JPH03210467A - Apparatus for simultaneously and automatically measuring acidity of fermented milk and concentration of l-lactic acid - Google Patents

Abstract

PURPOSE:To simultaneously and automatically measure the acidity of fermented milk and the concn. of L-lactic acid by mounting a sample flow passage, an acidity measuring cell, a buffer solution supply passage, an L-lactic acid concn. measuring cell and an operational processor. CONSTITUTION:The acidity measuring cell 13 provided on the way of a sample flow passage 12 has volume larger than that of an injected sample solution 1 and can dilute the sample solution 1 and a carrier liquid 2 in a predetermined ratio. A buffer solution supply passage 17 supplying a pH buffer solution meets with the downstream part of the acidity measuring cell 13 of the sample flow passage 12 and, on the downstream side of this this confluent part, an L-lactic acid concn. measuring cell 19 is arranged through a mixer 18. An operational processor 23 calculates the acidity of the sample solution 1 from the measured conductivity of the sample solution and calculates the concn. of L-lactic acid in the sample solution from the measured concn. of hydrogen peroxide therein.

Description

DETAILED DESCRIPTION OF THE INVENTION [Field of Industrial Application] The present invention relates to an apparatus for simultaneously measuring the acidity and L-lactic acid concentration of fermented milk such as yogurt.

[Conventional technology]

Fermented milk such as yogurt is produced by fermenting formula or skim milk powder with lactic acid bacteria, but it is essential to measure acidity and L-lactic acid concentration to control the fermentation process and quality.

The measurement of acidity and L-lactic acid concentration is important because the quality of fermented dairy products is greatly influenced by acidity and L-lactic acid concentration. In other words, the acidity of fermented milk depends on the concentration of citric acid contained in the milk formula and lactic acid produced during fermentation, and is greatly involved in the sourness of fermented milk products. In addition, the lactic acid contained in fermented milk has two optical isomers, L-type and D-type.
Although L-lactic acid is useful for the human body, it is known that the compatibility between the two differs depending on the type and proportion of lactic acid bacteria used for fermentation and the fermentation process.

The acidity of fermented milk is determined by the lactic acid concentration (defined as a candle) calculated from the amount of I M NaOH (■) required to neutralize 100% of fermented milk using a food analysis method. It was measured by neutralization titration or potentiometric titration using the potentiometric method.

L-lactic acid concentration was determined by manual analysis of L-lactate dehydrogenase and L-lactate concentration.
- Decompose L-lactic acid using lactate oxidase and measure by absorbance measurement, or measure using a hydrogen peroxide electrode equipped with an immobilized enzyme membrane in which L-lactate oxidase is immobilized on a plastic membrane, etc. It had been.

That is, the hydrogen peroxide concentration generated by the following enzymatic reaction was measured, and the L-lactic acid concentration was determined from this; It is necessary to take a sample from a fermented milk cup in quantitative quantities using a pipette or balance, and manually measure the acidity and L-lactic acid concentration separately, which not only is a complicated operation and requires a lot of time and manpower. There were problems such as being susceptible to human error. In addition, since the acidity and L-lactic acid concentration of fermented milk change with fermentation time, it is desirable to measure and control the acidity and L-lactic acid concentration of samples taken at about the same time. This required more manpower, which caused an increase in management costs.

[Problem to be solved by the invention]

In view of such conventional circumstances, the present invention has been developed to improve the acidity of fermented milk and L-
It is an object of the present invention to provide a device that can measure lactic acid concentration almost simultaneously and automatically.

[Means to solve the problem]

In order to achieve the above objectives, the acidity and L-
The simultaneous automatic measurement device for lactic acid concentration is a flow analyzer that includes a sample liquid supply path for injecting the fermented milk sample liquid into the carrier liquid, a sample flow path for transferring the sample liquid injected into the carrier liquid, and a sample flow path. It is set up in the middle of
An acidity measurement cell with a larger capacity than the injected sample liquid equipped with a stirring device and a conductivity electrode, a buffer supply path for injecting a pH buffer into the sample flow path downstream of the acidity measurement cell, and a confluence of the buffer supply path. an L-lactic acid concentration measuring cell provided in the middle of the sample flow path after passing through a mixer downstream from the point and equipped with a hydrogen peroxide electrode having an enzyme membrane with immobilized L-lactic acid oxidase;
and an arithmetic processing device connected to the conductivity electrode and the hydrogen peroxide electrode, which calculates the acidity of the sample liquid from the measured conductivity and calculates the L-lactic acid concentration of the sample liquid from the measured hydrogen peroxide concentration. It is characterized by

Further, if a switching valve is provided in the middle of the sample liquid supply path, and the calibration liquid supply path is switchably connected to the sample liquid supply path by the switching valve, it is possible to automatically calibrate the apparatus before measurement.

