CN112790273A

CN112790273A - Method for recovering L-lactic acid fermentation thallus and continuously producing fermented feed

Info

Publication number: CN112790273A
Application number: CN202011641449.8A
Authority: CN
Inventors: 穆晓玲; 孟瑶; 王舒; 秦晴; 毕从胜; 常乐
Original assignee: Anhui BBCA Fermentation Technology Engineering Research Co Ltd
Current assignee: Anhui BBCA Fermentation Technology Engineering Research Co Ltd
Priority date: 2020-12-31
Filing date: 2020-12-31
Publication date: 2021-05-14

Abstract

The invention discloses a method for recovering L-lactic acid fermentation thalli and continuously producing fermented feed, which comprises the steps of size mixing, solid-liquid separation, mixing the obtained concentrated phase with feed raw materials for fermentation, and further performing continuous fermentation to obtain the fermented feed. According to the invention, by optimizing the size mixing mode, the filtering treatment mode and the fermentation mode, the size mixing cost can be saved, and the wastewater discharge can be reduced. The invention mixes the recovered L-lactic acid fermentation thalli and lactate compound with feed raw materials for continuous fermentation, can continuously produce fermented feed, and greatly reduces the fermentation period of the feed. The fermented feed provided by the invention can effectively improve the growth rate of fattening livestock and reduce the meat-feed ratio and the diarrhea rate.

Description

Method for recovering L-lactic acid fermentation thallus and continuously producing fermented feed

Technical Field

The invention belongs to the technical field of microbial fermentation, and relates to a method for recovering L-lactic acid fermentation thalli and continuously producing fermented feed.

Background

Bacillus coagulans is formally listed in feed additive variety catalog (2013) by Ministry of agriculture in 12 and 30 months in 2013. The bacillus coagulans as a novel microorganism feed additive can play a role in replacing resistance and reducing resistance. Meanwhile, the bacillus coagulans is facultative anaerobe, can grow in aerobic and anaerobic environments, can adapt to the hypoxic intestinal environment, has high tolerance to acid and bile, can reduce the pH value of the intestinal tract by the generated L-lactic acid, inhibits harmful bacteria, and can promote the growth and reproduction of beneficial bacteria such as bifidobacterium and the like. Bacillus coagulans is able to form spores, which is beneficial for restoring the microecological balance of the gastrointestinal tract compared to other non-lactic acid producing bacilli. The survival rate of the bacillus coagulans after being treated by artificial gastric juice (pH 1.4) for 2 hours is 95.29%, and the result of measuring the number of the viable bacteria after being treated in a drying oven at 55 ℃ for 60 hours shows that the viable bacteria are not less than 1 hundred million cfu/g.

Bacillus coagulans is not an indigenous micro-organism in the intestine, and its physiological role in the intestine is a result of secretion of various beneficial substances and synergistic effects with other probiotics in the intestine, not a substance. After the bacillus coagulans enters the stomach by oral administration, the bacillus coagulans is activated under the action of gastric juice, the spore coat expands, the shape of the spore is enlarged, the water content is increased, and the metabolism is accelerated. When bacillus coagulans enters the duodenum, its spores germinate into vegetative cells. Vegetative cells start to grow and reproduce after entering the small intestine, and reproduce for one generation in about 30 min. The bacillus coagulans consumes free oxygen to propagate in the intestinal tract after entering the intestinal tract, and is beneficial to the growth of anaerobic microorganisms such as lactobacillus and bifidobacterium, thereby regulating the balance of microbial flora in the intestinal tract, improving the immunity and disease resistance of organisms and reducing the occurrence of intestinal diseases. The bacillus coagulans can also secrete amylase and protease in the intestinal reproduction process, so that the digestion and absorption of organisms on nutrient substances are promoted; the produced B vitamins, amino acids, short chain fatty acids and other substances can increase the peristalsis speed of the small intestine, thereby improving the digestion function of the intestinal tract. In addition, after the bacillus coagulans is settled in the intestinal tract, a large amount of bacteriostatic substances such as coagulans and L-lactic acid which can inhibit harmful bacteria can be generated, so that the bacillus coagulans has a certain treatment effect on gastrointestinal inflammation.

