CN102776247B - Method for increasing content of propionic acid in batch fermentation acid-producing liquid - Google Patents
Method for increasing content of propionic acid in batch fermentation acid-producing liquid Download PDFInfo
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- CN102776247B CN102776247B CN201210249898.7A CN201210249898A CN102776247B CN 102776247 B CN102776247 B CN 102776247B CN 201210249898 A CN201210249898 A CN 201210249898A CN 102776247 B CN102776247 B CN 102776247B
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Abstract
The invention belongs to the technical field of environmental protection and relates to a method for increasing content of propionic acid in batch fermentation acid-producing liquid. The method includes the steps of firstly, mixing kitchen waste and thickened sludge, adding mixture into an anaerobic hydrolysis fermenter for anaerobic fermentation, centrifugally separating mixture, and taking supernate for standby; secondly, activating and enriching propionibacterium seed broth; thirdly, sterilizing the supernate obtained in the step 1, inoculating the propionibacterium seed broth obtained in the step 2 to the supernate, and fermenting in an anaerobic fermenter to generate propionic acid; and fourthly, centrifuging the fermented mixture to obtain the fermented supernate rich in propionic acid after fermentation. The sludge and the kitchen waste are subjected to primary fermentation under the weak alkaline condition, and microorganism extracellular enzyme hydrolysis of macromolecular organic matters such as microorganism exoenzyme hydrolysate, protein and fat is facilitated. The content of the propionic acid in the fermented supernate is 3.56-9.95g COD per L of total acid in the supernate.
Description
Technical field
The invention belongs to environmental protection technical field, relate to a kind of method improving propionic acid content in batch fermentation product acid solution.
Background technology
Current, body eutrophication is day by day serious, and the nitrogen in control sewage and the amount of phosphorus are the Critical policies preventing body eutrophication in the world.Applying biological dephosphorization denitrogenation technology effectively can reduce the content of phosphorus in sewage and nitrogen, and the voltaile fatty acid in waste water can be biological dephosphorize denitrification provides required carbon source, there is report (see Water Science and Technology, 1992,25,185-194) claim Biochemical method 1mg phosphorus needs 6 ~ 9mg short chain fatty acid (SCFAs).
But in the water inlet of China's many Shelter in South China Cities Sewage Plant, solvability COD(comprises SCFAs, particularly acetic acid and propionic acid) containing quantity not sufficient, be difficult to the needs meeting high-performance bio dephosphorization denitrogenation.For this reason, some investigators have studied and in batch reactor, the mud (comprising primary sludge and excess sludge) that Sewage Plant produces carried out to anaerobically fermenting and prepares the research of short chain fatty acid (see Chinese invention patent 200510110451.1; Environmental Science and Technology, 2006,40:2025-2029).
According to another bibliographical information (see Water Research, 2004,38:27-36; Bioresource Technology, 2008,99:4400-4407), under the prerequisite keeping sewage total COD concentration constant, the propionic acid/acetic acid increasing wastewater influent than from, the removal effect of nitrogen and phosphorus can be significantly improved.This shows, the propionic acid content increasing sewage is more conducive to the raising of biological removal of phosphorus in wastewater denitrification effect than the concentration increasing acetic acid.
Having been found that in research process in the past by adding carbohydrate or rubbish from cooking in sludge fermentation product acid system, the propionic acid content (Chinese invention patent 200810035585.5 in mud product acid solution can be improved; Environmental Science andTechnology, 2009,43 (12): 4373-4380).But the content of propionic acid is lower, only less than 50%.
Summary of the invention
The object of the invention is to the defect for overcoming prior art and a kind of method improving propionic acid content in batch fermentation product acid solution is provided.
For achieving the above object, the present invention is by the following technical solutions:
Improve the method that batch fermentation produces propionic acid content in acid solution, comprise following steps:
(1) mixture (first stage ferments) of pre fermentation thickened sludge and rubbish from cooking;
By rubbish from cooking, thickened sludge mixing, join anaerobically fermenting in anaerobic hydrolysis fermentor tank, then by mixture centrifugation, take out supernatant liquor stand-by;
(2) also enrichment propionibacterium seed liquor is activated;
(3) propionibacterium seed liquor fermented supernatant fluid is utilized to produce propionic acid (subordinate phase fermentation);
By the supernatant liquor sterilizing that step (1) obtains, in the propionibacterium seed liquor that access step (2) obtains, in anaerobic fermentation tank, propionic acid is produced in fermentation;
(4) after fermentation ends, fermenting mixture is centrifugal, obtain the fermented supernatant fluid being rich in propionic acid.
In described step (1), the water ratio of thickened sludge is 98%-99%, and total suspended solid concentration (TS) is 10-20g/L (VS/TS=0.71 ± 0.08), be preferably 15g/L(wherein VS be organic substance, TS is total suspended matter matter, and VS/TS is the organic content characterizing mixture).
Described rubbish from cooking, for after the refuses such as rejecting chopsticks, broken bone, plastics, the scraps of paper, fragment, through pulverizing, crossing 10 mesh sieves, obtains the concentrated rubbish from cooking that C/N is 16-20; Adding tap water makes water ratio be 80%; Rubbish from cooking main ingredient is rice, and meat, oils are auxiliary, greengrocery less (can ignore), and wherein, the dry weight of rice, meat, oils is than being (3 ~ 10): (0.5 ~ 5): (0 ~ 2); Preferred rice, the dry weight of meat, oils is than being 7:2:1.
