CN112063661B - Method for producing medium-chain carboxylic acid from agricultural wastes in fermentation mode and application - Google Patents
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Abstract
The invention belongs to the field of agricultural waste utilization, and particularly relates to a method for producing medium-chain carboxylic acid by agricultural waste in a fermentation mode and application of the medium-chain carboxylic acid. In the method, the fermentation substrate is a mixture of livestock and poultry manure and agricultural wastes, and the agricultural wastes are crop straws and/or fruit and vegetable wastes; the method comprises the following steps: A. a primary fermentation stage: the zymocyte is lactobacillus, the fermentation temperature is 45-60 ℃, and the pH value is 4.0-6.0; B. a secondary fermentation stage: adopting domesticated sludge as inoculum for anaerobic fermentation at 25-40 deg.C and pH of 4.0-7.0. The invention obtains the medium-chain carboxylic acid with high added value from the livestock and poultry manure and the agricultural waste by a two-stage gradient temperature control fermentation method under the condition of not adding an external electron donor, thereby providing a feasible new way for high-value utilization of the agricultural waste.
Description
Technical Field
The invention belongs to the field of agricultural waste utilization, and particularly relates to a method for producing medium-chain carboxylic acid by agricultural waste in a fermentation mode and application of the medium-chain carboxylic acid.
Background
The organic wastes in agricultural production and living such as livestock and poultry manure, crop straws, fruit and vegetable wastes and the like have large resource quantity and high content of organic components, and are good biomass resources. The technology of producing biogas by anaerobic digestion is a common method for treating the waste, but the single biogas product has low efficiency and weak market competitiveness. Compared with biogas, the medium-chain carboxylic acid (caproic acid, enanthic acid, caprylic acid and the like) produced by the two-stage anaerobic fermentation process of biomass resources has wider application and market value, and the process mainly comprises 2 steps: 1) the first-stage fermentation organic waste is directionally converted into short-chain carboxylic acid; 2) the secondary fermentation synthesizes the short-chain carboxylic acid into the medium-chain carboxylic acid through a carbon chain extension reaction.
In recent years, research on the production of medium-chain carboxylic acid by the conversion of organic waste has attracted much attention, for example, hucho and the like use excess sludge as a raw material, and the anaerobic fermentation liquid of the hucho and the like performs a chain extension reaction to obtain hexanoic acid (hucho, research on the production of medium-chain fatty acid from excess sludge based on anaerobic fermentation, university of Hunan, 2018). It has also been shown that high-concentration caproic acid can be produced by a two-stage anaerobic fermentation process using food waste and municipal organic domestic waste as raw materials.
However, the prior art chain carboxylic Acid transformation processes all require the exogenous addition of ethanol as An electron donor, and Chen et al, through full Life Cycle assessment analysis, have shown that ethanol is the most important Environmental factor and source of cost for the synthesis process (Chen W S, Strik D P, Buisman C J, et al, Production of pharmaceutical Acid from Mixed Organic Water [ J ] Environmental Science & Technology, 2017, 51(12): 7159) 7168.). Therefore, reducing the amount of ethanol added or not adding an exogenous electron donor is the key to improve the process economy.
In contrast, chinese patent CN104357496B discloses a method for synthesizing hexanoic acid by catalyzing lactic acid with microorganisms, and indicates that lactic acid can effectively replace ethanol, drive carbon chain elongation reaction, and promote the synthesis of medium-chain carboxylic acid. Chinese patent CN 105420168B also discloses a rumen clostridium for synthesizing hexanoic acid by using lactic acid and an application method thereof, which realizes the synthesis of hexanoic acid without adding ethanol. However, they have not been investigated for the simultaneous preparation of a plurality of medium chain carboxylic acids without the addition of ethanol.
