CN101845463A - Control method for producing 1,3-propanediol by substrate fed-batch fermentation on basis of growth and metabolism characteristics - Google Patents

Abstract

The invention discloses a control method for producing 1,3-propanediol by substrate fed-batch fermentation on the basis of growth and metabolism characteristics, belonging to the technical field of biochemical engineering. The method comprises the following steps of: firstly deriving substrate fed-batch control functions coupled to growth and metabolism characteristics of thallus: logarithm growth period of F1=k1C+k2F2 (1) and stabilization period of F1=b+k3F2 (2); and then fermenting through manual control to determine values of k1, k2, k3 and b, wherein the selection of the fed-batch scheme is based on the concentration of CO2 and comprises the steps of: executing a scheme (1) when Ct+1 > Ct; and executing a scheme (2) when Ct+1 < Ct; and finally building a set of on-line detection and automatic control fermentation system so that the concentration of the substrate in the fermentation process is always 18+/-2g/L, and the concentration of 2,3-PD is up to 82.64g/L, improved by 33.14 percent in comparison with 55.25g/L by manual control. The method realizes on-line automatic control of the concentration of the substrate in the fermentation process, solves the instability of manual control and effectively improves the concentration of 1,3-PD.

Description

Bottoms stream based on growth and metabolic characteristics adds fermentative production 1, the control method of ammediol

Technical field

The invention belongs to technical field of biochemical industry, relate to a kind of fermentative production 1, the control techniques of ammediol relates in particular to a kind of batch of formula stream and adds fermentative production 1, the ammediol process with cell growth and metabolic characteristics mutually the bottoms stream that joins of coupling add autocontrol method.

Background technology

1, and ammediol (English name: 1,3-Propanediol is called for short 1, is a kind of extremely important industrial chemicals 3-PD), has wide application field.1,3-PD can be used as the synthesis material of softening agent, washing composition, sanitas and emulsifying agent, also can be used as the performance that improves product as the additive in the products such as printer ink, sanitising agent, stablizer and fuel-cell fuel; 1,3-PD can be used as the intermediate of medicine and organic synthesis, is used for industries such as food, makeup and pharmacy; 1, the alternative ethylene glycol of 3-PD, 1, the how pure polyester of intermediate production such as 4-butyleneglycol and neopentyl glycol and extend agent as carbochain; 1, the topmost purposes of 3-PD is the monomer as synthesizing polyester and urethane, especially as new polyester---and the monomer of polytrimethylene terephthalate (PTT).PTT is the another extremely rising new polyester material after polyethylene terephthalate (PET), polybutylene terephthalate (PBT), the novel thermoplastic polyester material that is a kind of high-performance, easily processes, have excellent elasticity renaturation and low temperature dyeing performance, have broad application prospects in various fields such as carpet industry, dress materials and engineering thermoplasts.

At present, 1, the industrialized preparing process of 3-PD is divided into chemical synthesis and microbe fermentation method.Microbe fermentation method has because of it that raw material sources are extensive, operational condition is gentle and environmental pollution is little etc. that advantage is subjected to extensive concern.Criticize formula stream and add fermentative production 1,3-PD is the main mode of production of microbe fermentation method, because this mode can obtain higher production concentration and transformation efficiency.The concentration of substrate that batch formula stream adds in the fermenting process is to influence 1, the key controlled variable of 3-PD output.Removing E.I.Du Pont Company's employing at present in the world is that the substrate one-step fermentation produces 1 with glucose, and outside the 3-PD technology, other technologies all are that the substrate one-step fermentation produces 1,3-PD with glycerine.With glycerine is that fermenting substrate produces 1, and 3-PD is a typical growth coupling connection type process, and product forms with the cell growth closely related.The cell growth is closely related with concentration of substrate, and concentration of substrate is too high can cell growth to produce inhibition, and concentration is crossed low then restrictive cell growth, and two kinds of situations all are unfavorable for 1, and the production of 3-PD needs concentration of substrate is controlled in the reasonable range for this reason.But because glycerol concentration is difficult to on-line quick detection, so the automatic control that bottoms stream adds in the fermenting process is an insoluble critical problem always.Although all there is the scholar to attempt addressing this problem both at home and abroad, proposing glycerine stream the earliest as German scholar A.Reimann and H.Biebl adds and pH regulator strategy (the Biotech Lett of coupling connection mutually, 1996,18 (7): 827-832), control strategy (the food and fermentation industries that people such as Tsing-Hua University's journey cocoa adopt glycerine to combine with alkali coupling and constant speed stream glycerol adding, 2004,30 (4)), the big feedback feed supplement control techniques (publication number CN101153292A) that proposes that waits of the Tan Tian of Beijing University of Chemical Technology, but these methods all are rule of thumb to control glycerol concentration, and the cell that has no basis is grown and metabolic feature is accurately controlled automatically.The present invention has set up base consumption and thalli growth (by CO in the tail gas by the fermentation kinetics analysis ₂Concentration characterizes) and metabolism (characterizing) by the alkali lye consumption between funtcional relationship, and data fitting has been determined facies relationship numerical value by experiment, and then the bottoms stream of the different steps that obtained fermenting adds strategy, has realized the automatic control of concentration of substrate in the fermenting process.

