CN103614286A - Method and device for cultivating high-concentration photosynthetic bacteria with low illumination - Google Patents
Method and device for cultivating high-concentration photosynthetic bacteria with low illumination Download PDFInfo
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Abstract
The invention discloses a method and a device for cultivating high-concentration photosynthetic bacteria with low illumination. The device comprises an anode chamber and a cathode chamber, wherein the anode chamber is made from a transparent material, and is separated from the cathode chamber through an ion exchange membrane; the anode chamber and the cathode chamber are respectively provided with an anode and a cathode; the anode and the cathode are connected through an external resistor; the ratio of surface area of the anode to effective capacity of the anode chamber is more than 0.06/cm, and ratio of resistance of the external resistor to effective capacity of the anode chamber is 4-40 ohm/cm3; the anode chamber of the device is a fermentation tank; photosynthetic bacteria cultivated to a logarithmic phase are inoculated to the anode chamber at a volume percentage of 1-10%, and the anode chamber is sealed and is communicated to an external circuit to cultivate the bacteria at 20-37 DEG C with illumination of 50-500lux. The method disclosed by the invention can cultivate high-concentration photosynthetic bacteria under low inoculum size and low illumination, and can generate electric energy while reducing illumination energy consumption; and the method is wide in application prospect.
Description
Technical field
The invention belongs to microorganism field, particularly adopt low-light (level) to cultivate the device of high density photosynthetic bacterium and utilize this device to cultivate the method for high density photosynthetic bacterium.
Background technology
Photosynthetic bacterium (being called for short PSB) is to occur the earliest on the earth, occurring in nature ubiquity, the prokaryotic organism with original luminous energy synthetic system, photosynthetic bacterium is energy decomposing organic matter and the ammonia-state nitrogen absorbing in water body in growth and breeding process, hydrogen sulfide, the objectionable impuritiess such as nitrite, in its cell dry-matter, protein content is up to more than 60%, its Amino Acid Composition of Proteins From Thermophlles is more complete, in cell, also contain multivitamin, especially vitamin B group is very abundant, Vb2, folic acid, pantothenic acid, the content of vitamin H is also higher, also contain a large amount of carotenoid simultaneously, the physiologically active substances such as ubiquinone.Therefore, photosynthetic bacterium can be widely used in the fields such as plant husbandry, livestock industry, culture fishery and environment protection, has huge utility value and potentiality to be exploited.
Photosynthetic bacterium has multiple nutrients metabolic way, can be aerobic in illumination, grow under the condition such as illumination anaerobism, dark aerobic, dark anaerobism and double oxygen.In actual photosynthetic bacterium produces, adopt the hold concurrently mode of oxygen of illumination anaerobism or illumination to cultivate more, because sulphate reducing bacteria or methanogen may form dominant growth while adopting dark fermentation culture photosynthetic bacterium, cause cultivating unsuccessfully; And the growth that illumination can directed strengthening photosynthetic bacterium while adopting light fermentation culture, meanwhile, light suppresses the growth of the miscellaneous bacterias such as methanogen or sulphate reducing bacteria to a certain extent, makes the photosynthetic bacterium in system more easily form dominant growth.But, photosynthetic bacterium requires high to intensity of illumination, to successfully realize the light fermentation culture of photosynthetic bacterium, conventionally need the illuminance of 2000～5000lux, this Design and manufacture to culture apparatus has proposed higher requirement: adopt the bioreactor of external light source to produce, be subject to the restriction of reactor wall material and nutrient solution light transmission, the light utilization efficiency of light source is low, reaction solution uneven illumination is even, and illumination is difficult to reach the requirement of photosynthetic bacterium growth; While adopting built-in light source, because photosynthetic bacterium has phototaxis, be easy to make photosynthetic bacterium to focus on light source surface and form microbial film, cause the interior local temperature of reactor too high, hinder the further transmission of light simultaneously.Owing to being subject to the restriction of all factors in illumination in form, the photosynthetic bacterium of inside reactor is difficult to obtain sufficient illumination, and this has limited large-scale production and the application of photosynthetic bacterium.
