CN103698541A - Online glutamic acid fermentation detection system and online detection method using system - Google Patents
Online glutamic acid fermentation detection system and online detection method using system Download PDFInfo
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Abstract
The invention discloses an online glutamic acid fermentation detection system and an online detection method using the system, which is used for detecting the concentrations of glucose and glutamic acid on line during glutamic acid fermentation so as to solve the problems of disturbance caused due to time-varying characteristics, relevance and hysteresis of biochemical parameters and inevitable manual interference in a fermentation process of glutamic acid and low reliability in implementing a plurality of mathematical models which are generated with relevance to substrate consumption, bacteria growth and product generation. The online detection method for the concentrations of the glucose and the glutamic acid is finished under automatic control of programs completely; manual sampling is not required; meanwhile, the concentrations of the glucose and the glutamic acid can be simultaneously detected and data can be input to a computer to be saved; the glutamic acid fermentation dynamics is researched according to the concentrations of the glucose and the glutamic acid in different fermentation stages, so that the fermentation process is well controlled.
Description
Technical field
The present invention relates to glutamic acid fermentation technical field, particularly a kind of glutamic acid fermentation on-line detecting system and adopt the online method detecting of this system.Glutamic acid fermentation process biomass online test method of the present invention, can robotization complete the detection to multiple biological parameter, has avoided human error to simplify mensuration operation; And simultaneously detection substrate, target product.
Background technology
The object of fermentation is to accumulate in a large number the needed microbial metabolic products of people; The manual control of metabolism is to break artificially the Metabolism control system of microorganism, and the direction that metabolism is wished towards people is carried out.
Biological manufacture is to utilize the biocatalytic Activity of cell or enzyme to carry out large-scale material processing and the advanced production mode transforming, be the hi-tech industry based on modern biotechnology development, be widely used in many key areas such as food, medicine, feed, papermaking, leather, weaving, chemical industry, daily use chemicals, the energy and environmental protection
Fermentation dynamics is one of theoretical foundation of biological respinse engineering science, is the science of the interaction change with time between the various environmental factors of research and cell (microorganism) metabolic activity.The mutual relationship, environmental factor that particular content has growth rate, substrate consumption speed and a product formation rate of thalline in the balance, sweat of quality in microorganism growth process is on three's impact and the factor that affects reaction rate.Reinforcement is to the dynamic (dynamical) research of sweat, can be simulation, optimization and the control based theoretical of industrial fermentation, to deep understanding and grasp sweat, transform and promote traditional mode of production, realize efficient, low consumption that large scale fermentation is produced, the international competitiveness that improves industry has important effect.
In recent years, adopt widely people's concern of optimization of computer-controlled fed batch fermentation process.It is larger that yet the on-line optimization of realizing sweat is controlled difficulty, main cause is: (1) sweat is dynamic non-linear process, (2) lack accurate process model, (3) model parameter is variable, (4) lack reliable biology sensor, make some important state variables as cell concentration, production concentration, substrate concentration cannot on-line measurement.For the Main Biochemical that characterizes sweat state, as glucose acid invert ratio, acid production rate, residual sugar etc., substantially by manually sampling and analyze to laboratory more at the scene, and disposable only to analyze a kind of biochemical parameter.The disturbances such as the time variation of model parameter, correlativity, hysteresis quality and inevitable human intervention, the reliability that some mathematical models that generate about base consumption, thalli growth, product are put into practice is less, and the control model that causes much fermenting does not reach the effect of expection in actual production.
The Computer Control Technology of sweat is the inevitable approach that fermentation industry changes the mode of production, enhances productivity from now on, and the basis of realizing this goal is exactly to set up mathematical model accurately.According to current achievement in research, aspect model sets up, lack online measuring technique and the method for biological (biochemistry) parameter of sweat, this has become one of key issue of restriction Computer Control Technology widespread use.
Summary of the invention
The object of the present invention is to provide a kind of glutamic acid fermentation on-line detecting system and adopt the online method detecting of this system, to solve in glutamic acid fermentation process time variation due to biochemical parameter, correlativity, hysteresis quality and the inevitably disturbance such as human intervention, and the less problem of reliability that puts into practice of some mathematical models generating about base consumption, thalli growth, product.