Summary of the invention

It is that fermenting substrate produces 1 with glycerine that the technical problem to be solved in the present invention provides a kind of, and the bottoms stream of 3-PD adds autocontrol method, mainly comprises three aspect contents: (1) and thalli growth characteristic and the metabolic characteristic bottoms stream of coupling connection mutually add the control function derivation; (2) the bottoms stream rate of acceleration is with tail gas CO ₂Concentration and pH value be the segmentation control scheme of coupling connection mutually; (3) detection and stream add automatic control automatically.

Among the present invention in the fermenting process stream of substrate glycerine add scheme and depend on CO in the fermentation tank tail gas ₂The variation of concentration, the stream rate of acceleration of substrate is then passed through CO ₂The wear rate of concentration and alkali lye is determined by corresponding functional relation.CO in tail gas ₂Concentration is increases trend gradually, when thalli growth is in logarithmic phase, and base consumption speed and CO ₂Concentration and alkali lye wear rate are relevant; And CO ₂Concentration is downtrending gradually, and when thalli growth was in stable or decline phase, base consumption speed and alkali lye wear rate were linear.According to these two relational expressions, constitute automatic control system by computer, transmitter and corresponding handling equipment, realize 1, the automatic control that bottoms stream adds in the ammediol fermenting process, concentration of substrate is controlled at a reasonable levels, both help the cell growth, helped the formation of target product again.

For achieving the above object, the technical solution adopted in the present invention is, derive earlier with thalli growth characteristic and metabolic characteristic mutually the bottoms stream that joins of coupling add control function, carry out the anaerobically fermenting experiment by Artificial Control then, preliminary determine that bottoms stream adds in the control function and the cell growth parameter relevant with metabolic characteristics, and add scheme according to the characteristics design bottoms stream of different times cell growth, again this bottoms stream is added automatic control system and be used for fermenting experiment;

Step 1, derivation and thalli growth and the metabolic characteristic bottoms stream of coupling connection mutually add control function.

In the fermenting process because continue to flow glycerol adding and alkali lye cause fermentating liquid volume constantly to change.A substrate part that consumes is used for synthetic thalline, and another part is used to keep cell activities and generates meta-bolites.Suppose that fermenting process meets the non-structural models of balanced growth, then base consumption can by under the description that establishes an equation:

$-\frac{\mathrm{dS}}{\mathrm{dt}}=\frac{1}{Y_X^{*}}\frac{\mathrm{dX}}{\mathrm{dt}}+\mathrm{mX}+X\underset{i=1}{\overset{N}{\mathrm{\&Sigma;}}}\frac{q_{\mathrm{Pi}}}{Y_{\mathrm{Pi}}}---\left(1\right)$

---glycerine consumption (g) X---total biomass (g) wherein: S

Y _X ^*---thalline yield coefficients (g/g) m---keeps constant (h ^-1)

Y _Pi---yield coefficients (g/g) q of i kind product _Pi---the specific production rate (g/g.h) of i kind product

T---fermentation time (h) N---meta-bolites kind sum

The variation that batch formula stream adds concentration of substrate in the fermenting process is because thalli growth and metabolic consumption are to cause fermentating liquid volume to change the dilution effect that is produced because bottoms stream adds on the other hand on the one hand.Therefore, the stream rate of acceleration of substrate should satisfy following formula:

${\mathrm{<figure-callout\; id="1"\; label="C\; S\; F"\; filenames="HDA0000021176250000021.png"\; state="\{\{state\}\}">C</figure-callout>}}_S{F}_{}{}_1=\frac{\mathrm{dS}}{\mathrm{dt}}+C_0\frac{\mathrm{dV}}{\mathrm{dt}}---\left(2\right)$

Fermentating liquid volume changes available following formula to be described:

$\frac{\mathrm{dV}}{\mathrm{dt}}=\frac{{\mathrm{<figure-callout\; id="1"\; label="F"\; filenames="HDA0000021176250000021.png"\; state="\{\{state\}\}">F</figure-callout>}}_1}{{\mathrm{\&rho;}}_1}+\frac{{\mathrm{<figure-callout\; id="2"\; label="F"\; filenames="HDA0000021176250000021.png"\; state="\{\{state\}\}">F</figure-callout>}}_2}{{\mathrm{\&rho;}}_2}---\left(3\right)$

C wherein _SBe the massfraction (g/g) of glycerine in the current adding substrate, C ₀Be substrate glycerol concentration (g/L) in the fermented liquid, V is fermentating liquid volume (L), F ₁Be bottoms stream rate of acceleration (g/h), F ₂Be alkaline stream rate of acceleration (g/h), ρ ₁Represent substrate proportion (g/L), ρ ₂Represent alkali lye proportion (g/L).

Keep pH value in the fermented liquid 7.0 ± 0.02 by adding alkaline solution in the fermenting process, the relational expression that then corresponding alkali consumption and cellular metabolism generate total acid content is:

$\frac{C_2{\mathrm{<figure-callout\; id="2"\; label="F"\; filenames="HDA0000021176250000021.png"\; state="\{\{state\}\}">F</figure-callout>}}_2}{{\mathrm{\&rho;}}_2}=X\underset{j=1}{\overset{n}{\mathrm{\&Sigma;}}}{\mathrm{Ka}}_j\frac{q_j}{m_j}---\left(4\right)$

C wherein ₂Be the volumetric molar concentration (mol/L) of alkaline solution, q _jBe the j kind organic acid specific production rate (g/g.h) that bacterial metabolism produces, m _jAnd K _AjBe respectively j kind organic acid molar mass and the coefficient that dissociates.Consolidated equation (1)～(4) can draw bottoms stream rate of acceleration equation:

${\mathrm{<figure-callout\; id="1"\; label="F"\; filenames="HDA0000021176250000021.png"\; state="\{\{state\}\}">F</figure-callout>}}_1=a\frac{\mathrm{dX}}{\mathrm{dt}}+\mathrm{bX}+c{F}_2---\left(5\right)$

Wherein $a=\frac{{\mathrm{\&rho;}}_1}{C_S{\mathrm{\&rho;}}_1-C_0}\frac{1}{Y_X^{*}},$ $b=m+\underset{k=1}{\overset{K}{\mathrm{\&Sigma;}}}\frac{q_k}{Y_k},$ $c=\frac{{\mathrm{\&rho;}}_1}{C_S{\mathrm{\&rho;}}_1-C_0}(\frac{C_0}{{\mathrm{\&rho;}}_2}+\frac{{\mathrm{<figure-callout\; id="2"\; label="C"\; filenames="HDA0000021176250000021.png"\; state="\{\{state\}\}">C</figure-callout>}}_2}{{\mathrm{\&rho;}}_2}\frac{\mathrm{\&Sigma;}\frac{q_j}{Y_j}}{\mathrm{\&Sigma;}{K}_{\mathrm{aj}}\frac{q_j}{m_j}})$

q _kBe the specific production rate (g/g.h) of k kind non-organic acid product, Y _kBe its yield coefficients.

Show by historical analysis of experimental data: a item proportion in the formula (5) is very little, can ignore, and therefore equation (5) further can be reduced to:

F ₁＝bX+cF ₂ (6)

According to the thalli growth situation fermenting process being divided into two stages controls: the fs is a logarithmic phase, and this stage thalline is grown fast, and biomass changes with CO in the tail gas ₂Change in concentration is linear dependence; Subordinate phase is stationary phase, and this stage biomass changes less relatively, can suppose that X is a constant.Therefore equation (6) can further be rewritten as:

Logarithmic phase F ₁=k ₁C+k ₂F ₂(7)

Stationary phase F ₁=b+k ₃F ₂(8)

Wherein C is CO ₂Concentration, F ₁Be bottoms stream rate of acceleration (g/h), F ₂Be alkaline stream rate of acceleration (g/h), k ₁, k ₂, k ₃, b is constant.

Step 2, acquisition segmentation control function equation

At first add the formula stream of criticizing and add fermentation, and write down related data related in the step 1 during the fermentation by the Artificial Control bottoms stream.Its method is as follows:

(1) preparation of seed liquor: preparation seed culture medium, 115 ℃～121 ℃ sterilization 15～20min; Cray Bai Shi bacillus CGMCC 2028 bacterial strains are inserted seed culture medium, under 37 ℃, 100～200rpm condition, cultivate 10～15h.