Generally, the products of the organic matter first-order anaerobic acidification process are mainly volatile short-chain carboxylic acids, and chinese patents CN 105132475A, CN 100526469C, CN 103923951A, CN 103509829A and CN 105603011 a disclose methods for producing volatile short-chain carboxylic acids such as acetic acid and butyric acid from wastes such as residual sludge and kitchen waste. Also, chinese patent CN 104911128B discloses a method for converting glucose/fructose and the like into lactic acid by using thermophilic anaerobic bacillus, but no report has been found on the research of directly using agricultural wastes such as livestock and poultry manure as raw materials and obtaining various medium-chain carboxylic acids by directional conversion without adding an external electron donor, and the idea of adding an external electron donor during the fermentation process of agricultural wastes such as poultry manure is still used, so that the production cost needs to be further reduced.
Disclosure of Invention
The invention provides a scheme capable of reducing the cost of producing the medium-chain carboxylic acid by oriented conversion of agricultural biomass resources, in order to overcome the defect that exogenous electron donors such as ethanol, lactic acid or methanol are required to be added in the process of producing the medium-chain carboxylic acid by two-stage fermentation of agricultural wastes such as livestock and poultry manure.
Specifically, the technical scheme of the invention is as follows:
a method for producing medium-chain carboxylic acid by agricultural wastes in a fermentation mode is characterized in that fermentation substrates are a mixture of livestock and poultry manure and agricultural wastes, and the agricultural wastes are crop straws and/or fruit and vegetable wastes; the method comprises the following steps:
A. a primary fermentation stage: the zymocyte is lactobacillus, the fermentation temperature is 45-60 ℃, and the pH value is 4.0-6.0;
B. a secondary fermentation stage: adopting domesticated sludge to carry out anaerobic fermentation, wherein the fermentation temperature is 25-40 ℃, and the pH value is 4.0-7.0.
The invention provides a scheme for realizing high-efficiency production of medium-chain carboxylic acid without adding an external electron donor by carrying out intensive research on the prior art for preparing the medium-chain carboxylic acid by taking livestock and poultry manure and other agricultural wastes as raw materials. Specifically, the lactic acid and various volatile fatty acids with specific proportional relation can be directionally generated in the primary fermentation stage by selecting different fermenting agents in each stage and matching with different control of stages of the fermentation process, so that the simultaneous and efficient production of various medium-chain carboxylic acids is realized in the subsequent secondary fermentation stage.
The volatile fatty acid is acetic acid, propionic acid, butyric acid and the like. The medium chain carboxylic acid is n-hexanoic acid, n-heptanoic acid, n-octanoic acid, etc.
Preferably, in the invention, the fermentation temperature in the primary fermentation stage is 50-55 ℃, and the pH value is 4.5-5.5, and the condition can improve the substrate competitive power of lactic acid producing bacteria, so as to be beneficial to enrichment and fermentation of dominant lactic acid bacteria and realize directional conversion of lactic acid and various volatile fatty acids in a specific proportion.
The fermentation temperature of the secondary fermentation stage is 25-35 ℃, the pH value is 5.0-6.5, so that the yield and the production efficiency of caproic acid are improved, and the propionic acid and the lactic acid are subjected to reverse beta oxidation to obtain heptanoic acid on the basis of ensuring the high-efficiency production of caproic acid and caprylic acid.
In the invention, the fermentation substrate is a mixture of livestock and poultry manure and crop straws, and the carbon-nitrogen ratio of the mixture is (30-40): 1;
or the fermentation substrate is a mixture of livestock and poultry manure and fruit and vegetable wastes, and the carbon-nitrogen ratio of the mixture is (30-40): 1.
The invention finds that when the proportion of the livestock and poultry manure and the agricultural wastes in the fermentation substrate is further controlled to be in the carbon-to-single ratio limited by the invention, the method can be helpful for obtaining lactic acid and various volatile fatty acids in a specific proportion, and further ensure the production of the final medium-chain carboxylic acid.
