CN105755049A - Method of preparing hydrogen by using xylose as substrate for fermentation - Google Patents
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Abstract
The invention relates to a method of preparing hydrogen by using xylose as a substrate for fermentation.The method particularly includes: adopting an xylose solution as the substrate and anaerobic digestion sludge, Clostridium butyricum INET1 or Enterococcus faecium INET2 as hydrogen-generating strains, wherein the xylose solution is an xylose water solution or cellulose hydrolyzate; fermenting under anaerobic conditions to prepare hydrogen, wherein maximum hydrogen yield of xylose is 105-330 mL/100mL and hydrogen generating efficiency of each gram of xylose is 66-228 mL.By implementing the method, the problem that the cellulose hydrolyzate cannot be fully utilized due to the fact that xylose is difficult to be utilized by microorganisms is solved effectively.Xylose is a main product of plant cellulose, and plant fiber raw materials are rich in resource, low in price and easy to get; xylose which is an important hydrolyzate of plant cellulose is used as the substrate for biological hydrogen generation, so that raw material cost of biological fermentation for hydrogen generation can be lowered and full utilization of cellulose raw materials can be realized.
Description
Technical field
The invention belongs to fermentative hydrogen production technical field, particularly to a kind of method preparing hydrogen for fermenting substrate with xylose.
Background technology
Hydrogen Energy is secondary energy sources, and current process for making hydrogen mainly has water electrolysis hydrogen production, pyrolysis hydrogen manufacturing, photochemical hydrogen manufacturing, Estimation of Nuclear Hydrogen Production, fossil fuels hydrogen manufacturing and biological hydrogen production etc..Water electrolysis hydrogen production is applied relatively wide at present, and comparative maturity.The efficiency of water electrolysis hydrogen production is generally 75～85%, and its technical process is simple, pollution-free, but power consumption is big, and therefore its application receives certain restriction.Photochemistry hydrogen manufacturing is with water for raw material, the method for photocatalysis Decomposition hydrogen making.Fossil fuel hydrogen manufacturing is the topmost method of current hydrogen making, and it is with coal, oil, natural gas for raw material.Utilizing nuclear energy and decomposing water with solar energy hydrogen manufacturing is the problem that people make great efforts to explore.
Comparing above-mentioned process for making hydrogen, fermentative hydrogen production method has reaction condition gentleness, and consume energy low and feature of environmental protection high, prepares hydrogen by fermentation method, it is possible to farthest play the environmental benefit of hydrogen.
Substrate is the key factor of restriction biological hydrogen production development, only adopts cheap substrate just can reduce hydrogen manufacturing cost, promotes the application of this technology.China, as a large agricultural country, has arable land more than 1,500,000,000 mu, and every annual meeting produces substantial amounts of Plant fiber's garbage.According to incompletely statistics, straw is often only by China, the yield of cot one just can reach more than 700,000,000 ton.These organic major parts, all without being utilized effectively, are mainly disposed in the way of on-site incineration, are not only wasted resource, and environment is caused pollution.If fermentation and hydrogen production can be used it for, then cellulosic is just obtained in that and utilizes more fully.
Lignocellulose is as the key component of agricultural crop straw, after physics, Chemical Pretreatment or employing enzymolysis processing, the crude fibre (lignin and cellulose) of 40% and the hemicellulose of 20% can be obtained, hemicellulose and cellulose are hydrolyzed to monosaccharide molecule under the effect of enzyme or acid, it is mainly glucose and xylose, wherein glucose can be utilized by almost all of microorganism, and account for the pentose of 30～40%, namely xylose is but difficult to be utilized by microorganism, there is also the compound of substantial amounts of possible microbiostatic activity in cellulosic hydrolysate simultaneously.The utilization of xylose in cellulosic hydrolysate, is emphasis and the focus of research at present.The cellulosic hydrolysate that utilizes of report produces research paper and the patent of hydrogen at present, its essence is and utilizes the glucose in hydrolyzed solution as fermentation substrate.Such as, studies have found that, xylose in the available cellulosic hydrolysate of some is thermophilic H_2-producing strains grows, but do not report utilize wherein xylose product hydrogen usefulness (Zhu Yuhong. hot fermentation H_2-producing strains W16 and fermentation and hydrogen production characteristic [D] thereof. Harbin Institute of Technology, 2008；Liu Yan. thermophilic anaerobic clostridium degraded cellulose and hydrogen-producing characteristic research [D]. Shandong University, 2008.)；The research of Niu Kun then finds, xylose in cellulosic hydrolysate can suppress by affecting the metabolic pathway of microorganism its produce hydrogen effect (the dark fermentation and hydrogen production process analysis of cattle female .KlebsiellapneumoniaeECU-15 bacterial strain and utilize the experimentation [D] of ligno-cellulose hydrolysate. East China University of Science, 2010.).Therefore, that cellulosic hydrolysate efficiently utilizes it is used for producing hydrogen as energy source by the xylose in cellulosic hydrolysate it is crucial that utilize xylose therein, the resource efficiency of cellulose substances can be substantially improved undoubtedly.