Technical scheme of the present invention is achieved in that
A glutamic acid fermentation on-line detecting system, comprises sampling pump, and described sampling pump inlet end has fermentation tank by pipeline communication, and described sampling pump endpiece has sampling pool by pipeline communication, and there is waste liquid drain pump described sampling pool bottom by pipeline communication;
Diluting tank, be arranged on described sampling pool right side and with described sampling pool on same straight line, as the mixing pit of dilution and sample, its bottom is by waste liquid drain pump inlet end described in pipeline communication, and the endpiece of described waste liquid drain pump is by pipeline communication waste liquid pool;
Dilution fillup valve, is fixedly connected with mechanical arm, for injecting dilution to described diluting tank;
Biology sensor reaction tank, is arranged on described diluting tank right side, and its bottom is by the pipeline communication pond scavenging pump endpiece that responds, and described reaction tank scavenging pump inlet end has damping fluid storage tank by pipeline communication; Its bottom is by the respond inlet end of pond drain pump of another pipeline communication, and the endpiece of described reaction tank drain pump is by waste liquid pool described in pipeline communication; In described biology sensor reaction tank, be provided with post glucose oxidase enzyme membrane the first Catalase, post the second Catalase of dglutamic oxidase enzyme membrane; Described the first Catalase and the second Catalase are electrically connected with ADS converting system, and described ADS converting system connects the control system of computing machine;
Standard specimen pond, is arranged between described diluting tank and described sampling pool, demarcates the standard solution of described the first Catalase and described the second Catalase for splendid attire;
Sampling valve, is fixedly connected with described mechanical arm, for the sample that extracts described sampling pool, diluting tank or standard specimen pond to described biology sensor reaction tank;
Control system, is electrically connected to described sampling pump, controls sampling interval, the sampling quantity of described sampling pump; Be electrically connected to described mechanical arm, control the direction of motion of described mechanical arm; Be electrically connected to described dilution fillup valve, control dilution injection rate IR and injection phase; Be electrically connected to described sampling valve, control sampling location, sampling quantity and the injection phase of described sampling valve; Be electrically connected to and control described sampling pump, reaction tank drain pump, waste liquid drain pump, reaction tank scavenging pump.
Preferably, also comprise colorimetric diluting tank, described colorimetric diluting tank comprises the pond body of splendid attire sample, described pond body sidewall is oppositely arranged light source emitter and light source absorption plant, described light source is 570nm light source, the sample of described light source in the body of described pond be laggard enters light source absorption plant, by the OD value of light source absorption plant test sample and light absorption signal is transmitted by ADS converter and be stored to computing machine; Body bottom, described pond is by the entrance of waste liquid drain pump described in pipeline communication.
Further, described biology sensor reaction tank side wall upper part is communicated with level control (LC) pipe, and the described level control (LC) pipe other end is communicated with described waste liquid pool.
Further, described sampling pump, waste liquid drain pump, reaction tank drain pump and reaction tank scavenging pump are peristaltic pump.
Further, described diluting tank, colorimetric diluting tank and/or biology sensor reaction tank bottom are provided with rabbling mechanism, and described rabbling mechanism is controlled by described control system.
Adopt described glutamic acid fermentation on-line detecting system to detect online the method for glutamic acid and concentration of glucose, it is characterized in that, comprise the following steps:
1) calibration: described control system is controlled described sampling valve and extracted 10-60uL standard solution to described biology sensor reaction tank by described standard specimen pond, simultaneously by described reaction tank scavenging pump, in described bio-sensing reaction tank, pump into quantitative buffer solution, measure the concentration of glucose and glutamic acid in described standard solution; METHOD FOR CONTINUOUS DETERMINATION at least twice, double measuring result error is less than setting value, and calibration is passed through;
2) OD value test: it is 7-255min that sample interval is set, described control system extracts fermentation liquor in described fermentation tank to described sampling pool according to sampling pump described in the time interval controls arranging; Described sampling valve extracts described in 10-60 μ L fermentation liquor to described colorimetric diluting tank by described sampling pool, then described dilution fillup valve extraction dilution quantitatively dilutes described fermentation liquor to described colorimetric diluting tank, and described fermentation liquor OD value directly shows on computers;
3) sample test: the sample solution that described sampling valve extracts 10-60 μ L by described colorimetric diluting tank is to described diluting tank; Described control system is controlled described dilution fillup valve and inject dilution in described diluting tank, according to extension rate in advance, further the fermentation liquor in described colorimetric diluting tank is quantitatively diluted, and described extension rate is 0-500 times; After dilution, sample solution after described sampling valve is diluted by described diluting tank extraction is to described biology sensor reaction tank, by described reaction tank scavenging pump, in described biology sensor reaction tank, pump into quantitative buffer solution, the concentration of test glucose and glutamic acid is also stored to computing machine simultaneously;
Be completed at every turn, described waste liquid drain pump and reaction tank drain pump automatically start under the control of described control system, solution in described sampling pool, colorimetric diluting tank, diluting tank and/or biology sensor reaction tank is evacuated to waste liquid pool, starts described sampling pump simultaneously the solution in described sampling pool is evacuated to described waste liquid pool;
4) clean: after being completed, described reaction tank scavenging pump extracts damping fluid to described biology sensor reaction tank, to clean described biology sensor reaction tank, after cleaning, by described reaction tank drain pump, described damping fluid is evacuated to described waste liquid pool.