(2) preparation of fermention medium: preparation fermention medium, 115 ℃～121 ℃ sterilization 15～20min.The initial concentration of substrate glycerine is 20～80g/L.

(3) anaerobically fermenting: insert in (2) cultivating the seed culture fluid that obtains in (1), inoculum size 5～25% is 150～450rpm, anaerobism (0.1～1vvm N at pH value 6.50～7.50,37 ℃ of temperature, mixing speed ₂) the condition bottom fermentation, concentration of substrate is 10～40g/L in manual control fermentation's liquid, fermentation stops fermentation when proceeding to 25～35h.In the fermenting process, the consumption of alkali lye and the consumption of substrate are measured by the constant speed pump; CO in the tail gas ₂Concentration is passed through CO ₂The analyser on-line measurement; Meta-bolites detects by gas-chromatography and high performance liquid chromatography.

Record CO by above-mentioned anaerobically fermenting ₂Concentration, base consumption speed and alkali wear rate, then according to the equation in the step 1 (7) and (8) respectively to the CO of thalline in logarithmic phase and stationary phase ₂Concentration, base consumption speed and alkali wear rate data are carried out match, determine k ₁, k ₂, k ₃With the value of b, can obtain the stream of substrate and add the controlling party formula in the different fermentations stage.

Experimental result shows: at thalline logarithmic phase, CO ₂Concentration is and increases progressively trend, and the trend that tapers off in stationary phase.Therefore, bottoms stream adds the control Scheme Selection and depends on CO in the fermenting process ₂The variation of concentration: when thalli growth is in logarithmic phase, CO ₂Concentration increases gradually, and the bottoms stream rate of acceleration is by CO ₂Concentration and alkali lye wear rate calculate; When thalli growth is in stationary phase, CO ₂Concentration reduces gradually, and the stream rate of acceleration of substrate is calculated by the alkali lye wear rate.

Step 3, the foundation and the control fermentation automatically of fermentation control system automatically

Controlling System of the present invention comprises fermentor tank, computer, pH electrode, CO ₂Analyser, alkali lye transferpump and substrate transferpump.Automatically control fermentation pH by this system, and alkali speed F mended in record ₂CO ₂The CO that analyser detects ₂Concentration value is sent to computer, relatively more different by analysis CO constantly ₂After the variation of concentration, instruction substrate transferpump adds scheme by homogeneous turbulence not and carries out feed supplement, and the concentration of substrate in the fermented liquid is maintained in the reasonable range.

Bottoms stream adds automatic control and online detection, and process is as follows:

1) seed culture fluid is inserted in the fermention medium of fermentor tank 1 and begin to carry out anaerobically fermenting;

2) in the process that fermentation is carried out, by the pH value of pH electrode 2 detection fermented liquids, its value is delivered in the computer 8 through the fermentor tank electric control box, and pH value deviation is ± 0.02;

3) computer 8 is judged according to the pH value in the fermented liquid, when it is lower than set(ting)value, opens alkali lye feed adjustment valve 4, alkali lye is delivered to fermentor tank 1 by alkali lye transferpump 5 by alkali lye storage tank 11, get back to set(ting)value until the pH value, close alkali lye transfer valve 4, alkali speed F mended in record simultaneously ₂

4) pass through CO ₂The CO that analyser 7 detects in the vapor pipe 6 ₂Concentration, its value is transferred into computer 8;

5) computer 8 is controlled Scheme Choice according to the logical program of accompanying drawing 1: as t+1 CO constantly ₂Concentration value C _T+1Greater than t concentration value C constantly _tThe time (this moment cell be in logarithmic phase), press equation (7) calculating bottoms stream rate of acceleration F ₁Otherwise, i.e. C _T+1＜C _tThe time (cell is in stationary phase), press equation (8) and calculate bottoms stream rate of acceleration F ₁

6) the computer 8 bottoms stream rate of acceleration F definite suddenly according to previous step ₁, substrate feed adjustment valve 10 is opened in control, and substrate glycerine is delivered to fermentor tank 1 through substrate transferpump 9 by substrate storage tank 12, until next control cycle;

7) repeating step (2) to (6) is until fermentation ends.

Fermenting experiment: seed liquor, fermention medium preparation, fermentation culture are identical with method described in the step 2, adopt method described in the step 3 to carry out substrate control in the fermenting process, finish experiment behind the fermentation 30h, and sampling detects 1,3-PD concentration.