The crop straw is one or more of corn straw, wheat straw and rice straw; the fruit and vegetable waste is rotten fruits, seedlings, vines, leaves generated in the planting process of fruits or vegetables and tailings generated in the processing process; the total solid content of the crop straws is 30-70%, the volatile solid content is 85-95% TS (based on the total solid content), the crude fiber content is 35-55%, and the carbon-nitrogen ratio is (45-60): 1; the total solid content of the fruit and vegetable waste is 2-20%, the volatile solid content is 90-98% TS (based on the total solid content), the total sugar content is 10-20% (based on fresh weight), and the carbon-nitrogen ratio is (40-100): 1.
in the invention, the lactic acid bacteria comprise one or more of bifidobacterium, streptococcus thermophilus, lactobacillus plantarum, lactobacillus acidophilus, lactobacillus rhamnosus and lactobacillus bulgaricus.
In the step A of the invention, the fermentation concentration is 2% VS-15% VS (calculated by volatile solid); the inoculation amount of the bacterial liquid is 2-8% (V/V), and the number of viable bacteria in the bacterial liquid is 106-109 CFU/mL。
In the step B, the inoculation amount of the domesticated sludge is 5-20% (V/V), and preferably 10%. The preparation method comprises the following steps:
taking ethanol and/or lactic acid as an electron donor, and domesticating the granular sludge in an anaerobic fermentation tank for treating the organic wastewater under a fermentation system with the temperature of 25-40 ℃ and the pH value of 5.0-6.5.
The main microorganism species in the domesticated sludge is carbon chain extension bacteria. The domestication time is about 2 months, which aims to realize the purpose of enriching the carbon chain extension flora.
The main flora in the acclimated sludge is 10-25% of Clostridium kluyveri (Clostridium kluyveri), 20-30% of prevotella (prevotella ceae), 10-15% of vibrio succinivibrio (succinivibrio acid), 5-10% of bacteroides (bacteroides), 5-10% of treponema flexneri (spirochaete), 2-8% of rikenella (rikenella), 2-5% of bacillus (fibrobacter), 2-5% of lachnospira (lachnospira), 2-5% of ruminobacteria (ruminococcus) and 10-15% of the rest.
The invention also provides the application of the method in the production of medium-chain carboxylic acid.
In addition, in consideration of the fact that the short-chain carboxylic acid obtained in the primary fermentation stage has wide application value in the industry in the method for producing the medium-chain carboxylic acid by the two-stage gradient temperature-control mixed fermentation of the livestock and poultry manure and other agricultural wastes, the invention also provides a method for producing the short-chain carboxylic acid such as lactic acid and the like by utilizing the high-temperature mixed fermentation of the livestock and poultry manure and other agricultural wastes, which comprises the following steps:
Wherein the steps
The lactic acid fermentation bacteria mainly comprise bifidobacterium, streptococcus thermophilus, lactobacillus plantarum, lactobacillus acidophilus, lactobacillus rhamnosus, lactobacillus bulgaricus and the like.
Wherein the steps
The short-chain carboxylic acid such as lactic acid is produced in the fermentation stage, specifically, lactic acid producing bacteria liquid is inoculated according to the volume ratio of 2-8%, the fermentation temperature is controlled to be 45-60 ℃, and the pH value is controlled to be 4.0-6.0.
The invention has the beneficial effects that:
the invention provides a method for producing medium-chain carboxylic acid by utilizing two-stage gradient temperature-control mixed fermentation of livestock and poultry manure and other agricultural wastes, fermentation substrates are the livestock and poultry manure, crop straws, fruit and vegetable wastes and the like, the raw material sources are wide, no exogenous electron donor is required to be added in the medium-chain carboxylic acid conversion process, the operation cost is low, and the added value of the produced carboxylic acid product is high; can effectively realize the harmless treatment and high-value utilization of agricultural organic wastes such as livestock and poultry manure and the like. The method has the advantages of simple process operation, easy control of anaerobic fermentation conditions, low economic cost and good application prospect. The invention can convert more than 90 percent of endogenous short-chain carboxylic acid into medium-chain carboxylic acid with high added value, and provides an economic and feasible new way for high-value utilization of agricultural wastes.