Summary of the invention
Not enough for prior art, the invention provides a kind of method preparing hydrogen for fermenting substrate with xylose.
A kind of method preparing hydrogen for fermenting substrate with xylose, comprises the following steps:
1) preculture of hydrogenogen kind；
2) with xylose solution for substrate, add nutritional solution and prepare fermentation and hydrogen production culture medium, by step 1) in pre-incubated hydrogenogen kind be inoculated in fermentation and hydrogen production culture medium, nitrogen more than stripping 3min is to provide oxygen-free environment, and reactor is placed in water bath with thermostatic control shaking table, and temperature controls at 25～40 DEG C, shaking speed is 80～120r/min, regulate initial pH=6.0～9.0 of fermentation liquid, carry out ferment for hydrogen production, collect fermentation gas and remove CO2Obtain H2。
Step 1) described in hydrogenogen kind be the anaerobically digested sludge of irradiated pretreatment, or the mixed bacterial of Clostridium butyricum (Clostridiumbutyricum) INET1 and enterococcus faecalis (EnterococcusFaecium) INET2, or single culture Clostridium butyricum INET1, or single culture enterococcus faecalis INET2；The described Clostridium butyricum INET1 deposit number at China Committee for Culture Collection of Microorganisms's common micro-organisms center is CGMCC1.5199, and preservation date is on June 3rd, 2015；The described enterococcus faecalis INET2 deposit number at China Committee for Culture Collection of Microorganisms's common micro-organisms center is CGMCC1.15321, and preservation date is on June 3rd, 2015.The address at China Committee for Culture Collection of Microorganisms's common micro-organisms center is Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3.
The gamma-ray irradiation that described irradiation pretreatment is 5kGy dosage processes, and adopts60Co source is as radioactive source.
Described Clostridium butyricum INET1 and enterococcus faecalis INET2 separates in the anaerobically digested sludge processed by the gamma-ray irradiation through 5kGy dosage to obtain, and radioactive source is60Co。
The preculture of the anaerobically digested sludge of irradiated pretreatment, nutrient media components is: xylose 20g/L, peptone 10g/L, yeast powder 0.5g/L, nutritional solution 10mL/100mL；
Clostridium butyricum INET1, enterococcus faecalis INET2 preculture, nutrient media components is: glucose 50g/L, peptone 10g/L, yeast powder 0.5g/L, nutritional solution 10mL/100mL.
Step 2) described in xylose solution be xylose aqueous solution or cellulosic hydrolysate, described xylose solution is cellulosic hydrolysate is use cellulosic hydrolysate directly prepare fermentation and hydrogen production culture medium.
The component of described fermentation and hydrogen production culture medium is: xylose solution 90mL/100mL, nutritional solution 10mL/100mL.
In fermentation and hydrogen production culture medium, the inoculative proportion of hydrogenogen kind is 10mL/100mL.
The component of described nutritional solution is: NaHCO340g/L, NH4Cl5g/L, NaH2PO4·2H2O5g/L, K2HPO4·3H2O5g/L, FeSO4·7H2O0.25g/L, MgCl2·6H2O0.085g/L, NiCl2·6H2O0.004g/L。
In described fermentation and hydrogen production culture medium, xylose concentration is 5～50g/L, and the maximum hydrogen output of xylose is 105～330mL/100mL, and hydrogen generation efficiency is 66～228mL/g xylose.
The invention have the benefit that ligno-cellulose hydrolysate is that substrate carries out producing hydrogen, hydrogenogen kind of the present invention, be possible not only to make full use of glucose therein, it is also possible to effectively utilize xylose therein to carry out producing hydrogen.Xylose is as the important hydrolyzate of lignocellulose, hydrogen is prepared for fermenting substrate with xylose, not only extend the substrate spectrum of biological hydrogen production, improve the utilization rate of plant fiber material, reduce the cost producing hydrogen substrate, also contribute to realize the minimizing of Plant fiber's garbage, resource, be really achieved the purpose turned waste into wealth.The successful Application of the present invention is significant to the industrialization realizing biological hydrogen production, will push directly on making full use of further of cellulose substances, disposes providing new method for plant waste.