In described step (1), thickened sludge and rubbish from cooking are by dry weight ratio (0.3 ~ 0.08): 1 mixes, the total chemical oxygen demand (COD) (TCOD) added after tap water dilution is 20-40g COD/L, and preferred thickened sludge and rubbish from cooking are 0.18:1 by dry weight ratio.
In described step (1), the temperature of anaerobically fermenting is 10 ~ 65 DEG C, and stirring velocity is 60-250rpm, with Ca (OH)
2the pH controlling fermentation is 5-12, and fermentation time is 2-5 days; Preferred leavening temperature is 25 DEG C, and the pH of fermentation is 7, and fermentation time is 4 days.
In described step (1), the rotating speed of centrifugation is 2000-4000rpm, and centrifugation time is 4-6min.
In described step (2), propionibacterium (SICC1.256) is activated rear enrichment from inclined-plane and is stored in the glycerine pipe storage substratum of 10%-30% (culture medium prescription is in table 1), Conservation environment is-50 to-80 DEG C; Be seeded to enrichment medium (culture medium prescription is in table 2) according to inoculum size 5-15% at every turn, in culturing bottle, be filled with N
2, stand-by after temperature 30 DEG C of enrichment 2-4 days, preferred enrichment 3 days.
In described step (3), the supernatant liquor that step (1) obtains is moved at 110 ~ 130 DEG C, sterilizing 10 ~ 40min under 0.06 ~ 0.16MPa; Be cooled to room temperature, join in anaerobic acid-production fermentor tank after regulating pH 6-8, in supernatant liquor, access propionibacterium by 5% ~ 15% inoculum size and carry out stirring anaerobically fermenting, the time of this anaerobically fermenting is 3 ~ 5 days, maintains pH=5.5 ~ 8.5(Ca (OH)
2regulate), temperature 25 ~ 35 DEG C.
In described step (4), centrifugal rotating speed is 500-2000rpm, centrifugation time 2-4min.
In described step (4), in fermented supernatant fluid, propionic acid content is 3.56 ~ 9.95gCOD/L.The kind of rubbish from cooking has a lot, and also have impact to the effect of fermentation, the carbohydrate of rice is more, more easily be converted into propionic acid, and the composition of protein and fat is also conducive to the metabolism growth of microorganism, experimentally result, more favourable to product propionic acid under this ratio
The invention has the beneficial effects as follows:
(1) mud and rubbish from cooking carry out first stage fermentation under weak basic condition, are conducive to the larger molecular organicses such as extracellular microbial exoenzyme hydrolysis sugar, protein, fat.
(2) propionic acid content that subordinate phase fermentation produces is that in the 61.2%-74.6%(fermented supernatant fluid of total acid, propionic acid content is 3.56 ~ 9.95gCOD/L of total acid in supernatant liquor), higher than the propionic acid content of bibliographical information a lot.
Accompanying drawing explanation
Fig. 1 is the technology schematic diagram that in the embodiment of the present invention, pure bacterium improves mud and rubbish from cooking mixed fermentation liquid propionic acid content.
Embodiment
The present invention is further illustrated below in conjunction with embodiment and accompanying drawing.
Rubbish from cooking in the following example, for after the refuses such as rejecting chopsticks, broken bone, plastics, the scraps of paper, fragment, through pulverizing, crossing 10 mesh sieves, obtains the concentrated rubbish from cooking that C/N is 16-20.Wherein the dry weight of rice, meat, oils is than being (3 ~ 10): (0.5 ~ 5): (0 ~ 2); Preferred rice, the dry weight of meat, oils is than being 7:2:1.
Embodiment 1
As shown in Figure 1:
(1) mixture (first stage ferments) of pre fermentation mud and rubbish from cooking;
Learning from else's experience, (reject rubbish from cooking in the present embodiment such as chopsticks, broken bone, plastics, the scraps of paper, fragment is after rejecting the refuses such as chopsticks, broken bone, plastics, the scraps of paper, fragment to the rubbish from cooking pulverized and sieved, through pulverizing, crossing 10 mesh sieves, obtain the concentrated rubbish from cooking that C/N is 16-20; Adding tap water makes water ratio be 80%; Rubbish from cooking main ingredient is rice, and meat, oils are auxiliary, rice, and the best dry weight of meat, oils is than being 7:2:1.) 0.41L, concentrate excess sludge (TSS 20g/L, VSS 14g/L, water ratio are 98%, pH=6.84) with 0.46L to mix (mud and rubbish from cooking dry weight are than being 0.18:1), add 1.63L tap water, be injected into 3L anaerobic fermentation tank (TCOD is 25g COD/L);
(25 DEG C) fermentation 2.5 days under room temperature, stirring velocity is 120rpm and uses Ca (OH)
2control pH is 8.5; Take out fermenting mixture at the centrifugal 5min of 3000rpm, the 2L centrifuged supernatant of acquisition.