At present, most of bacillus coagulans products take the effect in the form of a microbial preparation as a feed additive, the strain culture and enrichment process is complicated, the target income is single, and the bacillus coagulans product does not contain or contains a very small amount of lactic acid and lactate thereof as a food additive. The microbial feed yield is limited by the yield of bacillus coagulans, and the bacillus coagulans starts to carry out metabolic activity only when entering a digestive system, so that the action time is short, the effect is small, and the utilization rate is low.

Disclosure of Invention

In order to overcome the defects of the prior art, the invention solves the problems through the following technical scheme.

The invention provides a method for recovering L-lactic acid fermentation thalli and continuously producing fermented feed, which comprises the following steps:

(1) mixing the L-lactic acid fermentation liquor with the elution waste liquor of the ion exchange process to obtain mixed liquor;

(2) carrying out solid-liquid separation on the mixed slurry to obtain a clear liquid and a concentrated phase, and mixing the concentrated phase with feed raw materials for fermentation to obtain a seed fermented feed;

(3) and mixing the seed fermented feed serving as seeds with feed raw materials for continuous fermentation to continuously obtain the fermented feed.

In one embodiment of the present invention, the L-lactic acid-fermenting bacteria is Bacillus coagulans.

In one embodiment of the present invention, in step (1), the elution waste liquid of the ion exchange process is eluted with 1% -5% sulfuric acid or hydrochloric acid, and the L-lactic acid fermentation broth is slurried with the elution waste liquid to pH 4-6, preferably to pH 4.5-5. The elution in the ion exchange step is an elution of a cation exchange column performed in the ion exchange step in the L-lactic acid production and extraction step.

As an embodiment of the invention, in the step (2), the solid-liquid separation mode is centrifugation, pressure filtration or membrane filtration; preferably, the solid-liquid separation equipment is selected from one or more of a vacuum rotary drum filter, a disc type ultrafiltration centrifuge, a plate filter press and a ceramic membrane; more preferably, the solid-liquid separation equipment is a plate filter press or a ceramic membrane.

After solid-liquid separation, the yield of the L-lactic acid in the clear liquid can reach 90-98 percent, and the clear liquid can enter the subsequent extraction process; the concentrated phase is equivalent to the concentration of the slurry adjusting liquid and is concentrated to 5 to 50 times, and the content of bacillus coagulans in the concentrated phase reaches 10⁹-10¹²cfu/g。

In one embodiment of the invention, in the step (2), the mass of the concentrated phase is 0.5-5% of the mass of the feed raw materials during mixing, the fermentation time is 24-36 h, more preferably 24h, and the bacillus coagulans in the seed fermented feed is 10⁶-10⁸cfu/g。

In one embodiment of the invention, in the step (3), the mass of the seed fermented feed is 10-45% of the mass of the feed raw materials, preferably 10-30%, the fermentation time is 48-72 h, preferably 48h, so that the bacillus coagulans in the fermented feed is 10⁵-10⁶cfu/g。

As an embodiment of the invention, in the step (3), during the continuous fermentation process or after the continuous fermentation reaches a certain batch, when the content of the bacillus coagulans is less than or equal to 10⁵When cfu/g, 10% of the seed fermented feed is added, and the fermented feed can be continuously produced.

As an embodiment of the invention, the fermentation temperature of the invention is constant temperature 40-60 ℃, and the preferred fermentation temperature is constant temperature 50 ℃.

In one embodiment of the present invention, the L-lactic acid fermentation broth contains one or more of lactic acid, potassium lactate, sodium lactate, calcium lactate, magnesium lactate, ammonium lactate, and ferrous lactate. Furthermore, the fermented feed of the present invention may also contain one or more of lactic acid, potassium lactate, sodium lactate, calcium lactate, magnesium lactate, ammonium lactate, and ferrous lactate.

As an embodiment of the invention, the feed raw material contains 55-75% of corn flour, 8-16% of bran and 15-30% of soybean meal by mass of 100%; preferably, the feed raw material contains 65% of corn flour, 12% of bran and 23% of soybean meal by mass of 100%.

The invention also provides a fermented feed obtained by any one of the methods.

The invention also provides application of the fermented feed in improving the growth rate of fattening livestock, reducing the meat-feed ratio and/or reducing the diarrhea rate. Including but not limited to pigs, sheep and cattle.