Preferably, described dilution is the mixed aqueous solution of Sodium Benzoate and EDETATE SODIUM, and the mass concentration of described Sodium Benzoate is 2%, and the mass concentration of described EDETATE SODIUM is 1%.
Preferably, described buffer solution is phosphate buffer solution.
Preferably, in step 1), described twice measuring result error range of set value is 0.1-4%.
Preferably, step 2) in, the quantitative dilution of described fermentation liquor is to 10 times of fermentation liquor dilutions.
To described biology sensor reaction tank, pump into damping fluid, for regulating the pH value of biology sensor reaction tank, the pH in biology sensor reaction tank is maintained near glucose oxidase and the optimum pH of dglutamic oxidase on the one hand; On the other hand, damping fluid plays medium effect, because only extracting 10-60 μ L sample solution, tests to described biology sensor reaction tank by sampling valve, so few solution does not reach described the first Catalase and the second Catalase, cannot test, adopt buffer solution as medium, make described the first Catalase and the second Catalase by solution submergence, be convenient to test.
Adopt the principle of the first Catalase and the second Catalase test concentration of glucose and aminoglutaric acid concentration to be: the substrate in fermentation liquor and corresponding enzyme generation biochemical reaction, the H that enzyme reaction is emitted
2o
2the internal layer that sees through enzyme membrane loses electron reduction on platinum electrode surface and becomes H
2o, platinum electrode receiving and losing electrons generation current signal, this current signal and H
2o
2concentration proportional, and H
2o
2concentration and concentration of substrate are linearly proportional, by the size of the electric signal that relatively measured object and standard model produce, just can calculate the content of substrate in sample, then through ADS converting system, deposit this information in computer stored.
Beneficial effect of the present invention is:
Glutamic acid on-line detecting system of the present invention is simple in structure, this detection system of use of the present invention detects online the method for glucose and aminoglutaric acid concentration in glutamic acid fermentation process and by procedure auto-control, is completed completely, do not need hand sampling, and the inventive method can record the concentration of glutamic acid and glucose simultaneously and data be inputted to computing machine and preserve, according to different fermentations stage concentration of glucose and aminoglutaric acid concentration research glutamic acid fermentation dynamics, better controlled fermentation process.
Accompanying drawing explanation
Fig. 1 is the structural representation of glutamic acid fermentation on-line detecting system of the present invention;
Fig. 2 is control system and each parts annexation block diagram in glutamic acid on-line detecting system of the present invention.
Embodiment
Below the technical scheme in the embodiment of the present invention is clearly and completely described, obviously, described embodiment is only a part of embodiment of the present invention, rather than whole embodiment.Embodiment based in the present invention, those of ordinary skills, not making the every other embodiment obtaining under creative work prerequisite, belong to the scope of protection of the invention.
As shown in Figure 1, a kind of glutamic acid fermentation on-line detecting system, comprises sampling pool 4, colorimetric diluting tank 5, standard specimen pond 6, diluting tank 7 and the biology sensor reaction tank 8 of linearly arranging successively from left to right.Upper space at sampling pool 4, colorimetric diluting tank 5, standard specimen pond 6, diluting tank 7 and biology sensor reaction tank 8 arranges mechanical arm 1, is fixedly connected with dilution fillup valve 2 and sampling valve 3 on mechanical arm.Dilution fillup valve 2 injects colorimetric diluting tank 5 or diluting tank 7 by the dilution extracting under the drive of mechanical arm 1.Sampling valve 3 extracts sample in sampling pool 4, colorimetric diluting tank 5, standard specimen pond 6 or diluting tank 7 to biology sensor reaction tank 8 under the drive of mechanical arm.
