CN112553069A - Solid state fermentation method based on combination of multiple technologies of pulse magnetic field, pulse intense light and near infrared spectrum online detection - Google Patents
Solid state fermentation method based on combination of multiple technologies of pulse magnetic field, pulse intense light and near infrared spectrum online detection Download PDFInfo
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- C12M21/00—Bioreactors or fermenters specially adapted for specific uses
- C12M21/16—Solid state fermenters, e.g. for koji production
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12J—VINEGAR; PREPARATION OR PURIFICATION THEREOF
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- C12M35/00—Means for application of stress for stimulating the growth of microorganisms or the generation of fermentation or metabolic products; Means for electroporation or cell fusion
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- C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
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- C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
- C12M41/48—Automatic or computerized control
Abstract
The invention belongs to the field of microbial fermentation engineering, and relates to a solid state fermentation method based on the combination of pulse magnetic field, pulse intense light and near infrared spectrum online detection technologies; firstly, inoculating fermented seed liquid or vinegar grains into a fermentation substrate to obtain a mixture, stirring the mixture, and then spreading the mixture on a solid fermentation bed, wherein the thickness of the mixture is 0.2-0.5 m; the fermentation substrate is a mixture of oil cake and water or a wine and bran mixture in an acetic acid fermentation stage in the brewing process of edible vinegar; after the solid state fermentation is started, applying a pulsed magnetic field with the intensity of 80-240 Gs to the mixture every 5-10 h, and treating for 1-2 h; applying pulsed strong light sterilization treatment to the surface of the fermentation substrate every 5-6 h, wherein the flash irradiation frequency is 2-5 times, the flash irradiation energy is 100-500J, and the flash irradiation distance is fixed to be 5-10 cm; the characteristic parameters of the fermentation substrate are monitored in real time by utilizing the near infrared spectrum on-line detection technology, and the automatic intelligent control of the belt type solid state fermentation is realized.
Description
Technical Field
The invention belongs to the field of microbial fermentation engineering, and particularly relates to a solid-state fermentation method based on-line detection of a pulsed magnetic field, pulsed intense light and near infrared spectrum.
Background
As shown in the "white paper 2015 of Chinese Industrial biotechnology", the industrial energy consumption of China is huge and the water consumption is about 13 hundred million m3The amount of wastewater per year is about 10 hundred million m3In one year, the problems of environmental pollution and resource waste caused by the method become bottleneck problems restricting sustainable development of the biological industry in China. Compared with liquid fermentation, the solid fermentation system has almost no free water, the fermentation process is simple, and the external environment close to the natural growth of microorganisms can be provided. With the increasing deterioration of resources and environmental problems, the solid-state fermentation production mode draws the attention of academia and industry, and is widely applied in the fields of starter propagation, wine brewing, cake feed fermentation and the like. The traditional solid state fermentation has the disadvantages of backward equipment, long fermentation period, easy pollution of mixed bacteria, difficult control of fermentation conditions and incapability of meeting the requirements of modern production. The magnetic field is used as a food non-thermal processing technology, and has the advantages of small heating temperature rise of raw materials, high processing speed, no need of medium transfer, capability of acting on materials, wide application range and the like. In recent years researchers have found that appropriate magnetic field parameters can promote the growth and metabolism of microorganisms. Therefore, the application of the pulsed magnetic field to the microorganism in the solid-state fermentation process is expected to improve the fermentation efficiency. In addition, most of solid state fermentation is open fermentation, the surface of a fermentation substrate is easy to pollute mixed bacteria, the quality of a fermentation product is reduced, and a proper fermentation substrate surface sterilization method for controlling the fermentation quality is not available at present. Hitherto, the monitoring of the fermentation process has mostly been carried out by time-division sampling and then by conventional chemical methodsThe method and the spectrum method are used for off-line analysis, the operation is complex, the time consumption is long, the determination cost is high, and the analysis result has hysteresis.
Therefore, there is a need to solve the problems of low fermentation efficiency, easy contamination of mixed bacteria, and inability of monitoring fermentation process and quality during solid-state fermentation.