In some specific embodiments, because the period of continuously producing the fermented feed is short, only 2-4 fermenting tanks are needed to continuously produce the fermented feed, the equipment investment is greatly reduced, compared with the conventional premix, the concentrated phase of the L-lactic acid fermentation liquor used for producing the fermented feed belongs to a byproduct in the L-lactic acid production, and the cost price of the raw materials of the fermented feed is reduced by 6 percent compared with the cost of the premix. Compared with the conventional premix, the fermented feed can improve the daily food intake of the fattening pigs, meanwhile, macromolecules such as starch, cellulose, protein and the like in the fermented feed are metabolized into micromolecular active peptides, oligopeptides, monosaccharides and oligosaccharides which are easier to be digested and absorbed by animals, and simultaneously, a plurality of digestive enzymes, amino acids and vitamins generated by metabolism are absorbed and utilized by the animals as nutrient substances, so that the nutrition level and the feed utilization rate of the feed are obviously improved, and the comparison result of the fattening pig fed with the fermented feed with the conventional premix for 75 days shows that the weight gain of the fattening pigs is improved by 12.02%, the meat-feed ratio is reduced by 3.86%, and the diarrhea rate is reduced by 56.48%.

The method for recovering L-lactic acid fermentation bacteria and continuously producing fermented feed optimizes a size mixing mode, a bacteria filtering treatment mode and a fermentation mode of fermentation liquor on the basis of the existing L-lactic acid fermentation extraction technology, and applies the obtained L-lactic acid fermentation bacteria and lactate compound to the fermented feed. The invention uses the elution waste liquid of the cation exchange procedure in the L-lactic acid extraction process to size-mix and filter the fermentation liquor, saves the size-mixing cost, reduces the waste water discharge, and can recover the eluted cations to generate lactate to be used as the fermentation feed additive. The invention does not need heating and concentrated sulfuric acid for acid adjustment treatment for the thallus treatment in the fermentation liquor, obtains the viable bacteria and simultaneously avoids the burden of macromolecular organic matters generated by thallus breaking and death in the fermentation liquor on the subsequent extraction process. The invention mixes the recovered L-lactic acid fermentation thalli and lactate compound with feed raw materials in proportion for continuous fermentation, can continuously produce fermented feed, and greatly reduces the fermentation period of the feed. The fermented feed provided by the invention can effectively improve the growth rate of fattening livestock and reduce the meat-feed ratio and the diarrhea rate.

Drawings

FIG. 1 shows the results of the experiment in example 2 in which the test group and the control group were kept for 75 days.

FIG. 2 shows the results of measuring the microbial contents of the experimental groups in example 3.

FIG. 3 shows the results of measuring the microbial content of the control group in example 3.

Detailed Description

The invention is further illustrated by the following examples.

The L-lactic acid fermentation broth used in the following examples is a by-product in the production of L-lactic acid. The L-lactic acid fermentation liquor contains one or more of lactic acid, potassium lactate, sodium lactate, calcium lactate, magnesium lactate, ammonium lactate and ferrous lactate.

The elution waste liquid used in the following examples is obtained by eluting 1% -5% of sulfuric acid or hydrochloric acid on a cation exchange column in an ion exchange process in an L-lactic acid extraction process stage.

Example 1 recovery of L-lactic acid fermentation broth

The embodiment provides a method for recovering L-lactic acid fermentation broth thalli, which comprises the following steps:

(1) and (3) putting 20L of L-lactic acid fermentation liquor into a 50L size mixing tank, maintaining the constant temperature at 50 ℃, feeding the elution waste liquid at a constant speed while stirring, stably displaying the pH value to 4.5 by a pH electrode, and stopping feeding the elution waste liquid to obtain the L-lactic acid size mixing liquid.

(2) Adding the mixed slurry into a three-leg upper discharging centrifuge, adjusting the rotating speed, stopping centrifugation when the outflow volume of the clear liquid reaches 80% of the volume of the mixed slurry, and collecting the concentrated phase to obtain the L-lactic acid fermentation thallus concentrated solution.