[Effect]

In the present invention, by adopting the flow analysis method and making it into an apparatus, while rapidly collecting and transferring a certain amount of sample liquid, the acidity is first measured using a conductivity electrode using the dilution conductivity method, and then The L-lactic acid concentration is measured using a hydrogen peroxide electrode having an enzyme membrane with immobilized L-lactate oxidase. Therefore, it is possible to carry out two measurements of the acidity and L-lactic acid concentration of fermented milk continuously, almost simultaneously, and automatically.

In particular, regarding the measurement of acidity, we followed the idea of the general dilution conductivity method and focused on the fact that a linear correlation between acidity and conductivity can be obtained by diluting fermented milk appropriately. We have developed an optimal method for flow analysis in which a correlation equation is calculated between the acidity determined by the method and the conductivity determined by the conductivity electrode, and then the acidity is determined from the conductivity measured for the sample and the correlation equation. , adopted in the device of the present invention.

However, since the correlation equation between acidity and electrical conductivity differs depending on the type of fermented milk, it is necessary to obtain a correlation equation for each individual fermented milk.

〔Example〕

A specific example of the device of the present invention will be explained with reference to FIG.

This device includes a sample liquid supply path 9 in which a sample liquid 1 of fermented milk is transferred via a switching valve 11 by a pump 5, and injected into a carrier liquid 2 by an injector 10, and an injector 10 injected into the carrier liquid 2 transferred by a pump 6.
Sample flow M12. ! : Equipped with: Further, a calibration liquid supply path 22 is connected to the switching valve 11, and by switching the switching valve 11, the calibration liquid 3 can be supplied to the injector 10 through a part of the sample liquid supply path 9.

An acidity measuring cell 13 is provided in the middle of the sample flow path 12, and the acidity measuring cell 13 has a larger capacity than the sample liquid 1 injected by the injector 10, and adjusts the sample liquid 1 to a predetermined ratio with the carrier liquid 2. It can be diluted. The acidity measuring cell 13 has a dip-type conductivity electrode 14 using a PT electrode, and is equipped with a stirring device consisting of a stirring bar 15 and a magnetic stirrer 16. Further, downstream of the acidity measurement cell 13 in the sample flow path 12, a buffer supply path 17 for supplying pH buffer 4 by a pump 8 joins, and downstream from the confluence, a mixer 18 passes through the L-
A lactic acid concentration measuring cell 19 is installed. The L-lactic acid concentration measuring cell 19 is equipped with a hydrogen peroxide electrode 21 using PT as an anode and Ag as a cathode, and the hydrogen peroxide electrode 21 is an immobilized enzyme membrane 2 in which L-lactate oxidase is immobilized on a plastic membrane.
The sample liquid 1 is contacted through the sample liquid 1, and the HO concentration generated when L-lactic acid is oxidized by an enzymatic reaction is measured.

The conductivity electrode 14 of the acidity measurement cell 13 and the hydrogen peroxide electrode 21 of the L-lactic acid concentration measurement cell 19 are respectively connected to a processing unit 23, and the sample liquid is compared with the conductivity and hydrogen peroxide concentration at the time of calibration. The acidity is calculated from the measured conductivity of sample liquid 1, and the L-lactic acid concentration is calculated from the measured hydrogen peroxide concentration of sample liquid 1.

Example 1 Using the measuring device shown in FIG. 1, the acidity output and L-lactic acid concentration output of an L-lactic acid solution were determined.

The lactic acid solution as sample solution 1 contains L-lactic acid 5. O
Og was dissolved in 100% of ion-exchanged water, diluted as appropriate, and adjusted to various concentrations. Carrier liquid 2 is ion-exchanged water with an electrical conductivity of 1μ3/c or less, and pH buffer 4.
For the calibration solution 3, a 1 mol/l'J acid solution with a pH of 7.5 containing o, i% benzoic acid and 1 no//, / KCI was used, and a 0.5% L-lactic acid solution was used as the calibration solution 3.

First, the acidity and conductivity of L-lactic acid were measured using a conventional titration method and a conductivity electrode, respectively, a correlation equation between the two was obtained, and the acidity could be calculated from the measured conductivity using the arithmetic processing unit 23. .

Next, the sample liquid 1 supplied to the injector 10 (capacity 20 μl) through the sample liquid supply path 9 is transferred to the injector 10.
The samples were rotated and cut out, sandwiched between the carrier liquid 2 filling the sample channel 12, and supplied to the acidity measurement cell 13' with a flow rate of 10 mm/min. The sample liquid 1 was diluted with the carrier liquid 2 in the acidity measurement cell 13, and the conductivity of the diluted sample liquid was measured by the conductivity electrode 14. Next, the diluted sample liquid 1 is added to the conductivity measurement cell 1.
3 and from the buffer supply section 17 at a flow rate of 1 milnim.
mixed with pH buffer 4 which is continuously supplied at L
- The HO concentration was measured by the hydrogen peroxide electrode 21 in the lactic acid concentration measurement cell 19 (capacity 100μ).
! Prior to this measurement, calibration using calibration solution 3 was performed.