Disclosure of Invention
The invention aims to provide a solid-state fermentation method based on the combination of pulse magnetic field, pulse intense light and near infrared spectrum on-line detection technologies, which can promote the solid-state fermentation process of microorganisms, avoid the pollution of mixed bacteria and realize the on-line real-time monitoring of the solid-state fermentation process; improving the solid state fermentation efficiency, reducing the production cost and ensuring the quality of the fermentation product.
In order to achieve the above purpose, the technical scheme adopted by the invention is as follows:
the solid state fermentation method based on the combination of multiple technologies of pulse magnetic field, pulse intense light and near infrared spectrum online detection comprises the following steps:
(1) firstly, inoculating fermentation seed liquid or vinegar grains with a proper proportion into a fermentation substrate to obtain a mixture, uniformly stirring the mixture, and then spreading the mixture on a belt type solid state fermentation bed to enable the thickness of a material layer of the mixture to be 0.2-0.5 m; the fermentation substrate is a mixture of oil cake and water or a wine and bran mixture in an acetic acid fermentation stage in the brewing process of edible vinegar;
(2) after the solid state fermentation is started, applying a magnetic field to the mixture once every 5-10 h, wherein the magnetic field intensity is 80-240 Gs of a pulse magnetic field, and the treatment time is 1-2 h;
(3) after the solid-state fermentation is started, applying pulsed strong light sterilization treatment to the surface of a fermentation substrate every 5-6 hours, wherein the flash irradiation frequency is 2-5 times, the flash irradiation energy is 100-500J, and the flash irradiation distance is fixed to be 5-10 cm;
(4) the characteristic parameters of the fermentation substrate are monitored in real time by utilizing the near infrared spectrum on-line detection technology, and the automatic intelligent control of the belt type solid state fermentation is realized.
Preferably, the oil cake meal in the step (1) is any one or more of soybean meal, rapeseed meal, sunflower seed meal or peanut meal; the dosage ratio of the oil cake dregs to the water is 1g (1-2.0) mL.
Preferably, the wine-bran mixture in the step (1) is mixed according to the mass ratio of the bran to the fermented grains of 1:2 (w/w).
Preferably, the fermentation seed liquid in the step (1) is any one or more of bacillus subtilis, bacillus licheniformis and bacillus natto seed liquid.
Preferably, in the step (1), when the fermentation substrate is a wine-bran mixture, the fermentation substrate is correspondingly inoculated with vinegar mash; the mass ratio of the vinegar grains to the fermentation substrate is 1:100 (w/w).
Preferably, in the step (1), when the fermentation substrate is a mixture of oil cake and water, the fermentation substrate is correspondingly inoculated with fermentation seed liquid; the volume mass ratio of the fermentation seed liquid to the fermentation substrate is 5-10% (v/w).
The fermentation of the wine-bran mixture in the brewing process of the edible vinegar is to provide acetic acid bacteria for fermentation by utilizing mature vinegar mash.
Preferably, in the step (2), the pulsed magnetic field with the magnetic field intensity of 240Gs is processed for 1 h.
Preferably, in the step (2), the flash irradiation frequency is 5 times, the flash irradiation energy is 500J, and the flash irradiation distance is fixed at 10 cm.
Preferably, in step (4), the characteristic parameters of the fermentation substrate include pH, polypeptide content, acetic acid content or microbial biomass.