Detailed description of the invention
Below in conjunction with detailed description of the invention, the present invention will be further described.It is emphasized that the description below is merely exemplary, rather than in order to limit the scope of the present invention and application thereof.
Embodiment 1
With xylose for substrate, the anaerobically digested sludge inoculating irradiated pretreatment carries out fermentation and hydrogen production:
Anaerobically digested sludge adopts the gamma-rays of 5kGy dosage to carry out irradiation pretreatment, with60Co is as radioactive source.The anaerobically digested sludge of irradiation pretreatment carries out enrichment preculture, and pre-incubated nutrient media components is: xylose 20g/L, peptone 10g/L, yeast powder 0.5g/L, nutritional solution 10mL/100mL.After preculture, take 10mL and be inoculated in 100mL fermentation and hydrogen production culture medium, fermentation and hydrogen production nutrient media components is: xylose 50g/L, nutritional solution 10mL/100mL, using the HCl solution of 5mol/L or the NaOH solution of 5mol/L to regulate initial pH is 7.0, and nitrogen stripping inspection air-tightness also drives away oxygen in bottle, reaction bulb is placed in water bath with thermostatic control shaking table, at 35 DEG C, 100r/min reacts bottom fermentation and produces hydrogen, and interval 2h records gas production.Producing H-H reaction after 48h to terminate, gas chromatographic analysis finds in course of reaction without methane generation, it is seen that radiation treatment has killed methane backeria effectively.The largest cumulative hydrogen output that experiment obtains is 330mL/100mL, and hydrogen generation efficiency is 66mL/g xylose, and maximum hydrogen-producing speed is 20.3mL 100mL-1·h-1, lag time is 2.4h.
Embodiment 2
With xylose for substrate, the anaerobically digested sludge inoculating irradiated pretreatment carries out fermentation and hydrogen production:
Anaerobically digested sludge adopts the gamma-rays of 5kGy dosage to carry out irradiation pretreatment, with60Co is as radioactive source.The anaerobically digested sludge of irradiation pretreatment carries out enrichment preculture, and pre-incubated nutrient media components is: xylose 20g/L, peptone 10g/L, yeast powder 0.5g/L, nutritional solution 10mL/100mL.After preculture, take 10mL and be inoculated in 100mL fermentation and hydrogen production culture medium, fermentation and hydrogen production nutrient media components is: xylose 5g/L, nutritional solution 10mL/100mL, using the HCl solution of 5mol/L or the NaOH solution of 5mol/L to regulate initial pH is 7.0, and nitrogen stripping inspection air-tightness also drives away oxygen in bottle, reaction bulb is placed in water bath with thermostatic control shaking table, at 35 DEG C, 100r/min reacts bottom fermentation and produces hydrogen, and interval 2h records gas production.Producing H-H reaction after 48h to terminate, gas chromatographic analysis finds in course of reaction without methane generation, it is seen that radiation treatment has killed methane backeria effectively.The largest cumulative hydrogen output that experiment obtains is 114mL/100mL, and hydrogen generation efficiency is 228mL/g xylose, and maximum hydrogen-producing speed is 7.8mL 100mL-1·h-1, lag time is 1.8h.
Embodiment 3
With xylose for substrate, inoculation Clostridium butyricum INET1 carries out fermentation and hydrogen production:
Picking Clostridium butyricum INET1 bacterium colony from plating medium, is inoculated in 200mL enrichment culture liquid and carries out enrichment preculture, and enrichment culture liquid composition is as follows: glucose 50g/L, peptone 10g/L, yeast powder 0.5g/L, nutritional solution 10mL/100mL.Strain grows into logarithmic (log) phase through 8-10h, now, taking 10mL to be in the bacterium solution of logarithmic (log) phase and be inoculated in 100mL fermentation and hydrogen production culture medium, fermentation and hydrogen production nutrient media components is: xylose 10g/L, nutritional solution 10mL/100mL, using the HCl solution of 5mol/L or the NaOH solution of 5mol/L to regulate initial pH is 7.0, utilize nitrogen stripping inspection air-tightness and drive away oxygen in bottle, reaction bulb being placed in water bath with thermostatic control shaking table, at 35 DEG C, 100r/min condition bottom fermentation produces hydrogen, and interval 2h records gas production.Producing H-H reaction after 56h to terminate, the largest cumulative hydrogen output that experiment obtains is 105mL/100mL, and hydrogen generation efficiency is 105mL/g xylose, and maximum gas production rate is 9.3mL 100mL-1·h-1, lag time is 36.4h.