This glutamic acid fermentation on-line detecting system, by sampling pump 14, in fermentation tank, sample, sampling pump 14 inlet ends are by pipeline communication fermentation tank, and sampling pump 14 endpiece have sampling pool 4 by pipeline communication, and there is waste liquid drain pump 15 sampling pool 4 bottoms by pipeline communication.
When sample concentration is larger, need to dilute sample, diluting tank 7 is as the mixing pit of dilution and sample.Diluting tank 7 bottoms are by pipeline communication waste liquid drain pump 15 inlet ends, and the endpiece of waste liquid drain pump 15 is by pipeline communication waste liquid pool 19.
The bottom of biology sensor reaction tank 8 is by the respond endpiece of pond scavenging pump 17 of pipeline communication, and reaction tank scavenging pump 17 inlet ends have damping fluid storage tank 18 by pipeline communication; Biology sensor reaction tank 8 bottoms are by the respond inlet end of pond drain pump 16 of another pipeline communication, and the endpiece of reaction tank drain pump 16 is by pipeline communication waste liquid pool; In biology sensor reaction tank 8, be provided with post glucose oxidase enzyme membrane the first Catalase 9, post the second Catalase 10 of dglutamic oxidase enzyme membrane; The first Catalase 9 and the second Catalase 10 are electrically connected with ADS converting system, and ADS converting system connects the control system of computing machine.In addition, biology sensor reaction tank 8 side wall upper part are communicated with level control (LC) pipe 22, and level control (LC) pipe 22 other ends are communicated with waste liquid pool 19.
As shown in Figure 2, the control system in glutamic acid fermentation on-line detecting system of the present invention, is electrically connected to sampling pump 14, controls sampling interval, the sampling quantity of sampling pump 14; Be electrically connected to mechanical arm 1, control the direction of motion of mechanical arm 1; Be electrically connected to dilution fillup valve 2, control dilution injection rate IR and injection phase; Be electrically connected to sampling valve 3, control sampling location, sampling quantity and the injection phase of sampling valve; Be electrically connected to and control the first stirring machine 11, the second stirring machine 12, the 3rd stirring machine 13; Be electrically connected to and control the startup of reaction tank drain pump 16, waste liquid drain pump 15, reaction tank scavenging pump 17 and close.
With specific embodiment, further illustrate the present invention below:
Preparatory work of experiment:
1, seed culture: preparation seed culture medium (sugar 0.25%, dipotassium hydrogen phosphate 0.02%, magnesium sulfate 0.01%, urea 0.05%, corn steep liquor 0.03%), (120 ℃ of high-pressure steam sterilizing pans, sterilizing 20min), the corynebacterium glutamicum of cooling good access activation belongs to, and is placed on and on shaking table, cultivates 8h;
2, expand and cultivate: in fermentation tank, prepare fermentation medium (sugar 0.55%, dipotassium hydrogen phosphate 0.02%, magnesium sulfate 0.01%, urea 0.05%, corn steep liquor 0.05%), to and in fermentation medium, insert pH electrode and dissolved oxygen electrode, proofread and correct after ph electrode, the pipe outlets that fermentation tank is all tie, high-pressure sterilizing pot (120 ℃, 20min) in sterilizing, (ventilation is to ventilate in fermentation tank in cooling rear ventilation water flowing, guaranteeing has pressure in fermentation tank, prevents microbiological contamination; Water flowing is the recirculated water in fermentation tank bottom, because fermentation will be controlled temperature, when excess Temperature, recirculated water is connected automatically, is used for cooling), proofread and correct dissolved oxygen electrode, then carry out sterile working, the seed access fermentation tank by cultivating in shaking flask, expands and cultivates.Wherein 32 ℃ of fermentation temperatures, dissolved oxygen be greater than 15%, the stirring rate 250r/min of pH7.2, fermentation tank, these parameters also can be controlled by computer control system.