The invention has the following remarkable advantages:
(1) the invention applies the combination of multiple technologies of pulsed magnetic field enhanced fermentation, sterilization of the surface of a fermentation substrate by pulsed strong light and near infrared spectrum on-line detection to the microbial solid state fermentation process, and the invention is not reported in the open at home and abroad at present; the intensive pulse light sterilization is characterized in that strong white light flash of pulses is utilized to enable an inert gas lamp to emit light from ultraviolet rays to infrared rays which are similar to the solar spectrum but have stronger intensity so as to inhibit the growth and the propagation of microorganisms on the surfaces of food and packaging materials, solid surfaces, gases and transparent beverages, and the intensive pulse light sterilization is a novel safe, powerful and energy-saving cold sterilization technology; compared with other non-thermal sterilization technologies, the pulse highlight sterilization has the advantages of convenience in operation, high sterilization efficiency, capability of keeping food quality better and the like; meanwhile, a monitoring model is established by combining near-infrared real-time spectrum with a chemometrics method, so that online automatic intelligent control of the fermentation process can be realized
(2) The belt-type solid-state fermentation device is adopted, so that the space occupied by solid-state fermentation is saved; in addition, the microbial solid state fermentation period is long, the production efficiency is low, and the pulsed magnetic field is applied to the solid state fermentation process to stimulate the growth and metabolism of the microbes and improve the efficiency of the solid state fermentation.
(3) The solid state fermentation is mostly open type fermentation, the surface of the fermentation substrate is easy to pollute mixed bacteria, and the quality of the fermentation product is reduced.
(4) At present, no method for monitoring the solid-state fermentation process of microorganisms exists in actual production, the invention applies the near infrared spectrum on-line detection technology to the solid-state fermentation process, and realizes automatic intelligent control of the solid-state fermentation process by establishing a near infrared spectrum monitoring model of characteristic parameters of fermentation substrates.
(5) Through a contrast test, a pulsed magnetic field and pulsed intense light treatment are applied in the solid-state fermentation process of the oil cake dregs, and the polypeptide content of the fermented cake dregs is increased by 11.5-17.6% compared with that of an untreated group; the fermentation product has no mixed bacteria pollution; can establish a Si-PLS quantitative model for detecting the polypeptide content, the pH value and the microbial biomass of the cake and the near infrared spectrum. In the process of brewing the edible vinegar, a pulsed magnetic field and pulsed intense light treatment are applied to a fermentation substrate obtained after adding mature vinegar grains into a wine-bran mixture in an acetic acid fermentation stage, so that the yield of the acetic acid is increased by 4.5-8.3%; the fermentation product has no mixed bacteria pollution; can establish a Si-PLS quantitative model for detecting the content of acetic acid in the matrix and the near infrared spectrum.
Drawings
FIG. 1 is a schematic diagram of a multi-technology combined device for on-line detection of pulsed magnetic field, pulsed intense light and near infrared spectrum in the solid-state fermentation process of the invention; wherein 1-coil; 2, conveying a belt; 3-fermentation substrate; 4-pulsed strong light irradiation board; 5-near infrared spectrum probe; 6-signal acquisition system.
Detailed Description
The present invention will be further illustrated with reference to the following examples, but the present invention is not limited thereto.
FIG. 1 is a schematic diagram of a multi-technology combined device for online detection of pulsed magnetic field, pulsed intense light and near infrared spectrum in the solid-state fermentation process of the invention; the pulse magnetic field generating device consists of a coil 1 and is positioned below the fermentation substrate 3; meanwhile, a pulse strong light irradiation plate 4 is arranged above the fermentation substrate 3; a near infrared spectrum probe 2 of the near infrared spectrometer extends into the fermentation substrate 3 for detection, and meanwhile, the near infrared spectrometer is electrically connected with a signal acquisition system 6, and the signal acquisition system 6 acquires signals and performs information feedback control; furthermore, a conveyor belt 2 is provided for transporting the fermentation substrate 3.
Example 1:
(1) mixing the soybean meal and water according to the mass-volume ratio of 1g to 1.0mL, and mixing the soybean meal and the water according to the weight-volume ratio of the seed liquid: inoculating the bacillus subtilis seed solution into the mixture according to the volume mass ratio of (cake meal and water) of 5% (v/w), uniformly stirring, and then spreading on a belt type solid state fermentation bed, wherein the thickness of a material layer is 0.2 m;
(2) after the solid state fermentation is started, applying a pulse magnetic field with the magnetic field intensity of 80Gs for 1h every 5 h;
(3) after the solid-state fermentation is started, applying pulsed strong light sterilization treatment to the surface of the fermentation substrate every 5h, wherein the flash irradiation frequency is 2 times, the flash irradiation energy is 100J, and the flash irradiation distance is 5 cm;
(4) and monitoring the pH value change of the fermentation substrate by using a near infrared spectrum online detection technology. Through determination, the content of the polypeptide is increased by 11.5 percent compared with that of a group which is not applied with a pulse magnetic field; the flora analysis result shows that the fermented cake is free from mixed bacteria pollution; Si-PLS is adopted to establish a quantitative model for online pH detection by near infrared spectroscopy to obtain a pH correction and prediction model RcAnd RpAnd RMSECV and RMSEP are 0.9716 and 0.9616 and 1.872 and 1.911, respectively.
