心 | Simple culture method of photosynthetic bacteria with high cell concentration |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101407774A (en) * | 2008-11-20 | 2009-04-15 | 中国科学院广州地球化学研究所 | Preparation technique of photosynthetic bacteria preparation |
| CN102212491A (en) * | 2011-03-29 | 2011-10-12 | 天津市农业生物技术研究中心 | Simple culture method of photosynthetic bacteria with high cell concentration |
Non-Patent Citations (1)
| Title |
|---|
| Synchronous Growth of Rhodopseudomonas palustris from the Swarmer Phase;D. WESTMACOTT等;《Journal of General Microbiology》;19761231;117-125 * |
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