The glutamic acid fermentation on-line detecting system adopting detects the method for glutamic acid and concentration of glucose online, comprises the following steps:
1) calibration: control system is controlled sampling valve and extracted 10-60uL standard solution to biology sensor reaction tank by standard specimen pond, by reaction tank scavenging pump, in bio-sensing reaction tank, pump into quantitative buffer solution, the concentration of bioassay standard glucose in solutions and glutamic acid simultaneously; METHOD FOR CONTINUOUS DETERMINATION at least twice, double measuring result error is less than setting value, and calibration is passed through;
2) OD value test: it is 7-255min that sample interval is set, control system extracts fermentation liquor in fermentation tank to sampling pool according to the time interval controls sampling pump arranging; Sampling valve extracts 10-60 μ L fermentation liquor to colorimetric diluting tank by sampling pool, and then dilution fillup valve extraction dilution quantitatively dilutes fermentation liquor to colorimetric diluting tank, and fermentation liquor OD value directly shows on computers;
3) sample test: the sample solution that sampling valve extracts 10-60 μ L by colorimetric diluting tank is to diluting tank; Control system is controlled dilution fillup valve and inject dilution in diluting tank, according to the fermentation liquor in the further contrast colors diluting tank of extension rate in advance, quantitatively dilutes, and extension rate is 0-500 times; After dilution, sample solution after sampling valve is diluted by diluting tank extraction is to biology sensor reaction tank, by reaction tank scavenging pump, in biology sensor reaction tank, pump into quantitative buffer solution, the concentration of test glucose and glutamic acid is also stored to computing machine simultaneously;
Be completed at every turn, waste liquid drain pump and reaction tank drain pump automatically start under the control of control system, solution in sampling pool, colorimetric diluting tank, diluting tank and/or biology sensor reaction tank is evacuated to waste liquid pool, starts sampling pump simultaneously the solution in sampling pool is evacuated to waste liquid pool;
4) clean: after being completed, reaction tank scavenging pump extracts damping fluid to biology sensor reaction tank, to clean biology sensor reaction tank, after cleaning, by reaction tank drain pump, damping fluid is evacuated to waste liquid pool.
As preferred implementation, dilution is the mixed aqueous solution of Sodium Benzoate and EDETATE SODIUM, and the mass concentration of Sodium Benzoate is 2%, and the mass concentration of EDETATE SODIUM is 1%.
As preferred implementation, buffer solution is phosphate buffer solution.
As preferred implementation, in step 1), twice measuring result error range of set value is 0.1-4%.
As preferred implementation, step 2) in, the quantitative dilution of fermentation liquor be to 10 times of fermentation liquor dilutions.
Embodiment 1:
Selecting the bacterial classification of in Dec, 2012 switching is seed, adopts this glutamic acid fermentation on-line detecting system to detect online the method for glutamic acid and concentration of glucose, comprises the following steps:
1) calibration: system is controlled sampling valve 3 and extracted 40uL standard solution to biology sensor reaction tank 8 by standard specimen pond 4, simultaneously by reaction tank scavenging pump 17 to the buffer solution that pumps into 20mL in bio-sensing reaction tank 8, the concentration of bioassay standard glucose in solutions and glutamic acid; METHOD FOR CONTINUOUS DETERMINATION twice, twice measuring result error is less than 4%(setting value), system shows to calibrate to be passed through;
2) OD value test: it is 7min that sample interval is set, control system extracts fermentation liquor in fermentation tanks to sampling pool 4 according to the time interval controls sampling pump 14 arranging; Sampling valve 3 extracts 40 μ L fermentation liquors to colorimetric diluting tank 5 by sampling pool 4, and then dilution fillup valve 2 extraction dilutions dilute 10 times to 5 pairs of fermentation liquors of colorimetric diluting tank, and fermentation liquor OD value directly shows on computers;
3) sample test: the sample solution that sampling valve 3 extracts 40 μ L by colorimetric diluting tank 5 is to diluting tank 7; System is controlled dilution fillup valve 2 and inject dilution in diluting tank 7, and system is quantitatively diluted according to the fermentation liquor in the further contrast colors diluting tank of extension rate in advance, and it is 0 times (not diluting) that extension rate is set; Sampling valve 3 by diluting tank 7 draw samples solution to biology sensor reaction tank 8, simultaneously by reaction tank scavenging pump 17 to the buffer solution that pumps into 20mL in bio-sensing reaction tank 8, the concentration of test glucose and glutamic acid is also stored to computing machine;
Be completed at every turn, system starts waste liquid drain pump 15 and reaction tank drain pump 16 automatically, solution in sampling pool 4, colorimetric diluting tank 5, diluting tank 7 and/or biology sensor reaction tank 8 is evacuated to waste liquid pool, starts sampling pump 14 simultaneously the solution in sampling pool 4 is evacuated to waste liquid pool 19;
4) clean: after being completed, reaction tank scavenging pump 17 extracts damping fluid to biology sensor reaction tank 8, to clean biology sensor reaction tank 8, after cleaning, by reaction tank drain pump 16, damping fluid is evacuated to waste liquid pool 19.
Wherein, the consisting of of dilution: the mixed aqueous solution of Sodium Benzoate and EDETATE SODIUM, the mass concentration of Sodium Benzoate is 2%, the mass concentration of EDETATE SODIUM is 1%.