(2) activation and enrichment propionibacterium;
Propionibacterium (SICC1.256) is activated rear enrichment from inclined-plane and is stored in the glycerine pipe storage substratum of 20% (culture medium prescription is in table 1), Conservation environment is-80 DEG C; Before using, 20ml glycerine pipe is seeded to 200ml enrichment medium (culture medium prescription is in table 2) according to inoculum size 10%, in culturing bottle, is filled with N
2, in temperature 30 DEG C of enrichments after 3 days, obtain propionibacterium seed liquor.
(3) propionibacterium seed liquor fermented supernatant fluid is utilized to produce propionic acid (subordinate phase fermentation);
By the supernatant liquor arrived of step (1) sterilizing 20min under high-pressure sterilizing pot 121 DEG C, 0.110MPa, be cooled to room temperature, inject 2.5L anaerobic fermentation tank, with Ca (OH)
2pH is adjusted to 7, and accesses the propionibacterium seed liquor (inoculum size 10%) that step (2) obtains, anaerobism stirs fermentation 4 days, maintains pH=7(Ca (OH)
2regulate), temperature 30 DEG C.
(4) after fermentation ends, by mixture at the centrifugal 2min of 1000rmp, the propionic acid amount obtained in supernatant liquor is that 9.95gCOD/L(accounts for 74.6% of total short chain fatty acid).
Embodiment 2
(1) mixture (first stage ferments) of pre fermentation mud and rubbish from cooking;
Learn from else's experience rubbish from cooking (rejecting chopsticks, broken bone, plastics, the scraps of paper, the fragment etc.) 0.37L pulverized and sieved, concentrate excess sludge (TSS 20g/L, VSS 14g/L, water ratio are 98%, pH=6.84) with 0.7L to mix (mud and rubbish from cooking dry weight are than being 0.3:1), add 1.43L tap water, be injected into 3L anaerobic fermentation tank (TCOD is 25g COD/L);
(25 DEG C) fermentation 2.5 days under room temperature, stirring velocity is 250rpm, and uses Ca (OH)
2control pH is 8; Take out fermenting mixture at the centrifugal 5min of 3000rpm, the 2L centrifuged supernatant of acquisition.
(2) activation and enrichment propionibacterium;
Propionibacterium (SICC1.256) is activated rear enrichment from inclined-plane and is stored in the glycerine pipe storage substratum of 10% (culture medium prescription is in table 1), Conservation environment is-80 DEG C; Before using, 10ml glycerine pipe is seeded to 100ml enrichment medium (culture medium prescription is in table 2) according to inoculum size 10%, in culturing bottle, is filled with N
2, in temperature 30 DEG C of enrichments after 4 days, obtain propionibacterium seed liquor.
(3) propionibacterium seed liquor fermented supernatant fluid is utilized to produce propionic acid (subordinate phase fermentation);
By the supernatant liquor arrived of step (1) sterilizing 20min under high-pressure sterilizing pot 121 DEG C, 0.110MPa, be cooled to room temperature, inject 2.5L anaerobic fermentation tank, with Ca (OH) 2, pH is adjusted to 7, and access the propionibacterium seed liquor (inoculum size 10%) that step (2) obtains, anaerobism stirs fermentation 4 days, maintains pH=7(Ca (OH)
2regulate), temperature 30 DEG C.
(4) after fermentation ends, by mixture at the centrifugal 2min of 1000rmp, the propionic acid amount obtained in supernatant liquor is that 6.10gCOD/L(accounts for 68.2% of total short chain fatty acid).
Embodiment 3
(1) mixture (first stage ferments) of pre fermentation mud and rubbish from cooking;
Learn from else's experience rubbish from cooking (rejecting chopsticks, broken bone, plastics, the scraps of paper, the fragment etc.) 0.45L pulverized and sieved, concentrate excess sludge (TSS 20g/L, VSS 14g/L, water ratio are 98%, pH=6.84) with 0.22L to mix (mud and rubbish from cooking dry weight are than being 0.08:1), add 1.83L tap water, be injected into 3L anaerobic fermentation tank (TCOD is 25g COD/L);
(25 DEG C) fermentation 2.5 days under room temperature, stirring velocity is 60rpm, and uses Ca (OH)
2control pH is 8; Take out fermenting mixture at the centrifugal 5min of 3000rpm, the 2L centrifuged supernatant of acquisition.
(2) activation and enrichment propionibacterium;
Propionibacterium (SICC1.256) is activated rear enrichment from inclined-plane and is stored in the glycerine pipe storage substratum of 15% (culture medium prescription is in table 1), Conservation environment is-80 DEG C; Before using, 30ml glycerine pipe is seeded to 200ml enrichment medium (culture medium prescription is in table 2) according to inoculum size 15%, in culturing bottle, is filled with N
2, in temperature 30 DEG C of enrichments after 2 days, obtain propionibacterium seed liquor.
(3) propionibacterium seed liquor fermented supernatant fluid is utilized to produce propionic acid (subordinate phase fermentation);
By the supernatant liquor arrived of step (1) sterilizing 20min under high-pressure sterilizing pot 121 DEG C, 0.110MPa, be cooled to room temperature, inject 2.5L anaerobic fermentation tank, with Ca (OH) 2, pH is adjusted to 7, and access the propionibacterium seed liquor (inoculum size 10%) that step (2) obtains, anaerobism stirs fermentation 4 days, maintains pH=7(Ca (OH)
2regulate), temperature 30 DEG C.
