JP4001665B2 - Fermentation product culture management method - Google Patents

Description

[0001]
BACKGROUND OF THE INVENTION
The present invention relates to a method for managing the culture state of lactic acid bacteria in the culture process in producing various fermented products using lactic acid bacteria such as fermented milk. More specifically, the present invention relates to the FT-IR method. The present invention relates to a method for managing a culture process of lactic acid bacteria, and more particularly to a new culture management method for lactic acid bacteria that makes it possible to easily measure the coagulation rate of a culture and accurately predict the culture end point.
[0002]
[Prior art]
Generally, in the production of various products using lactic acid bacteria such as fermented milk, it is important to determine the end point of culture (culture end point) in the management of the culture process.
Conventionally, the management of fermentation, using as an index the concentration of sugar as a substrate, the concentration of lactic acid as a product produced as the fermentation progresses, and the acidity and pH of the fermentation liquor, i.e., The progress of fermentation and the timing of the end of fermentation are performed. In order to obtain these numerical values, conventionally, in the fermentation process, the fermentation liquor is once sampled at an appropriate stage, and then analyzed for each item according to a conventional method using, for example, a neutralization titration method. It was usual to perform quantification.
However, according to such a method, there is a problem that a titration error is large to measure each index value, and it takes a considerable amount of time and manpower, so that the culture process can be managed in a simple and rapid manner. Could not do.
On the other hand, the applicant previously disclosed in Japanese Patent Application Laid-Open No. 9-37770 focusing on the dissociation and non-dissociation type absorbance of lactic acid in the culture solution and the change in absorbance of sugar, and measuring the pH value using FT-IR. A management method is proposed.
[0003]
[Problems to be solved by the invention]
As described above, in view of the above conventional method, conventionally, in the culturing process of fermented milk or the like, the concentration of sugar, the concentration of lactic acid, the acidity, and the pH value as indicators for managing the culturing state of lactic acid bacteria are quickly, simply and accurately. Development of a method for measuring lactic acid bacteria in particular, and development of a method for culture control of a new lactic acid bacterium based on the development of a method for in-line measurement have been desired.
As a result of diligent research aimed at developing such a new culture management method for lactic acid bacteria, the present inventors have used the FT-IR method (Fourier transform infrared spectroscopy), and the infrared absorption of the fermentation broth. The spectrum was measured, and the absorbance of amide I near 1635 cm ⁻¹ and / or the absorbance of amide II near 1540 cm ⁻¹ in the culture was measured over time, and the coagulation of the culture was measured using the rate of change of the absorbance as an index. The rate was measured and it was confirmed that the end point of culture could be determined, and the present invention was completed.
In addition, according to the ATR-FT-IR method, which is a method of measuring the total reflection spectrum (ATR-FT-IR method) among the FT-IR methods, the numerical values of various indices can be measured in-line. In particular, it has been found that the culture end point can be accurately predicted, and that the culture management of lactic acid bacteria can be automated.
[0004]
An object of the present invention is to provide a method for managing the culture process in the production of various products using lactic acid bacteria such as fermented milk.
Another object of the present invention is to provide a new culture management method capable of measuring the coagulation rate of a culture and determining the culture end point in the lactic acid bacteria culture process.
[0005]
[Means for Solving the Problems]
The present invention for solving the aforementioned problems is, in the step of culturing fermentation products using lactic acid bacteria, using the FT-IR method, 1635 cm culture ^- absorbance of ¹ amide I and / or 1540 cm ^{- 1} absorbance of the amide II Is measured over time, and the culture of the fermented product is managed using the rate of change in absorbance as an index.
Furthermore, the onset bright, in the step of culturing the above fermentation product, using the FT-IR method, 1635 cm culture ^{- 1} absorbance of the amide I and / or 1540 cm ^{- 1} of the absorbance of the amide II over time The culture management method for a fermented product according to claim 1, wherein the measurement is performed, the rate of change in absorbance is used as an index, the solidification rate of the culture is measured, and the culture end point is predicted / determined.