Publication number is that the patent that CN101407774A and publication number are CN102326673A discloses the method that photosynthetic bacterium is cultivated in dark fermentation (unglazed photograph).Although realized the method for dark fermentation culture photosynthetic bacterium, but culturing process needs constantly to fermentor tank, to pass into sterile air, and strictly control dissolved oxygen concentration between 1～2mg/L, this has increased aeration energy consumption, the complicacy of equipment and the difficulty of operational administrative have been increased simultaneously.Publication number is that the patent of CN103086520A discloses a kind of device and technology of processing livestock breeding wastewater producing biodiesel while, although this technology Neng turns out photosynthetic bacterium in anolyte compartment, but need to provide higher illuminance (2000～10000lux), and culture cycle reaches 13d.Publication number is that the patent of CN101409355B discloses a kind of photosynthesis microorganism fuel cell; this technology can efficiently be processed waste water and produce electric energy; but the photosynthetic bacterium concentration of gained is on the low side; be only 2.8 hundred million/mL; and anode and cathode damping fluid needs high-temperature sterilization, production and the application of its mass-producing have been limited.
Therefore, be badly in need of setting up method and the culture apparatus of a kind of low-light (level), low inoculum size cultivation high density photosynthetic bacterium, utilize the method under low irradiance, to cultivate the photosynthetic bacterium of high density, produce electric energy simultaneously, there is very important theory and realistic meaning.
Summary of the invention
In view of this, one of object of the present invention is to provide the device that adopts low irradiance to cultivate high density photosynthetic bacterium, and its result is simple; Two of object of the present invention is to provide the method for utilizing said apparatus to cultivate high density photosynthetic bacterium.
For achieving the above object, provide following technical scheme:
1. adopt low irradiance to cultivate the device of high density photosynthetic bacterium, comprise anolyte compartment and cathode compartment, described anolyte compartment is used transparent material, between described anolyte compartment and cathode compartment, with ion-exchange membrane, separate, described anolyte compartment and cathode compartment are respectively arranged with anode and negative electrode, described anode is connected by external resistance with negative electrode, and the surface-area of described anode is greater than 0.06/cm with the ratio of anolyte compartment's useful volume, and the resistance of described external resistance is 4～40 Ω/cm with the ratio of anolyte compartment's useful volume
3.The surface-area of anode is 0.27 with ratio the best of anolyte compartment's useful volume, and the resistance of external resistance is 4.17～8.3 Ω better effects if with the ratio of anolyte compartment's useful volume, and the best is 4.17 Ω or 8.3 Ω.
In the present invention, by controlling the resistance of external resistance and the ratio of anolyte compartment's useful volume, regulate external resistance to derive the excess electron of anolyte compartment's microorganisms, the oxidation reduction potential control of Yi Cijiang anolyte compartment is in double oxygen level, makes it to be suitable for amphimicrobian photosynthetic bacterium growth and suppresses the varied bacteria growings such as the methanogen of strictly anaerobic and sulphate reducing bacteria.
Preferably, described transparent material is synthetic glass, toughened glass, polyvinyl chloride or acrylic.
Preferably, described anode is made by carbon-based material, platinum-base material or stainless material.
Preferably, described cathode compartment is air cathode, biological-cathode or electrolytic solution negative electrode.
Preferably, described ion-exchange membrane is anion-exchange membrane or cationic exchange membrane.
2. utilize described device to cultivate the method for photosynthetic bacterium, comprise the steps:
By the photosynthetic bacterium that is cultured to logarithmic phase by volume per-cent be 1%～10% to be seeded to the anolyte compartment of described device, sealing anolyte compartment, after connection external circuit, in temperature, be 20～37 ℃, illuminance is to cultivate under 50～500Lux condition, and photosynthetic bacterium culture is equipped with in described anolyte compartment.Optimal light illumination is 100-300lux, optimum inoculation amount be by volume per-cent for be 5%～8% inoculation.