(4) after fermentation ends, by mixture at the centrifugal 2min of 1000rmp, the propionic acid amount obtained in supernatant liquor is that 8.70gCOD/L(accounts for 63.9% of total short chain fatty acid).
Embodiment 4
(1) mixture (first stage ferments) of pre fermentation mud and rubbish from cooking;
Learn from else's experience rubbish from cooking (rejecting chopsticks, broken bone, plastics, the scraps of paper, the fragment etc.) 0.41L pulverized and sieved, concentrate excess sludge (TSS 20g/L, VSS 14g/L, water ratio are 98%, pH=6.84) with 0.46L to mix (mud and rubbish from cooking dry weight are than being 0.18:1), add 1.63L tap water, be injected into 3L anaerobic fermentation tank (TCOD is 25g COD/L);
(25 DEG C) fermentation 2.5 days under room temperature, stirring velocity is 120rpm, and uses Ca (OH)
2control pH is 5; Take out fermenting mixture at the centrifugal 3min of 2000rpm, the 2L centrifuged supernatant of acquisition.
(2) activation and enrichment propionibacterium;
Propionibacterium (SICC1.256) is activated rear enrichment from inclined-plane and is stored in the glycerine pipe storage substratum of 30% (culture medium prescription is in table 1), Conservation environment is-80 DEG C; Before using, 20ml glycerine pipe is seeded to 200ml enrichment medium (culture medium prescription is in table 2) according to inoculum size 10%, in culturing bottle, is filled with N
2, in temperature 30 DEG C of enrichments after 3 days, obtain propionibacterium seed liquor.
(3) propionibacterium seed liquor fermented supernatant fluid is utilized to produce propionic acid (subordinate phase fermentation);
By the supernatant liquor arrived of step (1) sterilizing 20min under high-pressure sterilizing pot 121 DEG C, 0.110MPa, be cooled to room temperature, inject 2.5L anaerobic fermentation tank, with Ca (OH) 2, pH is adjusted to 7, and access the propionibacterium seed liquor (inoculum size 10%) that step (2) obtains, anaerobism stirs fermentation 4 days, maintains pH=7(Ca (OH) 2 and regulates), temperature 30 DEG C.
(4) after fermentation ends, by mixture at the centrifugal 2min of 500rmp, the propionic acid amount obtained in supernatant liquor is that 3.56gCOD/L(accounts for 61.2% of total short chain fatty acid).
Embodiment 5
(1) mixture (first stage ferments) of pre fermentation mud and rubbish from cooking;
Learn from else's experience rubbish from cooking (rejecting chopsticks, broken bone, plastics, the scraps of paper, the fragment etc.) 0.41L pulverized and sieved, concentrate excess sludge (TSS 20g/L, VSS 14g/L, water ratio are 98%, pH=6.84) with 0.46L to mix (mud and rubbish from cooking dry weight are than being 0.18:1), add 1.63L tap water, be injected into 3L anaerobic fermentation tank (TCOD is 25g COD/L);
(25 DEG C) fermentation 2.5 days under room temperature, stirring velocity is 120rpm, and uses Ca (OH)
2control pH is 12; Take out fermenting mixture at the centrifugal 6min of 4000rpm, the 2L centrifuged supernatant of acquisition.
(2) activation and enrichment propionibacterium;
Propionibacterium (SICC1.256) is activated rear enrichment from inclined-plane and is stored in the glycerine pipe storage substratum of 20% (culture medium prescription is in table 1), Conservation environment is-80 DEG C; Before using, 20ml glycerine pipe is seeded to 200ml enrichment medium (culture medium prescription is in table 2) according to inoculum size 10%, in culturing bottle, is filled with N
2, in temperature 30 DEG C of enrichments after 3 days, obtain propionibacterium seed liquor.
(3) propionibacterium seed liquor fermented supernatant fluid is utilized to produce propionic acid (subordinate phase fermentation);
By the supernatant liquor arrived of step (1) sterilizing 20min under high-pressure sterilizing pot 121 DEG C, 0.110MPa, be cooled to room temperature, inject 2.5L anaerobic fermentation tank, with Ca (OH)
2pH is adjusted to 7, and accesses the propionibacterium seed liquor (inoculum size 10%) that step (2) obtains, anaerobism stirs fermentation 5 days, maintains pH=7(Ca (OH)
2regulate), temperature 30 DEG C.
(4) after fermentation ends, by mixture at the centrifugal 4min of 1500rmp, the propionic acid amount obtained in supernatant liquor is that 7.50gCOD/L(accounts for 68.9% of total short chain fatty acid).
Embodiment 6
(1) mixture (first stage ferments) of pre fermentation mud and rubbish from cooking;
Learn from else's experience rubbish from cooking (rejecting chopsticks, broken bone, plastics, the scraps of paper, the fragment etc.) 0.41L pulverized and sieved, concentrate excess sludge (TSS 20g/L, VSS 14g/L, water ratio are 98%, pH=6.84) with 0.46L to mix (mud and rubbish from cooking dry weight are than being 0.18:1), add 1.63L tap water, be injected into 3L anaerobic fermentation tank (TCOD is 25g COD/L);
(25 DEG C) fermentation 1.5 days under room temperature, stirring velocity is 120rpm, and uses Ca (OH)
2control pH is 8; Take out fermenting mixture at the centrifugal 5min of 3000rpm, the 2L centrifuged supernatant of acquisition.