Example 2:
(1) mixing rapeseed dregs and water according to the mass-volume ratio of 1g to 2.0mL, and mixing the rapeseed dregs and the water according to the ratio of seed liquid: inoculating the bacillus licheniformis seed solution into the mixture according to the volume mass ratio of (cake meal and water) of 10% (v/w), uniformly stirring, and spreading on a belt type solid state fermentation bed, wherein the thickness of a material layer is 0.5 m;
(2) after the solid state fermentation is started, applying pulsed magnetic field treatment with the magnetic field intensity of 240Gs every 10h for 1 h;
(3) after the solid-state fermentation is started, applying strong pulse light sterilization treatment to the surface of the fermentation substrate every 5h, wherein the flash irradiation frequency is 5 times, the flash irradiation energy is 500J, and the flash irradiation distance is 10 cm;
(4) and monitoring the polypeptide content change in the fermentation substrate by using a near infrared spectrum online detection technology. Through determination, the content of the polypeptide is improved by 17.6 percent compared with that of a group which is not applied with a pulse magnetic field; the flora analysis result shows that the fermented cake is free from mixed bacteria pollution; Si-PLS is adopted to establish a quantitative model for detecting the content of the polypeptide on line by near infrared spectroscopy, and a correction and prediction model R of the content of the polypeptide is obtainedcAnd RpAnd RMSECV and RMSEP are 0.9494 and 0.9570 and 2.452 and 2.061, respectively.
Example 3:
(1) mixing sunflower seed meal and water according to a mass-volume ratio of 1g: 1.5mL, and mixing according to the ratio of seed liquid: inoculating the bacillus natto seed liquid into the mixture according to the volume mass ratio of (cake meal and water) of 8% (v/w), uniformly stirring, and spreading on a belt type solid state fermentation bed, wherein the thickness of a material layer is 0.35 m;
(2) after the solid state fermentation is started, applying pulse magnetic field treatment with the magnetic field intensity of 160Gs every 8h for 2 h;
(3) after the solid-state fermentation is started, applying pulsed strong light sterilization treatment to the surface of the fermentation substrate every 6h, wherein the flash irradiation frequency is 4 times, the flash irradiation energy is 300J, and the flash irradiation distance is 7.5 cm;
(4) and monitoring the change of the microbial biomass in the fermentation substrate by using a near infrared spectrum online detection technology. Through determination, the content of the polypeptide is improved by 15.7 percent compared with that of a group which is not applied with a pulse magnetic field; the flora analysis result shows that the fermented cake is free from mixed bacteria pollution; method for establishing near infrared spectrum on-line detection of bacillus natto biomass by adopting Si-PLSQuantitative model, correction and prediction model R for obtaining biomasscAnd RpAnd RMSECV and RMSEP are 0.9402 and 0.9521 and 1.639 and 1.618, respectively.