Preferably, described photosynthetic bacterium is Purple Non-sulfur photosynthetic bacterium or purple sulfur bacteria, and in Purple Non-sulfur photosynthetic bacterium or purple sulfur bacteria, rhodospirillum, Rhodopseudomonas or Rhodomicrobium effect are better.Optimum is Rhodopseudomonas palustris, the red bacterium of class ball or its mixture.
Preferred, the culture of photosynthetic bacterium is that photosynthetic bacteria culture medium or VFA concentration are not less than the organic waste water that 2g/L, ammonia nitrogen concentration are not less than 0.5g/L.
Preferably, described photosynthetic bacteria culture medium component is as follows: sodium acetate 3g/L, NH
4cl1g/L, MgSO
40.2g/L, K
2hPO
40.2g/L, NaHCO
31g/L, yeast powder 0.5g/L, and regulate pH to 7.4.
Utilize method of the present invention to cultivate photosynthetic bacterium, its inoculum size still less, substratum or organic waste water are added to anolyte compartment, anolyte compartment and air are isolated, form anaerobic environment, photosynthetic bacterium in anolyte compartment is with small molecules acid, alcohol is substrate, oxidation of organic compounds produces electronics and proton, they arrive cathode compartment by external circuit and ion-exchange membrane respectively, thereby automatically maintain the double oxygen environment that is conducive to photosynthetic bacterium growth in anolyte compartment, therefore without the aeration redox potential of the hierarchy of control automatically just, and can under low light conditions, cultivate the photosynthetic bacterium of high density, final concentration is 3,000,000,000/mL, in reduction energy consumption, produce electric energy simultaneously.
Principle of work of the present invention:
Microbiological fuel cell in the starting period (MFC) only produces lower electric current in biomass accumulation process, has higher redox potential.Through cultivating domestication, levels of current raises, redox potential declines gradually, when external circuit adopts the resistance of suitable resistance, can slow down the decline of redox potential, the oxygen level of holding concurrently that maintains that makes it the long period, is conducive to the growth of amphimicrobian Purple Non-sulfur photosynthetic bacterium or purple sulfur bacteria.Meanwhile, the oxygen that this moderate redox potential existing in anolyte compartment and cross-film infiltration are come can disturb the metabolism of obligate anaerobic type microorganism (for example methanogen and sulphate reducing bacteria), makes it in the downtrod state of growing.The growth that can strengthen Purple Non-sulfur photosynthetic bacterium or purple sulfur bacteria by this process suppresses the growth of other miscellaneous bacteria, thereby can realize low-light (level), low inoculum size is turned out the photosynthetic bacterium of high density.
Beneficial effect of the present invention is: the invention discloses the method for cultivating photosynthetic bacterium, utilize this method can under low-light (level), low inoculum size condition, realize the cultivation of high density photosynthetic bacterium, produce electric energy simultaneously, the method is easy and simple to handle, economical and efficient, compares tool have the following advantages with existing photosynthetic bacterium culture technique:
1, the present invention adopts low irradiance to realize the high-density culture of photosynthetic bacterium
In prior art for reach strengthening photosynthetic bacterium growth object, the illuminance that conventional photosynthetic bacteria used for light fermentation adopts is conventionally at 1000～5000lux, and the present invention adopts the illumination of 50～500lux can reach the photosynthetic bacterium concentration that conventional high illumination cultivation is produced, energy consumption significantly saves lighting, and reduce the requirement to culture apparatus daylighting and light transmission, save production cost.
2, the present invention adopts the totally-enclosed high-density culture that realizes photosynthetic bacterium without aeration mode
The present invention adopts totally enclosed anaerobism to cultivate, by electrode output electronics in anolyte compartment, automatically control the redox potential in storehouse, without to device aeration, do not need to arrange accurate online dissolved oxygen Controlling System, can stop aeration energy consumption and reduce the complexity of device and the difficulty of operational administrative, in addition, without the totally enclosed of aeration, cultivate the hidden danger that can effectively stop living contaminants in culturing process.