Buffer solution is phosphate buffer solution.
Embodiment 2:
Selecting the bacterial classification of in February, 2013 switching is seed, adopts this glutamic acid fermentation on-line detecting system to detect online the method for glutamic acid and concentration of glucose, comprises the following steps:
1) calibration: system is controlled sampling valve 3 and extracted 40uL standard solution to biology sensor reaction tank 8 by standard specimen pond 4, simultaneously by reaction tank scavenging pump 17 to the buffer solution that pumps into 20mL in bio-sensing reaction tank 8, the concentration of bioassay standard glucose in solutions and glutamic acid; METHOD FOR CONTINUOUS DETERMINATION three times, double measuring result error is less than 4%(setting value), system shows to calibrate to be passed through;
2) OD value test: it is 255min that sample interval is set, control system extracts fermentation liquor in fermentation tanks to sampling pool 4 according to the time interval controls sampling pump 14 arranging; Sampling valve 3 extracts 60 μ L fermentation liquors to colorimetric diluting tank 5 by sampling pool 4, and then dilution fillup valve 2 extraction dilutions carry out 10 times of dilutions to 5 pairs of fermentation liquors of colorimetric diluting tank, and fermentation liquor OD value directly shows on computers;
3) sample test: the sample solution that sampling valve 3 extracts 40 μ L by colorimetric diluting tank 5 is to diluting tank 7; System is controlled dilution fillup valve 2 and inject dilution in diluting tank 7, and system is quantitatively diluted according to the fermentation liquor in the further contrast colors diluting tank of extension rate in advance, and extension rate is 500 times; After dilution, sample solution after sampling valve 3 is diluted by diluting tank 7 extractions is to biology sensor reaction tank 8, by reaction tank scavenging pump 17, in bio-sensing reaction tank 8, pump into quantitative buffer solution, the concentration of test glucose and glutamic acid is also stored to computing machine simultaneously;
Be completed at every turn, system starts waste liquid drain pump 15 and reaction tank drain pump 16 automatically, solution in sampling pool 4, colorimetric diluting tank 5, diluting tank 7 and biology sensor reaction tank 8 is evacuated to waste liquid pool, starts sampling pump 14 simultaneously the solution in sampling pool 4 is evacuated to waste liquid pool 19;
4) clean: after being completed, reaction tank scavenging pump 17 extracts damping fluid to biology sensor reaction tank 8, to clean biology sensor reaction tank 8, after cleaning, by reaction tank drain pump 16, damping fluid is evacuated to waste liquid pool 19.
Wherein, the consisting of of dilution: the mixed aqueous solution of Sodium Benzoate and EDETATE SODIUM, the mass concentration of Sodium Benzoate is 2%, the mass concentration of EDETATE SODIUM is 1%.Buffer solution is phosphate buffer solution.
Embodiment 3:
Selecting the bacterial classification of in June, 2013 switching is seed, adopts this glutamic acid fermentation on-line detecting system to detect online the method for glutamic acid and concentration of glucose, comprises the following steps:
1) calibration: system is controlled sampling valve 3 and extracted 40uL standard solution to biology sensor reaction tank 8 by standard specimen pond 4, simultaneously by reaction tank scavenging pump 17 to the buffer solution that pumps into 20mL in bio-sensing reaction tank 8, the concentration of bioassay standard glucose in solutions and glutamic acid; METHOD FOR CONTINUOUS DETERMINATION twice, twice measuring result error is less than 4%(setting value), system shows to calibrate to be passed through;
2) OD value test: it is 100min that sample interval is set, control system extracts fermentation liquor in fermentation tanks to sampling pool 4 according to the time interval controls sampling pump 14 arranging; Sampling valve 3 extracts 40 μ L fermentation liquors to colorimetric diluting tank 5 by sampling pool 4, and then dilution fillup valve 2 extraction dilutions carry out 10 times of dilutions to 5 pairs of fermentation liquors of colorimetric diluting tank, and fermentation liquor OD value directly shows on computers;
3) sample test: the sample solution that sampling valve 3 extracts 40 μ L by colorimetric diluting tank 5 is to diluting tank 7; System is controlled dilution fillup valve 2 and inject dilution in diluting tank 7, and system is quantitatively diluted according to the fermentation liquor in the further contrast colors diluting tank of extension rate in advance, and extension rate is 20 times; After dilution, sample solution after sampling valve 3 is diluted by diluting tank 7 extractions is to biology sensor reaction tank 8, by reaction tank scavenging pump 17, in bio-sensing reaction tank 8, pump into quantitative buffer solution, the concentration of test glucose and glutamic acid is also stored to computing machine simultaneously;
Be completed at every turn, system starts waste liquid drain pump 15 and reaction tank drain pump 16 automatically, solution in sampling pool 4, colorimetric diluting tank 5, diluting tank 7 and biology sensor reaction tank 8 is evacuated to waste liquid pool, starts sampling pump 14 simultaneously the solution in sampling pool 4 is evacuated to waste liquid pool 19;
4) clean: after being completed, reaction tank scavenging pump 17 extracts damping fluid to biology sensor reaction tank 8, to clean biology sensor reaction tank 8, after cleaning, by reaction tank drain pump 16, damping fluid is evacuated to waste liquid pool 19.