Drawings
FIG. 1 is a bar graph showing the yield of carboxylic acid per unit VS of material under different fermentation temperatures in example 1 and comparative example 1 of the present invention.
FIG. 2 is a graph showing the tendency of conversion of lactic acid and medium chain carboxylic acid in example 2 of the present invention.
FIG. 3 is a graph showing the tendency of conversion of lactic acid and medium chain carboxylic acid in example 3 of the present invention.
Detailed Description
Preferred embodiments of the present invention will be described in detail with reference to the following examples. It is to be understood that the following examples are given for illustrative purposes only and are not intended to limit the scope of the present invention. Various modifications and alterations of this invention will become apparent to those skilled in the art without departing from the spirit and scope of this invention.
EXAMPLE 1 production of short-chain carboxylic acids such as lactic acid by high-temperature fermentation
The embodiment comprises the following steps:
1. and (3) culturing lactic acid bacteria: preparing a lactic acid bacteria culture medium-MRS broth (Qingdao high-tech industrial park Haibo biotechnology, Inc.), inoculating Baishengyou lactic acid bacteria powder (4 g/L, purchased from Shangen kang biotechnology, Inc.), controlling the culture temperature and pH to 35-38 ℃ and 6.0-7.0 respectively, and culturing for 36 h.
2. Culturing the cultured lactobacillus to obtain a bacterial solution (viable count is 10)7CFU/mL) is inoculated into an anaerobic fermentation tank which takes pig manure and corn straws as substrates, the carbon-nitrogen ratio of the mixed materials is 40:1, and the inoculation amount is 4%. The fermentation concentration is 4% VS, the pH is controlled at 4.5-5.5, the high-temperature anaerobic fermentation is carried out, and the fermentation temperature is controlled at 50-55 ℃. After 2 weeks of continuous anaerobic acidification fermentation, the carboxylic acid yield per unit VS of fermentation raw material is shown in FIG. 1, and the total carboxylic acid yield, lactic acid yield and volatile fatty acid yield of the material under high-temperature fermentation conditions are 226.8, 214.9 and 23.0 mg/gVS respectively, wherein the volatile fatty acid isYields of acetic acid, propionic acid, butyric acid were 6.1, 3.4 and 13.5 mg/gVS, respectively.
Comparative example 1 production of short-chain carboxylic acids such as lactic acid by Medium temperature fermentation
The production procedure of this comparative example is the same as example 1, except that: the fermentation temperature in step 2 is controlled to be 35-38 ℃. After 2 weeks of continuous anaerobic acidification fermentation, the carboxylic acid yield per unit VS of fermentation feedstock is shown in FIG. 1, and the total carboxylic acid yield, lactic acid yield and volatile fatty acid yield of the material under mesophilic fermentation conditions are 107.9, 81.9 and 25.8 mg/gVS, respectively. As can be seen from FIG. 1, the total carboxylic acid yield and lactic acid yield of the material under the high temperature condition (example 1) were 2.1 and 8.3 times higher than those under the medium temperature fermentation (comparative example 1), respectively. And the yield ratio of lactic acid to other volatile fatty acids also changed greatly.