Example 4:
(1) adding mature vinegar grains into a wine-bran mixture (bran and wine grains are mixed according to the mass ratio of 1:2 (w/w)) in an acetic acid fermentation stage in the brewing process of edible vinegar, wherein the mass ratio of the vinegar grains to the wine-bran mixture is 1:100(w/w), uniformly stirring, and then spreading on a belt-type solid fermentation bed, wherein the thickness of a material layer is 0.2 m;
(2) after the solid state fermentation is started, applying a pulse magnetic field with the magnetic field intensity of 80Gs for treatment for 1h every 5 h;
(3) after the solid-state fermentation is started, applying pulsed strong light sterilization treatment to the surface of the fermentation substrate every 5h, wherein the flash irradiation frequency is 2 times, the flash irradiation energy is 100J, and the flash irradiation distance is 5 cm;
(4) and monitoring the change of the content of acetic acid in the fermentation substrate by using a near infrared spectrum online detection technology. Through determination, the content of acetic acid is improved by 4.5 percent compared with a group which is not applied with a pulse magnetic field; the flora analysis result shows that the fermentation product has no mixed bacteria pollution; Si-PLS is adopted to establish a quantitative model for online pH detection by near infrared spectroscopy to obtain a pH correction and prediction model RcAnd RpAnd RMSECV and RMSEP are 0.9513 and 0.9624 and 2.864 and 3.015, respectively.
Example 5:
(1) adding mature vinegar grains into a wine-bran mixture (bran and wine grains are mixed according to the mass ratio of 1:2 (w/w)) in an acetic acid fermentation stage in the brewing process of edible vinegar, wherein the mass ratio of the vinegar grains to the wine-bran mixture is 1:100(w/w), uniformly stirring, and then spreading on a belt-type solid fermentation bed, wherein the thickness of a material layer is 0.5 m;
(2) after fermentation is carried out for 10 hours, a pulsed magnetic field with the magnetic field intensity of 240Gs is applied for treatment for 1 hour;
(3) after the solid-state fermentation is started, applying strong pulse light sterilization treatment to the surface of the fermentation substrate every 5h, wherein the flash irradiation frequency is 5 times, the flash irradiation energy is 500J, and the flash irradiation distance is 10 cm;
(4) using near infrared spectroscopyThe change in acetic acid content in the fermentation substrate was monitored by a line detection technique. Through determination, the content of acetic acid is improved by 8.3 percent compared with a group which is not applied with a pulse magnetic field; the flora analysis result shows that the fermentation product has no mixed bacteria pollution; establishing a quantitative model for online detection of acetic acid content by near infrared spectroscopy by using Si-PLS to obtain a correction and prediction model R of the acetic acid contentcAnd RpAnd RMSECV and RMSEP are 0.9711 and 0.9584 and 3.061 and 3.184, respectively.
Example 6:
(1) adding mature vinegar grains into a wine-bran mixture (bran and wine grains are mixed according to the mass ratio of 1:2 (w/w)) in an acetic acid fermentation stage in the brewing process of edible vinegar, wherein the mass ratio of the vinegar grains to the wine-bran mixture is 1:100(w/w), uniformly stirring, and then spreading on a belt-type solid fermentation bed, wherein the thickness of a material layer is 0.35 m;
(2) after the solid state fermentation is started, applying pulse magnetic field treatment with the magnetic field intensity of 160Gs every 8h for 2 h;
(3) after the solid-state fermentation is started, applying pulsed strong light sterilization treatment to the surface of the fermentation substrate every 6h, wherein the flash irradiation frequency is 4 times, the flash irradiation energy is 300J, and the flash irradiation distance is 7.5 cm;
(4) and monitoring the change of the acetic acid content in the fermentation substrate by using a near infrared spectrum online detection technology. Through determination, the content of acetic acid is improved by 7.2 percent compared with that of a group which is not applied with a pulse magnetic field; the flora analysis result shows that the fermentation product has no mixed bacteria pollution; establishing a quantitative model for online detection of acetic acid bacteria biomass by near infrared spectroscopy by adopting Si-PLS to obtain a biomass correction and prediction model RcAnd RpAnd RMSECV and RMSEP are 0.9399 and 0.9578 and 2.883 and 2.965, respectively.
Description of the drawings: the above embodiments are only used to illustrate the present invention and do not limit the technical solutions described in the present invention; thus, while the present invention has been described in detail with reference to the foregoing embodiments, it will be understood by those skilled in the art that various changes may be made and equivalents may be substituted; all such modifications and variations are intended to be included herein within the scope of this disclosure and the present invention and protected by the following claims.