Wherein, the consisting of of dilution: the mixed aqueous solution of Sodium Benzoate and EDETATE SODIUM, the mass concentration of Sodium Benzoate is 2%, the mass concentration of EDETATE SODIUM is 1%.Buffer solution is phosphate buffer solution.
These are only preferred embodiment of the present invention, in order to limit the present invention, within the spirit and principles in the present invention not all, any modification of doing, be equal to replacement, improvement etc., within all should being included in protection scope of the present invention.
Claims (10)
1. a glutamic acid fermentation on-line detecting system, comprises sampling pump, and described sampling pump inlet end has fermentation tank by pipeline communication, described sampling pump endpiece has sampling pool by pipeline communication, there is waste liquid drain pump described sampling pool bottom by pipeline communication, it is characterized in that, also comprises:
Diluting tank, be arranged on described sampling pool right side and with described sampling pool on same straight line, as the mixing pit of dilution and sample, its bottom is by waste liquid drain pump inlet end described in pipeline communication, and the endpiece of described waste liquid drain pump is by pipeline communication waste liquid pool;
Dilution fillup valve, is fixedly connected with mechanical arm, for injecting dilution to described diluting tank;
Biology sensor reaction tank, is arranged on described diluting tank right side, and its bottom is by the pipeline communication pond scavenging pump endpiece that responds, and described reaction tank scavenging pump inlet end has damping fluid storage tank by pipeline communication; Its bottom is by the respond inlet end of pond drain pump of another pipeline communication, and the endpiece of described reaction tank drain pump is by waste liquid pool described in pipeline communication; In described biology sensor reaction tank, be provided with post glucose oxidase enzyme membrane the first Catalase, post the second Catalase of dglutamic oxidase enzyme membrane; Described the first Catalase and the second Catalase are electrically connected with ADS converting system, and described ADS converting system connects the control system of computing machine;
Standard specimen pond, is arranged between described diluting tank and described sampling pool, demarcates the standard solution of described the first Catalase and described the second Catalase for splendid attire;
Sampling valve, is fixedly connected with described mechanical arm, for the sample that extracts described sampling pool, diluting tank or standard specimen pond to described biology sensor reaction tank;
Control system, is electrically connected to described sampling pump, controls sampling interval, the sampling quantity of described sampling pump; Be electrically connected to described mechanical arm, control the direction of motion of described mechanical arm; Be electrically connected to described dilution fillup valve, control dilution injection rate IR and injection phase; Be electrically connected to described sampling valve, control sampling location, sampling quantity and the injection phase of described sampling valve; Be electrically connected to and control described sampling pump, reaction tank drain pump, waste liquid drain pump, reaction tank scavenging pump.
2. glutamic acid fermentation on-line detecting system as claimed in claim 1, it is characterized in that: also comprise colorimetric diluting tank, described colorimetric diluting tank comprises the pond body of splendid attire sample, described pond body sidewall is oppositely arranged light source emitter and light source absorption plant, described light source is 570nm light source, the sample of described light source in the body of described pond be laggard enters light source absorption plant, by the OD value of light source absorption plant test sample and light absorption signal is transmitted by ADS converter and be stored to computing machine; Body bottom, described pond is by the entrance of waste liquid drain pump described in pipeline communication.
3. glutamic acid fermentation on-line detecting system as claimed in claim 2, is characterized in that: described biology sensor reaction tank side wall upper part is communicated with level control (LC) pipe, and the described level control (LC) pipe other end is communicated with described waste liquid pool.
4. glutamic acid fermentation on-line detecting system as claimed in claim 3, is characterized in that: described sampling pump, waste liquid drain pump, reaction tank drain pump and reaction tank scavenging pump are peristaltic pump.