3, the present invention adopts low inoculum size to realize the high-density culture of photosynthetic bacterium
The fermentation of traditional light or the photosynthetic bacterium of secretly fermenting are cultivated needs higher inoculum size impel photosynthetic bacterium to form dominant growth, then utilizes the interspecific competition of photosynthetic bacterium to be used for suppressing the pollution of miscellaneous bacteria, and common initial inoculum need to reach 10%～30%.The present invention can effectively suppress the growth activity of the miscellaneous bacterias such as methanogen and sulphate reducing bacteria by set up electrode in culture apparatus, whole culturing process is difficult for by living contaminants, therefore the photosynthetic bacterium that adopts 1%～10% initial inoculum can produce certified concentration.
4. the present invention cultivates photosynthetic bacterium does not need high-temperature sterilization
The fermenting container that traditional fermentation process is used need to carry out autoclave sterilization processing, and utilize method of the present invention to cultivate photosynthetic bacterium, because the oxygen that can come by the redox potential that exists in anolyte compartment and cross-film infiltration can disturb obligate anaerobic type microorganism, thereby suppress the growth of miscellaneous bacteria, so fermenting container does not need to carry out sterilising treatment.
Accompanying drawing explanation
In order to make object of the present invention, technical scheme and beneficial effect clearer, the invention provides following accompanying drawing:
Fig. 1 be take photosynthetic bacterium number, voltage---the time curve of substratum during as substrate.
Fig. 2 be take the photosynthetic bacterium of alcohol waste water during as substrate and the percentage composition of miscellaneous bacteria.
Fig. 3 be take bacterial count---the time curve of alcohol waste water during as substrate.
Fig. 4 be take alcohol waste water redox potential---time curve in system during as substrate.
Embodiment
Below in conjunction with accompanying drawing, the preferred embodiments of the present invention are described in detail.The experimental technique of unreceipted actual conditions in embodiment, conventionally according to normal condition or the condition of advising according to manufacturer.
Adopt low irradiance to cultivate the device of high density photosynthetic bacterium, foster device comprises cathode compartment and anolyte compartment, wherein anolyte compartment is made by transparent material, two chamber interior specifications are 5cm * 5cm * 5cm, between two Room, by cationic exchange membrane, separate, between cationic exchange membrane and chamber, the two poles of the earth, accompany silicagel pad to keep sealing, every chamber packs 120mL solution into, in cathode compartment and anolyte compartment, be respectively arranged with negative electrode and anode, and connect by external resistance, external resistance resistance is set to 500 Ω, anode adopts carbon felt as electrode, specification is 4cm * 4cm, negative electrode adopts carbon-point as electrode.Anode chamber adds photosynthetic bacteria culture medium, and its composition is: sodium acetate 3g/L, NH
4cl1g/L, MgSO
40.2g/L, K
2hPO
40.2g/L, NaHCO
31g/L, yeast powder 0.5g/L, pH7.4.Cathode compartment adds negative electrode damping fluid, and its composition is: Tripotassium iron hexacyanide 16.45g/L, K
2hPO
413.6g/L.Then will the photosynthetic bacterium Rhodopseudomonas palustris that substratum anolyte compartment inoculation volume percent is 5% be housed, the concentration of Rhodopseudomonas palustris is about 3,000,000,000/mL(logarithmic phase), then at room temperature 20-37 ℃, illuminance, be to cultivate under 100lux condition, in culturing process, by volt ohm-milliammeter, regularly gather voltage, photosynthetic bacterium concentration detects and carries out according to photosynthetic bacterium microbial inoculum standard NY527-2002.