Example 2 two-stage fermentation to produce Medium chain Carboxylic acids
The primary fermented acidified solution obtained after 2 weeks of continuous anaerobic acidification fermentation in example 1 was transferred to a secondary fermentor, and inoculated with acclimated sludge having a carbon chain elongation flora (Clostridium kluyveri 23.4%, prevotella 22.3%, vibrio succinivibrio succinate 11.6%, bacteroides 8.2%, leptospira Spirochaetaceae 6.7%, rikenella 5.5%, cellulose fibrillobacter 3.8%, lachnospira Lachnospiraceae 2.4%, ruminococcus 3.6%, other about 12.5%), which was prepared by: using ethanol and/or lactic acid as electron donor, acclimating granular sludge in anaerobic fermentation tank for treating organic wastewater for 2 months at 25-40 deg.C and pH of 5.0-6.5. The inoculation amount is 10 percent (volume ratio), polyurethane biological filler is filled in a fermentation tank for microorganism attachment, the fermentation temperature is 30 +/-1 ℃, and the pH value in the secondary fermentation process is controlled to be 6.0 +/-0.2. After 1 week of continuous fermentation, more than 95% of lactic acid is converted into medium-chain carboxylic acid such as caproic acid and the like (see figure 2) through carbon chain extension, the concentrations of caproic acid, enanthic acid and caprylic acid are 4039.5, 813.0 and 690.3 mgCOD/L respectively, and calculation can further obtain that 866.0 mgCOD medium-chain carboxylic acid can be correspondingly converted and synthesized after 1g of lactic acid is degraded.
Example 3
The procedure of this example is the same as example 2, except that: the pH value in the secondary fermentation process is controlled to be 4.5 +/-0.2. Compared with example 2, the conversion rate of lactic acid is slower, only 2 medium-chain carboxylic acids of caproic acid and caprylic acid are detected in the secondary fermentation product, and the maximum concentration is 2302.8 and 562.3 mgCOD/L (see figure 3), and the calculation can further show that every 1g of lactic acid is degraded, the corresponding conversion can be carried out to synthesize 409.3 mgCOD medium-chain carboxylic acid.
Example 4
The procedure for producing a short chain carboxylic acid of this example is the same as in example 1 except that: the substrate is a mixture of pig manure and fruit and vegetable waste (TS: 6%, VS: 92% TS, total sugar content 10% fresh weight, C/N =60: 1), and the carbon-nitrogen ratio of the mixture is 40: 1. The bacterial liquid has viable count of 106CFU/mL Streptococcus thermophilus (purchased from Shangkang Biotechnology, Suzhou) Limited) has a fermentation concentration of 15% VS at the primary fermentation stage and a bacterial liquid inoculation amount of 8% (V/V), and after 2 weeks of continuous anaerobic acidification fermentation, the total carboxylic acid yield, the lactic acid yield and the volatile fatty acid yield of fermentation raw material unit VS are respectively 370.0, 327.1 and 43.9 mg/gVS. Wherein the yields of acetic acid, propionic acid and butyric acid in the volatile fatty acids were 18.4, 9.6 and 15.9 mg/gVS, respectively.
In this example, the acidified solution of the first fermentation obtained above was transferred to a second fermentation tank, and medium-chain carboxylic acid was produced by the method of example 2, and after 1 week of continuous fermentation, the concentrations of hexanoic acid, heptanoic acid, and octanoic acid were 7732.9, 823.6, and 932.5 mgCOD/L, respectively, and the yield of medium-chain carboxylic acid was 791.8 mgCOD medium-chain carboxylic acid/g lactic acid.
Comparative example 2
This comparative example produced medium chain acids directly from the substrate using a one-step process, the specific steps being as follows:
the fermentation raw materials and the concentration are the same as those in example 1, the fermentation temperature is 35 +/-1 ℃, the pH value is 5.5 +/-0.5, the medium-chain carboxylic acid is obtained by the one-step method, and lactic acid, acetic acid, butyric acid and n-hexanoic acid can be detected in the fermentation substrate after continuous 3-week anaerobic fermentation, wherein the concentrations of the lactic acid, the acetic acid, the butyric acid and the n-hexanoic acid are 550.2, 1360.0, 2470.8 and 620.5 mgCOD/L respectively.
Comparative example 3
The production procedure of this comparative example is the same as example 2, except that: the secondary fermentation temperature is 45-48 ℃. After 1 week of continuous fermentation, only hexanoic acid and octanoic acid were detected at concentrations of 443.5 and 108.7 mgCOD/L, respectively, with a medium chain carboxylic acid yield of only 95.2 mgCOD medium chain carboxylic acid/g lactic acid.