The present invention realized the on-line automatic control of concentration of substrate in the fermenting process, set up cell and added scheme and according to CO in the bottoms stream of different growing stages ₂Concentration determines that bottoms stream adds scheme, has simplified experimental implementation, has solved manually operated unstable, and has improved 1 effectively, the concentration of 3-PD.

Description of drawings

Fig. 1 is based on the fermentative production 1 of cell growth and metabolic characteristics, and the bottoms stream of 3-PD adds the automatic control program logical flow chart.

The bottoms stream that Fig. 2 is based on cell growth and metabolic characteristics adds production 1, the online substrate automatic control system device synoptic diagram of 3-PD.

Among the figure: 1 fermentor tank; The 2pH electrode; The 3pH signal converter; 4 alkali lye feed adjustment valves; 5 alkaline solution transferpumps; 6 vapor pipes; 7CO ₂Analyser; 8 computers; 9 substrate transferpumps; 10 substrate feed adjustment valves; 11 alkali lye storage tanks; 12 substrate storage tanks.

Wherein: solid line is represented liquid transmission pipeline, and long and short dash line is represented gas transfer pipeline, dotted line representation signal bang path.

Embodiment

Below be described in detail specific embodiments of the invention for combination technology scheme and accompanying drawing.

Fermentative production 1 provided by the invention, the bottoms stream of 3-PD adds autocontrol method, according to the characteristics of thalline logarithmic phase and stationary phase bottoms stream in its fermenting process is added and to be divided into two kinds of control schemes and to carry out, concentration of substrate in the fermented liquid is controlled in the constant scope, neither produce the substrate restriction, do not produce substrate yet and suppress, thereby improved 1, the ultimate density of 3-PD.

Bottoms stream provided by the invention adds automatic control embodiment and installs as depicted in figs. 1 and 2, and its implementation process is with summary of the invention 4.

Embodiment

1) bacterial classification: Cray Bai Shi bacillus (Klebsiella pneumoniae) CGMCC 2028

2) substratum:

(1) seed culture medium (g/L): glycerine: 20, MgSO ₄7H ₂O:0.2, KH ₂PO ₄: 1.3, (NH ₄) ₂SO ₄: 2.0, K ₂HPO ₄3H ₂O:4.56, yeast powder: 2, CaCO ₃: 2.Trace element A:2mL/L, Fe ²⁺Solution: 1mL/L, Ca ²⁺Solution: 1mL/L.

Trace element A (mg/L): ZnCl ₂: 70, MnCl ₂4H ₂O:100, H ₃BO ₃: 60, CoCl ₂6H ₂O:200, CuCl ₂6H ₂O:20, NiCl ₂6H ₂O:25, Na ₂MoO ₄2H ₂O:5.

Fe ²⁺Solution (L ^-1): FeSO ₄7H ₂O:5g, HCl (12M): 4mL

Ca ²⁺Solution (g/L): CaCl ₂: 20.

(2) fermention medium (g/L): glycerine: 40, (NH ₄) ₂SO ₄: 6.61, KH ₂PO ₄: 1.36, MgCl ₂6H ₂O:0.26, CaCl ₂: 0.29, citric acid: 0.42, yeast powder: 1.Trace element B:5mL/L.

Trace element B (g/L): ZnCl ₂: 0.68, MnCl ₂4H ₂O:0.17, H ₃BO ₃: 0.06, CuCl ₂2H ₂O:0.47, Na ₂MoO ₄2H ₂O:0.005, FeCl ₂4H ₂O:3.97, CoCl ₂6H ₂O:0.47.HCl(12M)：10mL/L.

3) alkali lye: 5mol/LNaOH solution.

4) add substrate: 95% glycerine

5) detection of substrate and meta-bolites in the fermented liquid

A. the mensuration of glycerol concentration: sodium periodate oxidation

B. the mensuration of cell density: spectrophotometer method

C. organic acid content testing: high performance liquid chromatography

1. The pretreatment: in 12, the centrifugal 10min of 000rpm gets supernatant liquor with fermented liquid, Deproteinization, and suitably after the dilution, the moisture film of crossing 0.22 μ m is standby;

2. sample determination: sample size 10 μ L, ultraviolet detection wavelength 214nm adopts Kromasil C ₁₈Post, moving phase are 0.2% phosphoric acid: acetonitrile=96.5: 3.5 (V/V), flow velocity are 1.0mL/min, 20 ℃ of column temperatures.