According to above-mentioned condition, carry out domestication and the enrichment of production with photosynthetic bacteria electro-biofilm, inoculate and cultivate photosynthetic bacterium about about one week, more than voltage reaches and is stabilized in 600mV, now by emptying fresh substratum and the Rhodopseudomonas palustris of rejoining of anolyte compartment's nutrient solution, when reaching 4,000,000,000/mL, photosynthetic bacterium concentration anolyte compartment's bacterium liquid can be discharged to results photosynthetic bacterial thallus, repeat to add substratum and photosynthetic bacterium can realize the fed batch cultivation of photosynthetic bacterium, continuous 4 thalline results and cultivation results are shown in Fig. 1.As seen from the figure, photosynthetic bacterium Rhodopseudomonas palustris substantially without lag period, is bred rapidly after inoculation in this culture system, and after inoculation, the concentration of 3d photosynthetic bacterium can reach 4,000,000,000/mL, and output voltage is basicly stable in 680mV left and right.
The present embodiment Anodic surface-area is 0.27/cm with the ratio of anolyte compartment's useful volume, and external resistance resistance is 4.17 Ω/cm with the ratio of anolyte compartment's useful volume
3.
In the present embodiment, transparent material can be used synthetic glass, toughened glass, polyvinyl chloride or acrylic, and anode can also be used the electrode of being made by carbon-based material, platinum-base material or stainless material except carbon brush.
Adopt the device of low irradiance cultivation high density photosynthetic bacterium identical with embodiment 1, difference is that cathode compartment is air cathode, separate by anion-exchange membrane chamber, the two poles of the earth, external resistance resistance is set to 1000 Ω， anolyte compartments and adds 120mL alcohol waste water, and its composition is: chemical oxygen demand (COD) 10g/L, general volatile organic acid 6g/L, ammonia nitrogen 0.5g/L, pH7.2, negative electrode adopts 0.5mg/cm
2carry platinum carbon paper as electrode, specification is 4cm * 4cm.The inoculation volume percent photosynthetic bacterium that is 8% is the red bacterium of class ball and Rhodopseudomonas palustris quantity than being the mixture of 1:1, bacterial concentration is about 3,000,000,000/mL(logarithmic phase), then under being 20-30 ℃, illuminance 300lux condition, temperature cultivates, in culturing process, voltage regularly gathers by volt ohm-milliammeter, total bacterial concentration is according to 660nm place absorbance detection, and photosynthetic bacterium concentration detects and carries out according to photosynthetic bacterium microbial inoculum standard NY527-2002.
Meanwhile, according to method same as described above, the identical air cathode MFC of two covers is set, carries out domestication and the enrichment of production with photosynthetic bacteria electro-biofilm, when domestication and enrichment culture, the external circuit of a set of MFC disconnects, as blank.Latter about 10 days of inoculation, more than voltage reaches and is stabilized in 700mV, now rejoin respectively fresh substratum and photosynthetic bacterium by two covering device inner anode chamber nutrient solutions are emptying, start formally to carry out photosynthetic bacterium cultivation, in culturing process, detect voltage and bacterial concentration, result as in Figure 2-4.
From Fig. 2 and Fig. 3, after inoculation, microorganism breeds rapidly, the bacterial count after inoculation 4d in two systems all can reach 2,500,000,000/more than mL.But in the MFC disconnecting at external circuit, have more serious living contaminants phenomenon, although total plate count has reached 2,700,000,000/mL, its photosynthetic bacterium content only has 25%; And in external resistance is the air cathode MFC of 1000 Ω, total bacteria has reached 5,400,000,000/mL, wherein the content of photosynthetic bacterium has reached 99%.As shown in Figure 4, in air cathode MFC, the electronics that bacterial oxidation substrate produces can guiding system outside, make redox potential in the system double oxygen condition (redox potential is between 200～-300mV) in suitable photosynthetic bacterium growth, and the growth of the anaerobism miscellaneous bacterias such as inhibition methanogen, sulphate reducing bacteria, thereby realize the directed effect of oxygen photosynthetic bacterium growth of holding concurrently of strengthening.Therefore, even become to be grouped under the condition of complicacy, photosynthetic bacterium inoculum size lower (8%) and low irradiance (300lux) at waste water, still can guarantee the dominant growth of photosynthetic bacterium, make the final concentration of photosynthetic bacterium reach 5,300,000,000/mL.