(2) activation and enrichment propionibacterium;
Propionibacterium (SICC1.256) is activated rear enrichment from inclined-plane and is stored in the glycerine pipe storage substratum of 20% (culture medium prescription is in table 1), Conservation environment is-80 DEG C; Before using, 30ml glycerine pipe is seeded to 300ml enrichment medium (culture medium prescription is in table 2) according to inoculum size 10%, in culturing bottle, is filled with N
2, in temperature 30 DEG C of enrichments after 3 days, obtain propionibacterium seed liquor.
(3) propionibacterium seed liquor fermented supernatant fluid is utilized to produce propionic acid (subordinate phase fermentation);
By the supernatant liquor arrived of step (1) sterilizing 40min under high-pressure sterilizing pot 110 DEG C, 0.06MPa, be cooled to room temperature, inject 2.5L anaerobic fermentation tank, with Ca (OH)
2pH is adjusted to 7, and accesses the propionibacterium seed liquor (inoculum size 15%) that step (2) obtains, anaerobism stirs fermentation 4 days, maintains pH=7(Ca (OH)
2regulate), temperature 30 DEG C.
(4) after fermentation ends, by mixture at the centrifugal 3min of 1000rmp, the propionic acid amount obtained in supernatant liquor is that 5.70gCOD/L(accounts for 69.2% of total short chain fatty acid).
Embodiment 7
(1) mixture (first stage ferments) of pre fermentation mud and rubbish from cooking;
Learn from else's experience rubbish from cooking (rejecting chopsticks, broken bone, plastics, the scraps of paper, the fragment etc.) 0.41L pulverized and sieved, concentrate excess sludge (TSS 20g/L, VSS 14g/L, water ratio are 98%, pH=6.84) with 0.46L to mix (mud and rubbish from cooking dry weight are than being 0.18:1), add 1.63L tap water, be injected into 3L anaerobic fermentation tank (TCOD is 25g COD/L);
(25 DEG C) fermentation 5 days under room temperature, stirring velocity is 120rpm, and uses Ca (OH)
2control pH is 8; Take out fermenting mixture at the centrifugal 5min of 3000rpm, the 2L centrifuged supernatant of acquisition.
(2) activation and enrichment propionibacterium;
Propionibacterium (SICC1.256) is activated rear enrichment from inclined-plane and is stored in the glycerine pipe storage substratum of 20% (culture medium prescription is in table 1), Conservation environment is-80 DEG C; Before using, 10ml glycerine pipe is seeded to 200ml enrichment medium (culture medium prescription is in table 2) according to inoculum size 5%, in culturing bottle, is filled with N
2, in temperature 30 DEG C of enrichments after 4 days, obtain propionibacterium seed liquor.
(3) propionibacterium seed liquor fermented supernatant fluid is utilized to produce propionic acid (subordinate phase fermentation);
By the supernatant liquor arrived of step (1) sterilizing 10min under high-pressure sterilizing pot 130 DEG C, 0.116MPa, be cooled to room temperature, inject 2.5L anaerobic fermentation tank, with Ca (OH)
2pH is adjusted to 7, and accesses the propionibacterium seed liquor (inoculum size 10%) that step (2) obtains, anaerobism stirs fermentation 4 days, maintains pH=7(Ca (OH)
2regulate), temperature 30 DEG C.
(4) after fermentation ends, by mixture at the centrifugal 3min of 1000rmp, the propionic acid amount obtained in supernatant liquor is that 7.32gCOD/L(accounts for 64.2% of total short chain fatty acid).
Embodiment 8
(1) mixture (first stage ferments) of pre fermentation mud and rubbish from cooking;
Learn from else's experience rubbish from cooking (rejecting chopsticks, broken bone, plastics, the scraps of paper, the fragment etc.) 0.41L pulverized and sieved, concentrate excess sludge (TSS 20g/L, VSS 14g/L, water ratio are 98%, pH=6.84) with 0.46L to mix (mud and rubbish from cooking dry weight are than being 0.18:1), add 1.63L tap water, be injected into 3L anaerobic fermentation tank (TCOD is 25g COD/L);
In temperature 10 DEG C fermentation 2.5 days, stirring velocity was 120rpm, and uses Ca (OH)
2control pH is 8; Take out fermenting mixture at the centrifugal 5min of 3000rpm, the 2L centrifuged supernatant of acquisition.
(2) activation and enrichment propionibacterium;
Propionibacterium (SICC1.256) is activated rear enrichment from inclined-plane and is stored in the glycerine pipe storage substratum of 10% (culture medium prescription is in table 1), Conservation environment is-80 DEG C; Before using, 30ml glycerine pipe is seeded to 200ml enrichment medium (culture medium prescription is in table 2) according to inoculum size 15%, in culturing bottle, is filled with N
2, in temperature 30 DEG C of enrichments after 2 days, obtain propionibacterium seed liquor.