Claims (9)
1. A solid state fermentation method based on-line detection of multiple technologies of a pulse magnetic field, pulse intense light and near infrared spectrum is characterized by comprising the following steps:
(1) firstly, inoculating fermentation seed liquid or vinegar grains with a proper proportion into a fermentation substrate to obtain a mixture, uniformly stirring the mixture, and then spreading the mixture on a belt type solid state fermentation bed to enable the thickness of a material layer of the mixture to be 0.2-0.5 m; the fermentation substrate is a mixture of oil cake and water or a wine and bran mixture in an acetic acid fermentation stage in the brewing process of edible vinegar;
(2) after the solid state fermentation is started, applying a magnetic field to the mixture once every 5-10 h, wherein the magnetic field intensity is 80-240 Gs of a pulse magnetic field, and the treatment time is 1-2 h;
(3) after the solid-state fermentation is started, applying pulsed strong light sterilization treatment to the surface of a fermentation substrate every 5-6 hours, wherein the flash irradiation frequency is 2-5 times, the flash irradiation energy is 100-500J, and the flash irradiation distance is fixed to be 5-10 cm;
(4) the characteristic parameters of the fermentation substrate are monitored in real time by utilizing the near infrared spectrum on-line detection technology, and the automatic intelligent control of the belt type solid state fermentation is realized.
2. The solid-state fermentation method based on the combination of the pulsed magnetic field, the pulsed intense light and the near infrared spectrum for on-line detection of the multiple technologies according to claim 1, wherein the oil cake meal in the step (1) is any one or more of bean meal, rapeseed meal, sunflower seed meal or peanut meal; the dosage ratio of the oil cake dregs to the water is 1g (1-2.0) mL.
3. The solid-state fermentation method based on the combination of the pulsed magnetic field, the pulsed intense light and the near infrared spectrum online detection technologies as claimed in claim 1, wherein the wine-bran mixture in the step (1) is mixed according to a mass ratio of bran to fermented grains of 1: 2.
4. The solid-state fermentation method based on the combination of the pulsed magnetic field, the pulsed intense light and the near infrared spectrum for on-line detection of the multiple technologies as claimed in claim 1, wherein the fermented seed liquid in the step (1) is any one or more of bacillus subtilis, bacillus licheniformis and bacillus natto seed liquids.
5. The solid-state fermentation method based on the combination of the pulsed magnetic field, the pulsed intense light and the near infrared spectrum for on-line detection of the multiple technologies as claimed in claim 1, wherein in the step (1), when the fermentation substrate is a wine-bran mixture, the fermentation substrate is correspondingly inoculated with vinegar grains; the mass ratio of the vinegar grains to the fermentation substrate is 1: 100.
6. The solid state fermentation method based on the combination of the pulsed magnetic field, the pulsed intense light and the near infrared spectrum for on-line detection of the multiple technologies as claimed in claim 1, wherein in the step (1), when the fermentation substrate is a mixture of the oilseed residue and water, the fermentation substrate is correspondingly inoculated with fermentation seed liquid; the volume mass ratio of the fermentation seed liquid to the fermentation substrate is 5-10%.
7. The solid state fermentation method based on the combination of pulsed magnetic field, pulsed intense light and near infrared spectrum online detection technologies as claimed in claim 1, wherein in step (2), the pulsed magnetic field with the magnetic field intensity of 240Gs is processed for 1 h.
8. The solid state fermentation method based on the combination of pulsed magnetic field, pulsed intense light and near infrared spectroscopy for on-line detection of multiple technologies as claimed in claim 1, wherein in step (2), the number of flash irradiation is 5, the flash irradiation energy is 500J, and the flash irradiation distance is fixed at 10 cm.
9. The solid state fermentation method based on the combination of pulsed magnetic field, pulsed intense light and near infrared spectroscopy for on-line detection of multiple technologies as claimed in claim 1, wherein in step (4), the characteristic parameters of the fermentation substrate include pH, polypeptide content, acetic acid content or microbial biomass.
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