The present embodiment Anodic surface-area is 0.27/cm with the ratio of anolyte compartment's useful volume, and external resistance resistance is 8.3 Ω/cm with the ratio of anolyte compartment's useful volume
3.
Finally explanation is, above preferred embodiment is only unrestricted in order to technical scheme of the present invention to be described, although the present invention is described in detail by above preferred embodiment, but those skilled in the art are to be understood that, can to it, make various changes in the form and details, and not depart from the claims in the present invention book limited range.
Claims (10)
1. adopt low-light (level) to cultivate the device of high density photosynthetic bacterium, it is characterized in that: comprise anolyte compartment and cathode compartment, described anolyte compartment is used transparent material, between described anolyte compartment and cathode compartment, with ion-exchange membrane, separate, described anolyte compartment and cathode compartment are respectively arranged with anode and negative electrode, described anode is connected by external resistance with negative electrode, and the surface-area of described anode is greater than 0.06/cm with the ratio of anolyte compartment's useful volume, and the resistance of described external resistance is 4～40 Ω/cm with the ratio of anolyte compartment's useful volume
3.
2. device according to claim 1, is characterized in that: described transparent material is synthetic glass, toughened glass, polyvinyl chloride or acrylic.
3. device according to claim 1, is characterized in that: described anode is made by carbon-based material, platinum-base material or stainless material.
4. according to the device described in claim 1-3 any one, it is characterized in that: described cathode compartment is air cathode, biological-cathode or electrolytic solution negative electrode.
5. device according to claim 1, is characterized in that: described ion-exchange membrane is anion-exchange membrane or cationic exchange membrane.
6. utilize the method that device is cultivated high density photosynthetic bacterium described in claim 1-5 any one, it is characterized in that, comprise the steps:
By the photosynthetic bacterium that is cultured to logarithmic phase by volume per-cent be 1%～10% to be seeded to the anolyte compartment of described device, sealing anolyte compartment, after connection external circuit, in temperature, be 20～37 ℃, illuminance is to cultivate under 50～500lux condition, and photosynthetic bacterium culture is equipped with in described anolyte compartment.
7. method according to claim 6, is characterized in that: described illuminance is 100～300lux.
8. method according to claim 6, is characterized in that: described photosynthetic bacterium is Purple Non-sulfur photosynthetic bacterium or purple sulfur bacteria.
9. according to the method described in claim 6-8 any one, it is characterized in that: the culture of photosynthetic bacterium is that photosynthetic bacteria culture medium or VFA concentration are not less than the organic waste water that 2g/L, ammonia nitrogen concentration are not less than 0.5g/L.
10. method according to claim 9, is characterized in that: described photosynthetic bacteria culture medium component is as follows: sodium acetate 3g/L, NH
4cl1g/L, MgSO
40.2g/L, K
2hPO
40.2g/L, NaHCO
31g/L, yeast powder 0.5g/L, and regulate pH to 7.4.
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Cited By (3)
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| CN109250805A (en) * | 2018-09-05 | 2019-01-22 | 太原理工大学 | It is a kind of while handling high concentration starch water and optical microbial fuel cell and its application without carbon source containing nitrate wastewater |
| CN117886434A (en) * | 2024-02-22 | 2024-04-16 | 广东工业大学 | Recycling treatment device and treatment method for aged garbage leachate |
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| CN202333046U (en) * | 2011-09-05 | 2012-07-11 | 浙江工商大学 | Microbial fuel cell for denitrificating and desulphurating |
| CN103086520A (en) * | 2013-01-23 | 2013-05-08 | 中国科学院城市环境研究所 | Device and a method for producing biodiesel in a coupling way by treating livestock and poultry breeding wastewater |
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