Embodiment 4
With xylose for substrate, inoculation enterococcus faecalis INET2 carries out fermentation and hydrogen production:
Picking dung clostridium perfringens enterotoxin INET2 bacterium colony from plating medium, is inoculated in 200mL enrichment culture liquid and carries out enrichment preculture, and enrichment culture liquid composition is as follows: glucose 50g/L, peptone 10g/L, yeast powder 0.5g/L, nutritional solution 10mL/100mL.Strain grows into logarithmic (log) phase through 8-10h, now, taking 10mL to be in the bacterium solution of logarithmic (log) phase and be inoculated in 100mL fermentation and hydrogen production culture medium, fermentation and hydrogen production nutrient media components is: xylose 10g/L, nutritional solution 10mL/100mL, using the HCl solution of 5mol/L or the NaOH solution of 5mol/L to regulate initial pH is 7.0, utilize nitrogen stripping inspection air-tightness and drive away oxygen in bottle, reaction bulb being placed in water bath with thermostatic control shaking table, with 35 DEG C, 100r/min conditioned response, interval 2h records gas production.Producing H-H reaction after 70h to terminate, the largest cumulative hydrogen output that obtains of experiment is 3mL/100mL, illustrates that single culture enterococcus faecalis INET2 carries out the ability of xylose fermentation for producing hydrogen weak.
Embodiment 5
With xylose for substrate, inoculation Clostridium butyricum INET1 and enterococcus faecalis INET2 carries out fermentation and hydrogen production simultaneously:
Picking Clostridium butyricum INET1 and enterococcus faecalis INET2 bacterium colony from plating medium, being inoculated in 200mL enrichment culture liquid respectively and carry out enrichment preculture, enrichment culture liquid composition is as follows: glucose 50g/L, peptone 10g/L, yeast powder 0.5g/L, nutritional solution 10mL/100mL.Two strain strains all grow into logarithmic (log) phase through 8-10h, now, take 5mL respectively to be in two kinds of bacterium solution of logarithmic (log) phase and be inoculated in 100mL wood-sugar fermentation culture medium, fermentation and hydrogen production nutrient media components is: xylose 10g/L, nutritional solution 10mL/100mL, using the HCl solution of 5mol/L or the NaOH solution of 5mol/L to regulate initial pH is 7.0, utilize nitrogen stripping inspection air-tightness and drive away in bottle after oxygen, reaction bulb is placed in water bath with thermostatic control shaking table, with 35 DEG C, 100r/min conditioned response, interval 2h records gas production.Producing H-H reaction after 64h to terminate, the largest cumulative hydrogen output that experiment obtains is 114mL/100mL, and hydrogen generation efficiency is 114mL/g xylose, and maximum hydrogen-producing speed is 10.8mL 100mL-1·h-1, lag time is 49.7h.
Embodiment 6
With ligno-cellulose hydrolysate for substrate, the anaerobically digested sludge inoculating irradiated pretreatment carries out fermentation and hydrogen production:
Anaerobically digested sludge adopts the gamma-rays of 5kGy dosage to carry out irradiation pretreatment, with60Co is as radioactive source.The anaerobically digested sludge of irradiation pretreatment carries out enrichment preculture, and pre-incubated nutrient media components is: xylose 20g/L, peptone 10g/L, yeast powder 0.5g/L, nutritional solution 10mL/100mL.After preculture, take 10mL and be inoculated in 100mL fermentation and hydrogen production culture medium, fermentation and hydrogen production nutrient media components is: ligno-cellulose hydrolysate 90mL/100mL, nutritional solution 10mL/100mL, in ligno-cellulose hydrolysate, xylose concentration is 18g/L, utilize xylose therein to produce hydrogen for substrate utilization xylose therein for fermenting substrate, and utilize the fermentation and hydrogen productions such as glucose therein simultaneously.Using the HCl solution of 5mol/L or the NaOH solution of 5mol/L to regulate initial pH is 7.0, utilize nitrogen stripping inspection air-tightness and drive away oxygen in bottle, reaction bulb being placed in water bath with thermostatic control shaking table, at 35 DEG C, 100r/min reacts bottom fermentation and produces hydrogen, and interval 2h records gas production.Producing H-H reaction after 60h to terminate, the utilization rate of glucose is reached 90%, the hydrogen that deduction glucose etc. produces, in experiment, the largest cumulative hydrogen output of xylose is 152mL/100mL, and hydrogen generation efficiency is 94mL/g xylose.