The contents of lactic acid and Bacillus coagulans in the L-lactic acid tempered slurry, the concentrated phase and the clear solution were measured, respectively, and the results are shown in Table 1.

Table 1: experimental result for recovering L-lactic acid fermentation broth thalli filtered by centrifugal machine

	L-lactic acid (g/L)	Number of cells (cfu/g)
			L-lactic acid size mixing liquid	185.3	2×10⁹
Dense phase	193.2	1×10¹⁰
			Clear liquid	184.6	Not detected out

Example 2 preparation of fermented feed

This example provides a method of preparing a fermented feed comprising the steps of:

(1) and (3) in a 500L fermentation tank containing L-lactic acid fermentation liquor after L-lactic acid fermentation is finished, keeping the constant temperature at 50 ℃, feeding the elution waste liquid at a constant speed while stirring, stopping feeding the elution waste liquid when the pH value is stably displayed by a pH electrode to 4.5-5.0, and obtaining the L-lactic acid mixed liquor.

(2) Adding the L-lactic acid slurry regulating liquid into the ceramic membrane, adjusting the frequency of a circulating pump, controlling the pressure of an inlet and an outlet of the membrane core to make the flow rate of the clear liquid consistent with that of the slurry regulating liquid, adding 30% dialysis water when the flux of the clear liquid is obviously reduced, stopping filtering when the content of the L-lactic acid in the flowing clear liquid is less than 5%, and collecting a concentrated phase to obtain the L-lactic acid fermentation thallus concentrated solution.

The volumes of the conditioned L-lactic acid solution, the supernatant and the concentrated phase, the L-lactic acid content and the cell number were measured, respectively, and the results are shown in Table 2.

Table 2: experimental result for recovering L-lactic acid fermentation broth thalli by adopting ceramic membrane filtration

	L-lactic acid (g/L)	Number of cells (cfu/g)	Volume (L)
				L-lactic acid size mixing liquid	187.9	2×10⁹	412
Dense phase	47.5	3×10¹⁰	28
				Clear liquid	161.9	Not detected out	470

(3) Taking 3kg of concentrated phase and 297kg of feed raw materials (65% of corn flour, 12% of bran and 23% of soybean meal) to mix and match, and fermenting in a fermentation tank for 24 hours to obtain the seed fermented feed.

(4) 300kg of seed fermented feed and 2700kg of feed raw materials (65% of corn flour, 12% of bran and 23% of soybean meal) are mixed and proportioned, and fermented for 48 hours in a fermentation tank to obtain the fermented feed.

(5) Drying fermented feed, and packaging to obtain fermented feed containing Bacillus coagulans of 1 × 10⁶cfu/g, L-lactic acid content 102.5 mg/kg.

Example 3 continuous production of fermented feed

This example provides a method for continuous production of fermented feed comprising the steps of:

(1) 300m of L-lactic acid-containing fermentation broth at the end of L-lactic acid fermentation³And (3) in a fermentation tank, keeping the constant temperature at 50 ℃, feeding the elution waste liquid at a constant speed while stirring, stably displaying the pH value to 4.5-5.0 by a pH electrode, and stopping feeding the elution waste liquid to obtain the L-lactic acid size mixing liquid.

(2) And pumping a part of the slurry mixing liquid to a diaphragm type automatic discharging plate type filter press, and respectively collecting the filter press liquid and the filter cake.

(3) And mixing the filter cake and the other part of the mixed slurry according to the mass ratio of 1:1, mixing the mixed slurry with feed raw materials (65% of corn flour, 12% of bran and 23% of soybean meal) according to the mass ratio of 1:99, and fermenting the mixture in a fermentation tank for 24 hours to obtain the seed fermented feed.

(4) Mixing the seed fermented feed and feed raw materials (65% of corn flour, 12% of bran and 23% of soybean meal) according to a ratio of 1:9, and fermenting in a fermentation tank for 48h to obtain the fermented feed.

(5) Mixing the fermented feed and feed raw materials (65% of corn flour, 12% of bran and 23% of soybean meal) according to a mixing ratio of 3:7, and fermenting in a fermentation tank for 48h to continuously obtain the fermented feed.

(6) Drying fermented feed, and packaging to obtain fermented feed containing Bacillus coagulans of 6 × 10⁵cfu/g, L-lactic acid content 86.75 mg/kg.