In the present invention, as the FT-IR method, it is a preferred embodiment to measure the absorbance using the ATR-FT-IR method. In the measurement of absorbance by the FT-IR method, the absorbance of amide I at 1635 cm ⁻¹ and / or the absorbance of amide II at 1540 cm ^{−1 in} the culture is measured over time. In particular, when it is included in 1635 cm ⁻¹ or 1540 cm ⁻¹ , even the vicinity thereof is included in the scope of the present invention.
[0006]
DETAILED DESCRIPTION OF THE INVENTION
Next, the present invention will be described in more detail.
Although this invention is applied in the culture | cultivation process of various products using lactic acid bacteria, if this invention is a fermentation and culture | cultivation process containing lactic acid bacteria, it will be object regardless of the kind of the product. Hereinafter, culture of fermented milk will be described as an example.
In the production process of various products using lactic acid bacteria, the coagulation phenomenon of milk caused by the acid produced by lactic acid bacteria is observed as a change in absorbance around 1635 cm ⁻¹ and / or 1540 cm ⁻¹ . These two absorption bands are known as absorption of the amide of the protein molecule, and 1635 cm ⁻¹ is called amide I which is C═O stretching vibration, and 1540 cm ⁻¹ is called amide II of NH deformation vibration.
In the fermentation of fermented milk, the relationship between culture time, pH, and absorbance at 1635 cm ⁻¹ and 1540 cm ⁻¹ is shown in FIG. Absorbance was shown as the amount of change using the spectrum at 0 hours of culture as a reference. It was confirmed that the absorbance changed suddenly around pH 5.1 where milk coagulation began.
When the rate of change in absorbance is plotted against the incubation time, it is as shown in FIG. The rate of change of the two absorbances is mainly a dependent function of the milk protein clotting rate. In other words, it can be said that the rate of change in absorbance represents the rate of fermentation. Therefore, the culture end point can be determined from the rate of change in absorbance.
[0007]
Specifically, the culture end point can be determined by using either one or both of the following two functional expressions.
As the absorbance to be used, either one of the two absorption bands described above may be used, or both may be used, or the sum of the two absorbances may be used. However, if using low wavenumber precision spectrometer, when performing an operation which gives a change in absorbance of water, the absorbance of the amide I of 1635 cm ^-1, so influenced by the absorption of water 1650 cm ^-1 It is desirable to use only 1540 cm ⁻¹ .
t = f (v) ─────── (1)
t = f (s) ─────── (2)
Time from when the rate of change in absorbance reaches the maximum until the end of culture: t
Maximum change rate of absorbance: v
Area surrounded by the locus of change rate of absorbance and the time axis: s
Since the function f varies depending on the culture system, it is necessary to perform culture in advance and set it.
The present invention makes it possible to quantify the coagulation phenomenon of milk caused by the acid produced by lactic acid bacteria as the rate of change of the absorbance, measure the coagulation rate, and determine the culture end point. The present invention can be applied as a culture management method in a culture system of various products using lactic acid bacteria such as fermented milk regardless of the type of the product.
[0008]
【Example】
Hereinafter, the present invention will be specifically described based on examples.
Example 1
Streptococcus thermophilus (Streptococcus thermophilus) YIT2001 was cultured in a 20% strength non-fat dry milk aqueous solution at 37 ° C. with gentle stirring. Infrared spectra were measured using Fourier Transform Infrared Spectrophotometer FTS-65A (Bio-Rad Digilab) and FT-IR / ATR probe model DPR-210 (S) (Axion Analytical Inc. ) And continuous automatic in-line measurement of infrared absorption spectrum in the range of 1900 to 900 cm ⁻¹ . Data accumulation and analysis were examined using a computer (IBM-PC).
It is known that the culture end point changes depending on the activity and the amount of the starter. In order to derive the above-mentioned relational expression (1), several batch cultures were carried out while changing the activity of the starter. As a result, the following relational expression was obtained.
t = 0.45 / v ─────── (3)
Time from when the rate of change in absorbance reaches the maximum until the end of culture: t
Maximum change rate of absorbance: v
[0009]