(3) propionibacterium seed liquor fermented supernatant fluid is utilized to produce propionic acid (subordinate phase fermentation);
By the supernatant liquor arrived of step (1) sterilizing 20min under high-pressure sterilizing pot 121 DEG C, 0.110MPa, be cooled to room temperature, inject 2.5L anaerobic fermentation tank, with Ca (OH)
2pH is adjusted to 7, and accesses the propionibacterium seed liquor (inoculum size 10%) that step (2) obtains, anaerobism stirs fermentation 4 days, maintains pH=7(Ca (OH)
2regulate), temperature 30 DEG C.
(4) after fermentation ends, by mixture at the centrifugal 3min of 1000rmp, the propionic acid amount obtained in supernatant liquor is that 5.48gCOD/L(accounts for 71.2% of total short chain fatty acid).
Embodiment 9
(1) mixture (first stage ferments) of pre fermentation mud and rubbish from cooking;
Learn from else's experience rubbish from cooking (rejecting chopsticks, broken bone, plastics, the scraps of paper, the fragment etc.) 0.41L pulverized and sieved, concentrate excess sludge (TSS 20g/L, VSS 14g/L, water ratio are 98%, pH=6.84) with 0.46L to mix (mud and rubbish from cooking dry weight are than being 0.18:1), add 1.63L tap water, be injected into 3L anaerobic fermentation tank (TCOD is 25g COD/L);
In temperature 65 DEG C fermentation 2 days, stirring velocity was 120rpm, and uses Ca (OH)
2control pH is 8; Take out fermenting mixture at the centrifugal 6min of 4000rpm, the 2L centrifuged supernatant of acquisition.
(2) activation and enrichment propionibacterium;
Propionibacterium (SICC1.256) is activated rear enrichment from inclined-plane and is stored in the glycerine pipe storage substratum of 20% (culture medium prescription is in table 1), Conservation environment is-80 DEG C; Before using, 20ml glycerine pipe is seeded to 200ml enrichment medium (culture medium prescription is in table 2) according to inoculum size 10%, in culturing bottle, is filled with N
2, in temperature 30 DEG C of enrichments after 3 days, obtain propionibacterium seed liquor.
(3) propionibacterium seed liquor fermented supernatant fluid is utilized to produce propionic acid (subordinate phase fermentation);
By the supernatant liquor arrived of step (1) sterilizing 20min under high-pressure sterilizing pot 121 DEG C, 0.110MPa, be cooled to room temperature, inject 2.5L anaerobic fermentation tank, with Ca (OH)
2pH is adjusted to 7, and accesses the propionibacterium seed liquor (inoculum size 10%) that step (2) obtains, anaerobism stirs fermentation 4 days, maintains pH=7(Ca (OH)
2regulate), temperature 30 DEG C.
(4) after fermentation ends, by mixture at the centrifugal 3min of 1000rmp, the propionic acid amount obtained in supernatant liquor is that 9.45gCOD/L(accounts for 73.3% of total short chain fatty acid).
Embodiment 10
(1) mixture (first stage ferments) of pre fermentation mud and rubbish from cooking;
Learn from else's experience rubbish from cooking (rejecting chopsticks, broken bone, plastics, the scraps of paper, the fragment etc.) 0.41L pulverized and sieved, concentrate excess sludge (TSS 20g/L, VSS 14g/L, water ratio are 98%, pH=6.84) with 0.46L to mix (mud and rubbish from cooking dry weight are than being 0.18:1), add 1.63L tap water, be injected into 3L anaerobic fermentation tank (TCOD is 25g COD/L);
(25 DEG C) fermentation 2.5 days under room temperature, stirring velocity is 120rpm, and uses Ca (OH)
2control pH is 8; Take out fermenting mixture at the centrifugal 5min of 3000rpm, the 2L centrifuged supernatant of acquisition.
(2) activation and enrichment propionibacterium;
Propionibacterium (SICC1.256) is activated rear enrichment from inclined-plane and is stored in the glycerine pipe storage substratum of 20% (culture medium prescription is in table 1), Conservation environment is-80 DEG C; Before using, 30ml glycerine pipe is seeded to 300ml enrichment medium (culture medium prescription is in table 2) according to inoculum size 10%, in culturing bottle, is filled with N
2, in temperature 30 DEG C of enrichments after 3 days, obtain propionibacterium seed liquor.
(3) propionibacterium seed liquor fermented supernatant fluid is utilized to produce propionic acid (subordinate phase fermentation);
By the supernatant liquor arrived of step (1) sterilizing 20min under high-pressure sterilizing pot 121 DEG C, 0.110MPa, be cooled to room temperature, inject 2.5L anaerobic fermentation tank, with Ca (OH)
2pH is adjusted to 5.5, and accesses the propionibacterium seed liquor (inoculum size 15%) that step (2) obtains, anaerobism stirs fermentation 3 days, maintains pH=5.5(Ca (OH)
2regulate), temperature 25 DEG C.
(4) after fermentation ends, by mixture at the centrifugal 3min of 1000rmp, the propionic acid amount obtained in supernatant liquor is that 10.12gCOD/L(accounts for 72.1% of total short chain fatty acid).