D.1,3-PD assay: vapor-phase chromatography

1. The pretreatment: in 12, the centrifugal 10min of 000rpm gets supernatant liquor and carries out sample detection with fermented liquid;

2. sample determination: sample size 1 μ L, detector is FID, adopts the Chromsorb101 packed column, carrier gas is a nitrogen, 170 ℃ of column temperatures, 220 ℃ of injector temperatures, 200 ℃ of detector temperatures.

E.CO ₂Concentration determination: CO ₂Analyser

6) the concrete experimental procedure of present embodiment:

(1) anaerobically fermenting experiment

A. the preparation of seed liquor: the preparation seed culture medium, it is divided in 250mL shakes in the bottle, each liquid amount that shakes in the bottle is 100mL, 121 ℃ of sterilization 20min; 1mL Cray Bai Shi bacillus seed liquor is inserted in the seed culture medium, cultivate 12h and obtain seed liquor in 37 ℃, 150rpm shaking table anaerobism;

B. the preparation of fermention medium: preparation fermention medium, 121 ℃ of sterilization 20min.

C. anaerobically fermenting: insert in (2) cultivating the seed culture fluid that obtains in (1), inoculum size is 10%, at 7.00 ± 0.02,37 ℃ of pH values, 300rpm, anaerobism (0.1vvm N ₂) carrying out anaerobically fermenting under the condition, the permanent concentration of substrate of pre-control is 15～20g/L in the fermented liquid, fermentation time is about 30h.

D. the acquisition of cell growth and metabolic characteristics parameter: by experimental data is analyzed, obtain the mathematic(al) representation of thalline different growing stages bottoms stream rate of acceleration: logarithmic phase is F ₁=0.3043*C+1.0152*F ₂, be F stationary phase ₁=0.1137+1.3702*F ₂

(2) fermentation control experiment

A. the preparation of seed liquor: the preparation seed culture medium, it is divided in 250mL shakes in the bottle, each liquid amount that shakes in the bottle is 100mL, 121 ℃ of sterilization 20min; 1mL Cray Bai Shi bacillus seed liquor is inserted in the seed culture medium, cultivate 12h and obtain seed liquor in 37 ℃, 150rpm shaking table anaerobism;

B. the preparation of fermention medium: preparation fermention medium, 121 ℃ of sterilization 20min;

C. fermentation control experiment: insert in (2) cultivating the seed culture fluid that obtains in (1), inoculum size is 10%, at 7.00 ± 0.02,37 ℃ of pH values, 300rpm, anaerobism (0.1vvm N ₂) carrying out anaerobically fermenting under the condition, the concentration of substrate that adds in the automatic controlling system fermented liquid by bottoms stream is 18 ± 2g/L, ferments to 30h to finish.

D. experimental result: concentration of substrate is controlled to be 18 ± 2g/L all the time in the fermenting process; Not implementing bottoms stream, to add Automatic Control Strategy preceding 1, and 3-PD concentration is 55.25g/L, adds after the Automatic Control Strategy 1 and implement bottoms stream, and 3-PD concentration reaches 82.64g/L, has improved 33.14%.

Claims (1)

1. the bottoms stream based on growth and metabolic characteristics adds fermentative production 1, and the control method of ammediol is characterized in that:

(1) at the thalline logarithmic phase, the wear rate of base consumption speed and alkaline solution and CO ₂Funtcional relationship between the concentration is: F ₁=k ₁C+k ₂F ₂

In the thalline stable growth phase, the funtcional relationship between the consumption of base consumption amount and alkaline solution is:

F ₁＝b+k ₃F ₂；

Wherein: F ₁Be base consumption speed; C is CO ₂Concentration; F ₂Be the alkali lye wear rate; k ₁, k ₂, k ₃, b is constant, its numerical value obtains by the preliminary experiment data fitting;

(2) in the computer with the above-mentioned functional relation substitution fermentation control system that obtains, according to pH and CO ₂Concentration is controlled online fermentation;

Fermentation control system of the present invention comprises fermentor tank, computer, pH electrode, CO ₂Analyser, alkali lye transferpump and substrate transferpump; Automatically control fermentation pH by this system, and alkali speed F mended in record ₂CO ₂The CO that analyser detects ₂Concentration value is sent to computer, relatively more different by analysis CO constantly ₂After the variation of concentration, instruction substrate transferpump adds scheme by homogeneous turbulence not and carries out feed supplement, controls concentration of substrate in the fermented liquid with this.

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