Figure 2 shows the response curve of the acidity output and the L-lactic acid concentration output for the sample liquid obtained by two consecutive measurements by sampling the sample liquid 1 twice with the injector 10. Yes, beak ASB, C, and DSE have concentrations of 0.5%, 0.4%, 0°3%, 0.2%, and 0, respectively.
゜It corresponds to the acidity output of 1% L-lactic acid solution, and also the beak F
% GSH1 engineering and J have concentrations of 0.5%, 0.4%, and 0, respectively.
．． It corresponds to the L-lactic acid concentration output of 3%, 0.2%, and 0.1% L-lactic acid solutions.

The specific acidity and L-lactic acid concentration of the sample liquid can be determined from the peak height of each of these outputs, and it is important to note that each peak height is well proportional to the pre-adjusted L-lactic acid concentration of the sample liquid, and It can be seen from FIG. 2 that the measurement of acidity and L-lactic acid concentration can be carried out for each sample solution in a short time of about 5 degrees.

Example 2 Using the measuring device shown in FIG. 1, fermented milk was used as a sample solution at fixed intervals after the start of fermentation, and its acidity and L-lactic acid concentration were measured.

In addition, for fermentation, La\tobaci//us is used as a bacterial cell.
Bu/ga-ricus and 5tsrptocoacus
Thermophilua symbiotic cells were used, and a solution of 10.51% skim milk powder and 9.0% sugar dissolved in tap water was used as a culture medium.

First, the acidity and conductivity of the fermented milk at various concentrations were measured using a potentiometric titration device and a conductivity electrode, and the results showed that the correlation coefficient between acidity and conductivity was 0.997. The correlation shown in the figure was obtained. The correlation equation between acidity (X) and conductivity (Y) obtained from Figure 3 is Y -69,
It was 25x + 63.18.

A sample liquid of fermented milk was analyzed under the same measurement conditions and operations as in Example 1, and the acidity (X) was determined using the above correlation formula from the measured electrical conductivity, and the measured L-lactic acid concentration and fermentation Figure 4 shows the pH relationship determined by the pH electrode attached to the tank.

As can be seen from FIG. 4, as the fermentation time progresses, the acidity increases and the pH decreases. The L-bango concentration increases until about 3 hours after the start of fermentation, but remains approximately constant thereafter. The reason why the acidity further increases even after the L-lactic acid concentration becomes almost constant is due to the production of D-lactic acid.

〔Effect of the invention〕

According to the present invention, a sample liquid can be collected from fermented milk and its acidity and L-lactic acid concentration can be measured almost simultaneously and automatically within an extremely short period of time without requiring any pretreatment.

Therefore, the present invention is very effective for real-time monitoring of the fermentation process in lactic acid fermentation for yogurt production, for example, and for quality control of fermented dairy products.

[Brief explanation of drawings]

FIG. 1 is a schematic diagram showing a specific example of the apparatus of the present invention. Fig. 2 is a response curve of output of acidity and L-lactic acid concentration obtained in Example 1, Fig. 3 is a graph showing the correlation between acidity and conductivity obtained in Example 2, and Fig. 4 is a graph showing the correlation between acidity and conductivity obtained in Example 2. is a graph showing the relationship between acidity, L-lactic acid concentration, and pH obtained in Example 2. 1... Sample liquid 2... Carrier liquid 3... Calibration solution 4... pH buffer solution 5.6.7.8... Pump 9... Sample liquid supply path 10... Injector 11... Switching valve 12...
Sample channel 13... Acidity measurement cell 14... Conductivity electrode 15... Stirrer 16... Magnetic stirrer 17... Buffer supply channel 18... Mixer 19... L-
Lactic acid concentration measurement cell 20...immobilized enzyme membrane 21...hydrogen peroxide electrode 22
... Calibration liquid supply path 23 ... Arithmetic processing unit Fig. 2 ∆ time (minutes) Fig. 3 Revision (%)

Claims (2)

[Claims] (1) In the flow analysis device, a sample liquid supply path for injecting the sample liquid of fermented milk into the carrier liquid, a sample flow path for transferring the sample liquid injected into the carrier liquid, and a sample flow path provided in the middle of the sample flow path, An acidity measurement cell with a larger capacity than the injected sample liquid equipped with a stirring device and a conductivity electrode, a buffer supply path for injecting a pH buffer into the sample flow path downstream of the acidity measurement cell, and a confluence of the buffer supply path. An L-lactic acid concentration measuring cell is provided in the sample flow path after passing through a mixer downstream from the point, and is equipped with a hydrogen peroxide electrode having an enzyme membrane immobilized with L-lactate oxidase, Fermented milk characterized by comprising: an arithmetic processing device connected to a hydrogen oxide electrode, which calculates the acidity of the sample liquid from the measured conductivity and calculates the L-lactic acid concentration of the sample liquid from the measured hydrogen peroxide concentration. Simultaneous automatic measurement device for acidity and L-lactic acid concentration. (2) A switching valve installed in the middle of the sample liquid supply path,
The device for simultaneous automatic measurement of fermented milk acidity and L-lactic acid concentration according to claim (1), further comprising a calibration solution supply path switchably connected to the sample solution supply path by a switching valve. .