As an example of culture, a case of culture A having high starter activity and culture B having low activity will be described. When the titration acidity value of 20 was reached, the culture time of culture A was 20 hours and culture B was 25 hours. The culture time course of the sum of the absorbance at 1635 cm ^-1 and 1540 cm ^-1 in the case shown in the upper part of FIG. The lower part of FIG. 3 shows the rate of change in absorbance obtained by differentiating the time-dependent change in the culture with time in the sum of absorbances in FIG. The maximum value of the change rate of absorbance and the culture time when the maximum value are reached are −0.041 (Ab./h) and 9 h for culture A, −0.032 (Ab./h) for culture B, 10 hours. Substituting this maximum absorbance change rate into equation (3) yields 11 and 15, respectively.
Therefore, the total culture time was 20 hours for culture A at 11 + 9 and 25 hours for culture B at 15 + 10, which was consistent with the culture end point based on the titrated acidity value, indicating that the culture end point can be determined by the method of the present invention. .
[0010]
Example 2
Bibidobacterium breve YIT4065 was cultured in a 20% strength whole fat milk powder aqueous solution at 37 ° C. with gentle stirring. Infrared spectra were measured using Fourier Transform Infrared Spectrophotometer FTS-65A (Bio-Rad Digilab) and FT-IR / ATR probe model DPR-210 (S) (Axion Analytical Inc. ) And continuous automatic in-line measurement of the infrared absorption spectrum was performed in the range of 1900 to 900 cm ⁻¹ . Data accumulation and analysis were examined using a computer (IBM-PC).
When a titration acidity value of 20 was reached, the end point was reached after 48 hours of culture. Shows the rate of change in culture aging and absorbance of the sum of absorbance at 1635 cm ^-1 and 1540 cm ^-1 in this case is shown in FIG. 4. The absorbance change rate is 0.0175 (Ab./h) which is the maximum value after 27.5 hours from the start of the culture.
Therefore, in this culture system, the end point of culture could be determined by the following relational expression.
t = 0.35 / v
Time from when the rate of change in absorbance reaches the maximum until the end of culture: t
Maximum change rate of absorbance: v
[0011]
As described above, in the present invention, in the culturing process of various products using lactic acid bacteria such as fermented milk, it is necessary to monitor the culture state by pre-processing the measurement sample by using in-line infrared spectroscopy. In addition, since it can be monitored in a closed system, it has been shown that the risk of contamination is small, and it can be performed easily, quickly and fully automatically. Therefore, it has been clarified that the method of the present invention is extremely effective in systems such as lactic acid bacteria beverages and fermented milk that have conventionally been monitored for culture processes with titrated acidity.
[0012]
【The invention's effect】
As described above in detail, the present invention uses the FT-IR method in the culturing process of various products using lactic acid bacteria such as fermented milk, and the absorbance of amide I in the vicinity of 1635 cm ⁻¹ and / or 1540 cm. ^-1 , measuring the absorbance of amide II in the vicinity of time, measuring the coagulation rate of the culture using the rate of change of the absorbance as an index, and determining the end point of the culture, According to the present invention, (1) it is not necessary to pre-process the measurement sample, (2) the culture state of the fermentation product can be monitored in a closed system, and (3) the fermentation product culture process Special effects such as simple measurement of the coagulation rate of the culture and accurate prediction / determination of the culture end point, (4) easy and rapid culture management of the fermentation product by in-line Gain It is.
[Brief description of the drawings]
[1] in the culturing step of the fermented milk of Example 1, showing the incubation time and the pH value, and the absorbance of the relationship between 1635 cm ^-1 and 1540 cm ^-1.
FIG. 2 shows the results of plotting the rate of change in absorbance against the culture time in Example 1.
In [3] Example 1 shows the culture time course of the sum of the absorbance at 1635 cm ^-1 and 1540 cm ^-1.
FIG. 4 shows the rate of change in absorbance obtained by differentiating the change in culture with time in the sum of absorbance in FIG. 3 with respect to time.

Claims (2)

In the step of culturing fermentation products using lactic acid bacteria, using the FT-IR method, 1635 cm culture ^{- 1} amide I of absorbance and / or 1540 cm ^- over time by measuring the absorbance of ¹ amide II, the absorbance the rate of change as an index, the solidification rate of the culture was measured, originating酵製article method of culturing manage you and determining a culture endpoint. As FT-IR method, ATR-FT-IR method using, characterized rows Ukoto absorbance measurements, culture management method of fermentation product of claim 1 Symbol placement.

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