KR840000014B1 - Automation of glutamic acid fermentation process using exit gas analysis - Google Patents

Abstract

Glutamic acid fermentation process and measurement of cell concentration were automatized by exit gas analysis. Thus, Corynebacterium sp. was inoculated into 13L medium contg. molasses 2.6L, K3PO4 13.0g, MgSO4 5.2g, MnSo4 0.13g, Fe2So4 0.13g, Corn steep liquor 19.5g, and thiamine 2600μg and incubated at 31≰C in a 30L fermentor. Parts of the exitgas, according to the cell growth, were passed through CO2-analyzer and O2-analyzer. By the measurement of the produced O2 and CO2, penicillins or surfactants were automatically supplied into the fermentor.

Description

Automated glutamic acid fermentation process by fermentation exhaust gas analysis

1 is a glutamic acid fermentation process automation system.

2 is the relationship between the CO ₂ production rate and cell concentration.

The present invention relates to a method for automating a penicillin (or surfactant) administration process which is essential for an industrial glutamic acid fermentation process. Specifically, the present invention relates to a method for automating penicillin (or surfactant) administration that is important for glutamic acid fermentation by indirect measurement of growth and proliferation of microorganisms obtained from fermentation exhaust gas analysis.

Glutamic acid is industrially used to propagate microorganisms such as Corynebacterium, Brevibacterium, Microbacterium, and Micrococcus until the optimal cell weight is reached before penicillin (or surfactant ) Is produced by regulating cellular bermeabilit of microorganisms and discharging glutamic acid accumulated inside microorganisms out of microorganisms.

At this time, the molasses medium used industrially contains a large amount of growth factors, biotin and minerals, and the degree of growth and proliferation of microorganisms varies greatly according to the precision of timely and proper dose of penicillin (or surfactant). Therefore, glutamic acid production yield is also greatly affected.

In such a timely and proper dose administration process of penicillin (or surfactant), a sample of a fermentation tank is directly collected, and after measuring the extent of growth and proliferation of microorganisms using an analyzer such as a spectrophotometer, It has been done manually at the discretion of the technician.

However, the management of major processes by this method has been acting as a process instability due to many inconveniences in process and loss of work loss, and in fact, due to rapid cell growth at the time of penicillin (or surfactant) administration. Therefore, the difficulty of accurate and continuous cell determination in automating the entire process of glutamic acid fermentation was inevitable.

Thus, the present inventors this using a process computer to automate the conventional manual administration process, connect the glutamic acid fermentation exhaust gas line (on-line), a result of studying various Fermentation variables CO ₂ production rate (CER) The specific correlation between and the cell concentration could be derived, and from this result, the timely and proper dose of penicillin (or surfactant) could be automated to stabilize the process.

The content of the present invention will be described in detail as follows.

Microorganisms that produce glutamic acid multiply by maintaining appropriate temperature and pH in fermentation broth containing carbon source, nitrogen source, oxygen, and minerals, and produce water and carbon dioxide.

After removing the water contained in the fermented yeast exhaust gas, the CO ₂ production rate (CER) is calculated from the following equation (I) using an oxygen meter and a carbon dioxide meter.

Where F is the number of moles of air supplied to the fermenter (mole / min), V is the amount of fermentation broth (L), P _T is the voltage, P _O2 and P _CO2 are the partial pressures of oxygen and carbon dioxide contained in the exhaust gas, respectively. .

As described above, CERs that can be measured continuously from the beginning of fermentation have a linear relationship with the concentrations of the cells, as shown in FIG. 2. Therefore, the concentration of the cells can be estimated by continuously measuring the CERs.

Using the relationship between CER and cell concentration, penicillin (or surfactant) can be easily administered in a timely manner using a microprocessor or a process computer when the required cell concentration is reached during the fermentation process.

In addition, the oxygen measuring device and the carbon dioxide measuring device as described above are required for the measurement of the CER, but even if only the oxygen (or carbon dioxide) concentration in the exhaust gas is measured, the remaining concentration (carbon dioxide concentration) is estimated using Equation (II) and used in the present invention. Can be.