Claims (10)
1. the method preparing hydrogen for fermenting substrate with xylose, it is characterised in that comprise the following steps:
1) preculture of hydrogenogen kind；
2) with xylose solution for substrate, add nutritional solution and prepare fermentation and hydrogen production culture medium, by step 1) in pre-incubated hydrogenogen kind be inoculated in fermentation and hydrogen production culture medium, nitrogen more than stripping 3min is to provide oxygen-free environment, and reactor is placed in water bath with thermostatic control shaking table, and temperature controls at 25～40 DEG C, shaking speed is 80～120r/min, regulate initial pH=6.0～9.0 of fermentation liquid, carry out ferment for hydrogen production, collect fermentation gas and remove CO2Obtain H2。
2. method according to claim 1, it is characterized in that, step 1) described in hydrogenogen kind be the anaerobically digested sludge of irradiated pretreatment, or the mixed bacterial of Clostridium butyricum (Clostridiumbutyricum) INET1 and enterococcus faecalis (EnterococcusFaecium) INET2, or single culture Clostridium butyricum INET1, or single culture enterococcus faecalis INET2；The described Clostridium butyricum INET1 deposit number at China Committee for Culture Collection of Microorganisms's common micro-organisms center is CGMCC1.5199, and the described enterococcus faecalis INET2 deposit number at China Committee for Culture Collection of Microorganisms's common micro-organisms center is CGMCC1.15321.
3. method according to claim 2, it is characterised in that the gamma-ray irradiation that described irradiation pretreatment is 5kGy dosage processes, and adopts60Co source is as radioactive source.
4. method according to claim 2, it is characterised in that described Clostridium butyricum INET1 and enterococcus faecalis INET2 separates in the anaerobically digested sludge processed by the gamma-ray irradiation through 5kGy dosage to obtain, and radioactive source is60Co。
5. method according to claim 2, it is characterised in that the preculture of the anaerobically digested sludge of irradiated pretreatment, nutrient media components is: xylose 20g/L, peptone 10g/L, yeast powder 0.5g/L, nutritional solution 10mL/100mL；
Clostridium butyricum INET1, enterococcus faecalis INET2 preculture, nutrient media components is: glucose 50g/L, peptone 10g/L, yeast powder 0.5g/L, nutritional solution 10mL/100mL.
6. method according to claim 1, it is characterised in that step 2) described in xylose solution be xylose aqueous solution or cellulosic hydrolysate, described xylose solution is cellulosic hydrolysate is use cellulosic hydrolysate directly prepare fermentation and hydrogen production culture medium.
7. method according to claim 1, it is characterised in that the component of described fermentation and hydrogen production culture medium is: xylose solution 90mL/100mL, nutritional solution 10mL/100mL.
8. the method according to claim 5 or 7, it is characterised in that: the component of described nutritional solution is: NaHCO340g/L, NH4Cl5g/L, NaH2PO4·2H2O5g/L, K2HPO4·3H2O5g/L, FeSO4·7H2O0.25g/L, MgCl2·6H2O0.085g/L, NiCl2·6H2O0.004g/L。
9. method according to claim 1, it is characterised in that in described fermentation and hydrogen production culture medium, the inoculative proportion of hydrogenogen kind is 10mL/100mL.
10. method according to claim 1, it is characterised in that in described fermentation and hydrogen production culture medium, xylose concentration is 5～50g/L, the maximum hydrogen output of xylose is 105～330mL/100mL, and hydrogen generation efficiency is 66～228mL/g xylose.
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| CN113583921A (en) * | 2021-09-18 | 2021-11-02 | 北京北方华科科技有限公司 | Hydrogen producing bacterium and method for producing hydrogen by using same |
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| CN109825459A (en) * | 2019-03-29 | 2019-05-31 | 天津科技大学 | One plant of coupling produces the dissimilatory iron reduction bacterium of hydrogen |
| CN109825459B (en) * | 2019-03-29 | 2021-09-14 | 天津科技大学 | Dissimilatory iron reduction bacterium for coupling hydrogen production |
| CN113583921A (en) * | 2021-09-18 | 2021-11-02 | 北京北方华科科技有限公司 | Hydrogen producing bacterium and method for producing hydrogen by using same |
| CN117229936A (en) * | 2023-04-26 | 2023-12-15 | 淮阴师范学院 | Construction method and application of clostridium butyricum/CdS hybridization system |
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