5. glutamic acid fermentation on-line detecting system as claimed in claim 4, is characterized in that: described diluting tank, colorimetric diluting tank and/or biology sensor reaction tank bottom are provided with rabbling mechanism, and described rabbling mechanism is controlled by described control system.
6. adopt glutamic acid fermentation on-line detecting system as claimed in claim 5 to detect online the method for glutamic acid and concentration of glucose, it is characterized in that, comprise the following steps:
1) calibration: described control system is controlled described sampling valve and extracted 10-60uL standard solution to described biology sensor reaction tank by described standard specimen pond, simultaneously by described reaction tank scavenging pump, in described bio-sensing reaction tank, pump into quantitative buffer solution, measure the concentration of glucose and glutamic acid in described standard solution; METHOD FOR CONTINUOUS DETERMINATION at least twice, double measuring result error is less than setting value, and calibration is passed through;
2) OD value test: it is 7-255min that sample interval is set, described control system extracts fermentation liquor in described fermentation tank to described sampling pool according to sampling pump described in the time interval controls arranging; Described sampling valve extracts described in 10-60 μ L fermentation liquor to described colorimetric diluting tank by described sampling pool, then described dilution fillup valve extraction dilution quantitatively dilutes described fermentation liquor to described colorimetric diluting tank, and described fermentation liquor OD value directly shows on computers;
3) sample test: the sample solution that described sampling valve extracts 10-60 μ L by described colorimetric diluting tank is to described diluting tank; Described control system is controlled described dilution fillup valve and inject dilution in described diluting tank, according to extension rate in advance, further the fermentation liquor in described colorimetric diluting tank is quantitatively diluted, and described extension rate is 0-500 times; After dilution, sample solution after described sampling valve is diluted by described diluting tank extraction is to described biology sensor reaction tank, by described reaction tank scavenging pump, in described biology sensor reaction tank, pump into quantitative buffer solution, the concentration of test glucose and glutamic acid is also stored to computing machine simultaneously;
Be completed at every turn, described waste liquid drain pump and reaction tank drain pump automatically start under the control of described control system, solution in described sampling pool, colorimetric diluting tank, diluting tank and/or biology sensor reaction tank is evacuated to waste liquid pool, starts described sampling pump simultaneously the solution in described sampling pool is evacuated to described waste liquid pool;
4) clean: after being completed, described reaction tank scavenging pump extracts damping fluid to described biology sensor reaction tank, to clean described biology sensor reaction tank, after cleaning, by described reaction tank drain pump, described damping fluid is evacuated to described waste liquid pool.
7. the method for online detection glutamic acid as claimed in claim 6 and concentration of glucose, it is characterized in that: described dilution is the mixed aqueous solution of Sodium Benzoate and EDETATE SODIUM, the mass concentration of described Sodium Benzoate is 2%, and the mass concentration of described EDETATE SODIUM is 1%.
8. the method for online detection glutamic acid as claimed in claim 6 and concentration of glucose, is characterized in that: described buffer solution is phosphate buffer solution.
9. the method for online detection glutamic acid as claimed in claim 6 and concentration of glucose, is characterized in that: in step 1), described twice measuring result error range of set value is 0.1-4%.
10. the method for online detection glutamic acid as claimed in claim 6 and concentration of glucose, is characterized in that: step 2) in, the quantitative dilution of described fermentation liquor is to 10 times of fermentation liquor dilutions.
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| CN104007054A (en) * | 2014-06-13 | 2014-08-27 | 南京工业大学 | Online detection device and method for thallus concentration in biological fermentation process |
| CN104459169A (en) * | 2014-09-26 | 2015-03-25 | 浙江工商大学 | Concentration detection device and method for D-fructose solution |
| CN105890923A (en) * | 2014-12-11 | 2016-08-24 | 山东省科学院生物研究所 | Sterile trace on-line sampling device and sampling method |
| CN105890923B (en) * | 2014-12-11 | 2019-08-09 | 山东省科学院生物研究所 | A kind of sterile micro on-line period device and sampling method |
| CN107102086A (en) * | 2017-06-24 | 2017-08-29 | 深圳源广安智能科技有限公司 | A kind of Beer Brewage material quality evaluation system |
| CN107589669A (en) * | 2017-09-09 | 2018-01-16 | 北京化工大学 | It is a kind of to utilize the fermentation process intelligent monitoring method and system for measuring lag information |
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