(7) And (3) repeating the steps (1), (2) and (3) after each ten batches of fermented feed are continuously produced, and additionally adding 10% of seed fermented feed to obtain the continuous fermented feed.

Experimental example 1

Twenty-40 kg fattening pigs are fed for 75 days to 130kg for slaughter, the fermented feed (experimental group) and the feed raw materials (control group) of the fattening pigs in the example 2 are respectively fed for comparison, and the initial weight, slaughter weight and feed dosage of the fattening pigs are recorded and measured, and the result is shown in figure 1, wherein the slaughter weight p is less than 0.01, the weight gain p is less than 0.01, and the feed dosage p is less than 0.05.

Experimental example 2

Twenty-40 kg fattening pigs were fed for 75 days to 130kg slaughtered, and were fed with the fermented feed (experimental group) and the feed materials (control group) of example 3, respectively, and the initial weight, slaughtered weight, feed amount, and diarrhea rate of the fattening pigs were recorded and measured, and the results are shown in table 3. The microbial content was measured by taking a stool sample every 15 days, and the results are shown in fig. 2 and 3.

Table 3: the fermented feed has effect on growth of fattening pig

Note: wherein the weight p of the slaughtered animal is less than 0.01, and the feed dosage p is less than 0.05.

The results in table 3 show that compared with the control group, the weight gain of the fattening pig is improved by 12.02%, the meat-feed ratio is reduced by 3.86%, and the diarrhea rate is reduced by 56.48%.

The results of FIGS. 2 and 3 show that the Bacillus coagulans content of the experimental group was maintained at 10⁷-10⁸cfu/g, whereas the control group contained only 10% of Bacillus coagulans²-10³cfu/g. The tested group has obviously reduced Escherichia coli and salmonella, while the control group has no reduced Escherichia coli and salmonellaIs obvious.

Although the invention has been described in detail hereinabove by way of general description, specific embodiments and experiments, it will be apparent to those skilled in the art that many modifications and improvements can 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

1. A method for recovering L-lactic acid fermentation thalli and continuously producing fermented feed is characterized by comprising the following steps:

2. The method according to claim 1, wherein the L-lactic acid-fermenting bacteria is Bacillus coagulans.

3. The method according to claim 1, wherein the waste elution liquid from the ion exchange step is a waste elution liquid eluted with 1% -5% sulfuric acid or hydrochloric acid, and the L-lactic acid fermentation broth is slurried with the waste elution liquid to pH 4-6.

4. The method according to claim 1, wherein the solid-liquid separation mode is centrifugation, filter pressing or membrane filtration; preferably, the solid-liquid separation equipment is selected from one or more of a vacuum rotary drum filter, a disc type ultrafiltration centrifuge, a plate filter press and a ceramic membrane; more preferably, the solid-liquid separation equipment is a plate filter press or a ceramic membrane.

5. The method according to claim 1, characterized in that in the step (2), the mass of the concentrated phase is 0.5-5% of the mass of the feed raw materials during mixing;

preferably, the feed raw material contains 55-75% of corn flour, 8-16% of bran and 15-30% of soybean meal by mass of 100%;

preferably, the fermentation time is 24h-36 h.

6. The method according to claim 1, wherein in the step (3), the mass of the seed fermented feed is 10-45% of the mass of the feed raw materials during mixing;

preferably, the fermentation time is 48h-72 h;

more preferably, in the continuous fermentation process, when the content of the bacillus coagulans is less than or equal to 10⁵And (5) when cfu/g is reached, adding 10% of seed fermented feed to continuously produce the fermented feed.

7. The method according to claim 1, wherein the fermentation temperature is constant 40-60 ℃, preferably constant 50 ℃.

8. The method of any one of claims 1-7, wherein the L-lactic acid fermentation broth comprises one or more of lactic acid, potassium lactate, sodium lactate, calcium lactate, magnesium lactate, ammonium lactate, and ferrous lactate.

9. A fermented feed obtained by the method of any one of claims 1-8.

10. Use of the fermented feed according to claim 9 for increasing the growth rate, reducing the meat-to-feed ratio and/or reducing the diarrhea rate of fattening livestock.