Although the invention has been described in detail hereinabove with respect to a general description and specific embodiments thereof, it will be apparent to those skilled in the art that modifications or improvements may be made thereto based on the invention. Accordingly, such modifications and improvements are intended to be within the scope of the invention as claimed.
Claims (4)
1. A method for producing medium-chain carboxylic acid by agricultural wastes in a fermentation mode is characterized in that a fermentation substrate is a mixture of livestock and poultry manure and agricultural wastes, and the agricultural wastes are crop straws or fruit and vegetable wastes; the method comprises the following steps:
A. a primary fermentation stage: the zymocyte is lactobacillus, the fermentation temperature is 50-55 deg.C, and pH is 4.5-5.5;
B. a secondary fermentation stage: performing anaerobic fermentation by using domesticated sludge, wherein the fermentation temperature is 25-35 ℃, and the pH value is 5.0-6.5;
the medium chain carboxylic acids include n-hexanoic acid, n-heptanoic acid, and n-octanoic acid;
the fermentation substrate is a mixture of livestock and poultry manure and crop straws, and the carbon-nitrogen ratio of the mixture is (30-40) to 1;
or the fermentation substrate is a mixture of livestock and poultry manure and fruit and vegetable wastes, and the carbon-nitrogen ratio of the mixture is (30-40): 1.
2. The method of claim 1, wherein the lactic acid bacteria comprise one or more of bifidobacteria, streptococcus thermophilus, lactobacillus plantarum, lactobacillus acidophilus, lactobacillus rhamnosus, lactobacillus bulgaricus.
3. The method of claim 2, wherein in step a, the fermentation concentration is 2% VS-15% VS; the inoculation amount of the bacterial liquid is 2-8% (V/V), and the number of viable bacteria in the bacterial liquid is 106-109 CFU/mL。
4. The method according to claim 1, wherein in the step B, the inoculation amount of the acclimatized sludge is 5-20% (V/V), and the preparation method comprises the following steps:
taking ethanol and/or lactic acid as an electron donor, and domesticating the granular sludge in an anaerobic fermentation tank for treating the organic wastewater under a fermentation system with the temperature of 25-40 ℃ and the pH value of 5.0-6.5.
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| CN117737141B (en) * | 2024-02-19 | 2024-05-14 | 中国农业科学院农业环境与可持续发展研究所 | Method for regulating electron acceptor and electron donor ratio to promote production of medium-chain carboxylic acid |
| CN117737142B (en) * | 2024-02-20 | 2024-05-31 | 中国农业科学院农业环境与可持续发展研究所 | Method for producing medium-chain carboxylic acid by fermenting acidizing fluid with high-concentration agricultural source |
| CN118028387B (en) * | 2024-04-10 | 2024-06-25 | 中国农业科学院农业环境与可持续发展研究所 | Method for synthesizing polyhydroxyalkanoate by using low-pH lactic acid fermentation liquor through mixed flora |
| CN118064511B (en) * | 2024-04-18 | 2024-08-06 | 中国农业科学院农业环境与可持续发展研究所 | Storage method of agricultural waste and method for producing lactic acid by utilizing fermentation of agricultural waste |
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| EP3581659A1 (en) * | 2018-06-15 | 2019-12-18 | Politechnika Poznanska | Method for one-pot co-production of caproic acid and hydrogen |
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| CN107363076A (en) * | 2017-08-10 | 2017-11-21 | 中国科学院成都生物研究所 | A kind of reclaiming organic waste processing method |
| CN110643645A (en) * | 2019-10-31 | 2020-01-03 | 中国农业科学院农业环境与可持续发展研究所 | Method for producing medium-chain fatty acid by taking livestock and poultry manure as raw material through two-stage fermentation |
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