Embodiment 11
(1) mixture (first stage ferments) of pre fermentation mud and rubbish from cooking;
Learn from else's experience rubbish from cooking (rejecting chopsticks, broken bone, plastics, the scraps of paper, the fragment etc.) 0.41L pulverized and sieved, concentrate excess sludge (TSS 20g/L, VSS 14g/L, water ratio are 98%, pH=6.84) with 0.46L to mix (mud and rubbish from cooking dry weight are than being 0.18:1), add 1.63L tap water, be injected into 3L anaerobic fermentation tank (TCOD is 25g COD/L);
(25 DEG C) fermentation 1.5 days under room temperature, stirring velocity is 120rpm, and uses Ca (OH)
2control pH is 8; Take out fermenting mixture at the centrifugal 5min of 3000rpm, the 2L centrifuged supernatant of acquisition.
(2) activation and enrichment propionibacterium;
Propionibacterium (SICC1.256) is activated rear enrichment from inclined-plane and is stored in the glycerine pipe storage substratum of 20% (culture medium prescription is in table 1), Conservation environment is-80 DEG C; Before using, 30ml glycerine pipe is seeded to 300ml enrichment medium (culture medium prescription is in table 2) according to inoculum size 10%, in culturing bottle, is filled with N
2, in temperature 30 DEG C of enrichments after 3 days, obtain propionibacterium seed liquor.
(3) propionibacterium seed liquor fermented supernatant fluid is utilized to produce propionic acid (subordinate phase fermentation);
By the supernatant liquor arrived of step (1) sterilizing 20min under high-pressure sterilizing pot 121 DEG C, 0.110MPa, be cooled to room temperature, inject 2.5L anaerobic fermentation tank, with Ca (OH)
2pH is adjusted to 8.5, and accesses the propionibacterium seed liquor (inoculum size 15%) that step (2) obtains, anaerobism stirs fermentation 3 days, maintains pH=8.5(Ca (OH)
2regulate), temperature 35 DEG C.
(4) after fermentation ends, by mixture at the centrifugal 3min of 1000rmp, the propionic acid amount obtained in supernatant liquor is that 6.32gCOD/L(accounts for 68.2% of total short chain fatty acid).
Comparative example 1
Do not inoculate the situation of propionibacterium:
Learn from else's experience rubbish from cooking (rejecting chopsticks, broken bone, plastics, the scraps of paper, the fragment etc.) 0.25L pulverized and sieved, add 0.75L tap water, concentrate excess sludge (TSS 20g/L, VSS 14g/L, water ratio are 98%, pH=6.84) with 1.5L to mix (mud and rubbish from cooking dry weight are than being 0.9:1), be injected into 3L anaerobic fermentation tank (TCOD is 26.90g COD/L), (25 DEG C) fermentation 8 days under room temperature, stirring velocity is 120rpm, and uses Ca (OH)
2control pH is 8.
After fermentation ends, mixture is centrifugal, the propionic acid amount obtained in supernatant liquor is that 4.73gCOD/L(accounts for 49.1% of total short chain fatty acid)
The fermentation of comparative example 2 first stage is only with rubbish from cooking fermentation, and mud drops into considerably less, only does the inoculation of hydrolysis bacterium.
Learn from else's experience rubbish from cooking (rejecting chopsticks, broken bone, plastics, the scraps of paper, the fragment etc.) 0.57L pulverized and sieved, add 1.43L tap water, concentrate excess sludge (TSS 20g/L, VSS 14g/L, water ratio are 98%, pH=6.84) with 0.2L to mix (dry weight is than being 0.01:1), be injected into 3L anaerobic fermentation tank (TCOD is 39.08g COD/L), and use Ca (OH)
2control pH is 8, and under room temperature, (25 DEG C) fermentation is after 2.5 days, takes out fermenting mixture at the centrifugal 4min of 2000rpm, the 2L centrifuged supernatant sterilizing 20min under high-pressure sterilizing pot 121 DEG C, 0.110MPa obtained, be cooled to room temperature, pH is adjusted to 7, access 200ml enrichment bacterium liquid.Maintain reaction system pH=7, temperature 30 DEG C, anaerobism stirs fermentation 5 days.
After fermentation ends, mixture is centrifugal, the propionic acid amount obtained in supernatant liquor is that 1.15gCOD/L(accounts for 62.7% of total short chain fatty acid).
The fermentation of comparative example 3 first stage only uses sludge fermentation, does not relate to rubbish from cooking
Only get 2.5L and concentrate excess sludge (TSS 20g/L, VSS 14g/L, water ratio are 98%, pH=6.84), use Ca (OH)
2control pH is 8 mixing, under room temperature, (25 DEG C) fermentation is after 2.5 days, take out fermenting mixture at the centrifugal 5min of 2000rpm, the 2L centrifuged supernatant sterilizing 20min under high-pressure sterilizing pot 121 DEG C, 0.110MPa obtained, be cooled to room temperature, pH is adjusted to 7, and 200ml enrichment bacterium liquid is injected 2.5L anaerobic acid-production fermentor tank simultaneously.Maintain reaction system pH=7, temperature 30 DEG C, anaerobism stirs fermentation 5 days.
After fermentation ends, mixture is centrifugal, the propionic acid amount obtained in supernatant liquor is that 0.6gCOD/L(accounts for 20.1% of total short chain fatty acid).
Embodiment 1-10 greatly can improve propionic acid content compared with the prior art in comparative example 1.