P _CO2 = O.2lxP _T -P _O2 (II)

Table 1 shows the ratio of the oxygen concentration in the exhaust gas measured using the oxygen meter and the oxygen concentration calculated using the formula (II) from the carbon dioxide concentration measured by the carbon dioxide meter.

TABLE 1

Ratio of measured value and calculated value of oxygen concentration

As can be seen from Table 1, the relative error range between the actual measured value and the calculated value obtained from the calculation is within ± 3%, so the CER can be calculated and used industrially simply by the oxygen measuring device (or carbon dioxide measuring device).

Example 1

Using a 30 ℓ small fermenter, Corynebacterium spp. Was inoculated into industrial molasses containing Mg ⁺⁺ , Mn ⁺⁺ , Fe ⁺⁺ , PO ₄ ^-3 and air flow rate 1VVM, temperature 31 ℃, pH7.6 The mixture was grown at agitation speed of 400 rpm.

The composition of the medium used at this time is as follows.

Part of the exhaust gas generated along with the growth of the cells is passed through the dryer to the carbon dioxide meter and the oxygen meter as shown in FIG. 1, and the concentration of each cell is measured. The current generated by the meter is amplified by the signal conditioner. It is changed to voltage within ± 5V suitable for computer input and input to process computer through A / D converter. As the result of analysis and fermentation is progressed, the value of CER is continuously calculated and stored in process computer. When it arrives, the solenoid valve is operated by the output voltage from the D / A converter, so that penicillin (or surfactant) is automatically administered.

Table 2 shows the CER measured using the above-described formula during fermentation, and the error between the measured cell concentration and the measured cell concentration.

Here, the dry cell weight is the concentration of only cells excluding the solid component of the medium, and centrifuged at 3000 rpm for 15 minutes to wash the precipitated cells twice, followed by recentrifugation and drying at 105 ° C. for 20 hours.

The solid component of the medium was measured by phagocytosis as a blank.

TABLE 2

Cell mass calculated from CER and cell weight measured

At this time, the CER per unit cell calculated by computer was 0.473mmole / g cell / min, and the dry cell weight calculated based on this showed an error within ± 0.15g / ℓ with the measured dry cell weight.

Example 2

Under the same conditions as in Example 1, the air flow rate was increased to 0.5 VVM.

Table 3 shows the results of the comparison between the dry cell mass and indirectly calculated cell concentrations indirectly calculated by oxygen and carbon dioxide analysis in the exhaust gas in the same manner as in Table 2.

TABLE 3

Cell mass calculated from CER and contrast with cell mass measured

As shown in Table 3, even though the air flow rate was changed, the calculated dry cell mass was found to be in the error range of ± 0.1 g / l and the dry cell mass measured.

Therefore, indirect cell concentration measurement using CER according to the flow rate of air was possible, and the calculated CER per unit cell was 0.564mmole / gcell / min.

Example 3

Under the same conditions as in Example 1, growth was performed by varying the concentration per second, the temperature, and the stirring speed. Table 4 shows the difference between the calculated dry cell mass and the measured dry cell mass as shown in Tables 1 and 2, and shows the calculated CER per unit cell in each case.

More specifically, the value of CER per unit cell may vary depending on fermentation conditions or fermenter conditions, but the measurement of cell concentration according to the present invention shows an error of ± 0.1 g / L, which is a perfect penicillin (or surfactant). By automating the administration, it was possible to dramatically stabilize the process.

TABLE 4

Cell concentration measurement results according to fermentation conditions

Claims (3)

In the glutamic acid fermentation process using molasses as a carbon source, glutamic acid fermentation process by fermentation exhaust gas analysis is characterized in that the fermentation exhaust gas is analyzed by carbon dioxide and oxygen measuring instrument and the cell concentration is measured using the carbon dioxide production rate (CER). Automation method. The method for measuring cell concentration by fermentation exhaust gas analysis using only a carbon dioxide measuring device according to claim 1. The method for measuring cell concentration by fermentation exhaust gas analysis using only an oxygen measuring device according to claim 1.

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