Table 1 glycerin storage culture medium prescription
Table 2 enrichment culture based formulas
Above-mentioned is can understand and apply the invention for ease of those skilled in the art to the description of embodiment.Person skilled in the art obviously easily can make various amendment to these embodiments, and General Principle described herein is applied in other embodiments and need not through performing creative labour.Therefore, the invention is not restricted to embodiment here, those skilled in the art, according to announcement of the present invention, do not depart from improvement that scope makes and amendment all should within protection scope of the present invention.
Claims (11)
1. improve the method that batch fermentation produces propionic acid content in acid solution, it is characterized in that: comprise following steps:
(1) by rubbish from cooking, thickened sludge mixing, join anaerobically fermenting in anaerobic hydrolysis fermentor tank, then by mixture centrifugation, take out supernatant liquor stand-by;
(2) also enrichment propionibacterium seed liquor is activated;
(3), by the supernatant liquor sterilizing that step (1) obtains, in the propionibacterium seed liquor that access step (2) obtains, in anaerobic fermentation tank, propionic acid is produced in fermentation;
(4) after fermentation ends, fermenting mixture is centrifugal, obtain the fermented supernatant fluid being rich in propionic acid;
In described step (1), thickened sludge and rubbish from cooking are by dry weight ratio (0.08 ~ 0.18): 1 mixes, and the total chemical oxygen demand (COD) added after tap water dilution is 20-40g COD/L;
In described step (1), the temperature of anaerobically fermenting is 25 DEG C, and stirring velocity is 60-250rpm, with Ca (OH)
2the pH controlling fermentation is 8-8.5, and fermentation time is 2-5 days;
In described step (2), propionibacterium is activated rear enrichment from inclined-plane and is stored in the glycerine pipe storage substratum of 10%-30%, Conservation environment is-50 to-80 DEG C; Be seeded to enrichment medium according to inoculum size 5-15% at every turn, in culturing bottle, be filled with N
2, stand-by after temperature 30 DEG C of enrichment 2-4 days;
Described propionibacterium is SICC1.256;
The formula that glycerine pipe stores substratum is: yeast extract or corn steep liquor 5g/L, enzymic hydrolysis casein or pancreatin hydrolysis casein 10g/L, K
2hPO
4h
2o2.5g/L, KH
2pO
41.5g/L, glucose or Sodium.alpha.-hydroxypropionate 5g/L and glycerine 200g/L, and use KOH/HCL to regulate pH to be 6.8-7.2;
The formula of enrichment medium is: yeast extract or corn steep liquor 10g/L, enzymic hydrolysis casein or pancreatin hydrolysis casein 10g/L, K
2hPO
4h
2o2.5g/L, KH
2pO
41.5g/L, glucose or Sodium.alpha.-hydroxypropionate 20g/L, and use KOH/HCL to regulate pH to be 6.8-7.2; In described step (3), the supernatant liquor that step (1) obtains is moved at 110 ~ 130 DEG C, sterilizing 10 ~ 40min under 0.06 ~ 0.16MPa; Be cooled to room temperature, join in anaerobic acid-production fermentor tank after regulating pH 6-8, in supernatant liquor, access propionibacterium by 5% ~ 15% inoculum size and carry out stirring anaerobically fermenting, the time of this anaerobically fermenting is 3 ~ 5 days, Ca (OH)
2maintain pH=5.5 ~ 8.5, temperature 25 ~ 35 DEG C.
2. method according to claim 1, is characterized in that: in described step (1), and the water ratio of thickened sludge is 98%-99%, and total suspended solid concentration is 10-20g/L.
3. method according to claim 1, is characterized in that: in described step (1), total suspended solid concentration is 15g/L.
4. method according to claim 1, is characterized in that: described rubbish from cooking, for after rejecting chopsticks, broken bone, plastics, the scraps of paper, fragment refuse, through pulverizing, crossing 10 mesh sieves, obtains the concentrated rubbish from cooking that C/N is 16-20; Adding tap water makes water ratio be 80%; Wherein, the dry weight of rice, meat, oils is than being (3 ~ 10): (0.5 ~ 5): (0 ~ 2).
5. method according to claim 1, is characterized in that: the dry weight of rice, meat, oils is than being 7:2:1.
6. method according to claim 1, is characterized in that: in described step (1), thickened sludge mixes than 0.18:1 by dry weight with rubbish from cooking.
7. method according to claim 1, is characterized in that: in described step (1), the pH of fermentation is 8, and fermentation time is 4 days.
8. method according to claim 1, is characterized in that: in described step (1), and the rotating speed of centrifugation is 2000-4000rpm, and centrifugation time is 4-6min.
9. method according to claim 1, is characterized in that: in described step (2), be seeded to enrichment medium at every turn, in culturing bottle, be filled with N according to inoculum size 5-15%
2, in temperature 30 DEG C of enrichments 3 days.
10. method according to claim 1, is characterized in that: in described step (4), centrifugal rotating speed is 500-2000rpm, centrifugation time 2-4min.
11. methods according to claim 1, is characterized in that: in described step (4), in fermented supernatant fluid, propionic acid content is 3.56 ~ 9.95gCOD/L.
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| 响应面法优化产酸丙酸杆菌丙酸发酵条件的研究;刘寅;《食品工业科技》;20100531;第31卷(第5期);1.1材料与仪器，1.2实验方法 * |
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