Chapter 6 Changes Of Some Metabolites And PH
Oct 31, 2024
Chapter 6 Changes in some metabolites and pH
The metabolic changes of microorganisms during fermentation are very complex, and the metabolic mechanism is still unclear. Therefore, it is very meaningful to clarify the changes in the main metabolites of microbial enzymes during fermentation. This study took apple enzymes as the research object and tracked the changes in total acid, total sugar, alcohol content, and pH within 3 months during the fermentation process. It provides a certain data basis and technical basis for further solving the problems related to product quality such as the mechanism of the fermentation process, the proportion control of bacterial strains, and the research of functional ingredients.
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6.1 Materials and Methods
6.1.1 Materials
(1) Experimental raw materials
The raw materials required for the experiment are the same as 5.1.1 (1).
(2) Experimental instruments
The instruments required for the experiment are the same as 3.1.1 (2).
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6.1.2 Methods
(1) Determination of total acid content[71]
Direct titration method
(2) Determination of total sugar content[72]
Take 0.5 mL of fermentation broth and place it in a 50 mL volumetric flask. Add 2 mL of 6 mol/L hydrochloric acid solution and boil it in a water bath for 15 min. After cooling, add 1.8 mL of 6 mol/L sodium hydroxide solution and make up the volume. Take 2 mL of the liquid in the volumetric flask and place it in a 25 mL volumetric flask. Add 1.5 mL of DNS reagent and boil it in a water bath for 5 min. After cooling, make up the volume. Measure the absorbance at 520 nm. Calculate the total sugar content according to the linear equation of the standard curve.
(3) Determination of alcohol content[73]
The alcohol content was determined by the density bottle method.
(4) Change of pH
The pH value was directly measured using a pH meter.
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6.2 Results and Discussion
6.2.1 Total Acid
This study used the acid-base titration method to determine the total acid content. Apple enzyme was used as the research object. From the beginning of fermentation, the total acid content of apple enzyme was measured every 15 days, and the change curve of the total acid content was obtained, as shown in Figure 6.1.
6.2 Results and discussion
6.2.1 Total acid
This study used acid-base titration to determine the total acid content. Apple enzyme was used as the research object. From the beginning of fermentation, the total acid content of apple enzyme was measured every 15 days, and the variation curve of total acid content was obtained, as shown in Fig. 6.1.
Fig.6.1 The variation curves of total acid content
It can be seen from Fig. 6.1 that within 30 days of fermentation, the total acid content of the enzyme solution showed an upward trend, but after 30 days, the total acid content decreased and finally stabilized. In general, the total acid content accumulated rapidly within the first 15 days of fermentation, and the upward trend tended to be flat after 15 days. Compared with the case without adding bacteria, the total acid content was 24.14% higher on the 90th day of fermentation.
6.2.2 Total sugar
(1) According to the experimental results, the standard curve of glucose is shown in Fig.6.2.
Fig.6.2 The standard curve of glucose
As shown in Fig.6.2, the standard curve equation is y=0.6907x-0.0386, R2=0.9989. Where x is the glucose concentration (mg/mL) and y is the absorbance.
(2) Total sugar content
From the beginning of fermentation, the total sugar content of apple enzyme was measured every 15 days, and the change curve of total sugar content was obtained, as shown in Fig.6.3.
It can be seen from Figure 6.3 that as the fermentation time increases, the total sugar content of apple enzymes decreases significantly in the first 15 days of fermentation. After 15 days, the total sugar content gradually rises. It is speculated that the reason may be that in the first 15 days, the massive growth of microorganisms caused a large amount of sugar substances. After 15 days, the microorganisms decomposed the apple enzyme and produced sugary metabolites. The products fused with the enzyme, making the total sugar content of the enzyme gradually increase. The total sugar content decreased by 12.5% on the 90th day of fermentation with added strains compared to without added strains.
6.2.3 Alcohol Content
From the beginning of fermentation, the alcohol content of apple enzyme was measured every 15 days, and the change curve of alcohol content during fermentation was obtained, as shown in Figure 6.4.
As shown in Figure 6.4, in the first 15 days of fermentation, a large amount of alcohol was produced, so the alcohol content of enzyme was very high. After about 75 days, the alcohol content of enzyme reached below 1%vol. At this time, there was almost no alcohol taste in the taste of enzyme. Compared with the case without adding bacteria, the alcohol content of the case with added bacteria was 16.67% higher on the 90th day of fermentation.
6.2.4 pH variation law
The pH variation law of apple enzyme during fermentation is shown in Fig. 6.5.
Fig.6.5 The variation curves of pH
As shown in Fig. 6.5, the pH value of apple enzyme in the experimental group dropped rapidly from 5.4 to 3.8 within 15 days, while the pH value of the control group dropped from 5.4 to 4.0, and then tended to be moderate. During the experiment, we also found that a large number of bubbles were generated in the bottle from the 4th to the 8th day of enzyme production, indicating that the first 15 days were a stage of large-scale microbial reproduction and very active metabolism. After 15 days, it entered the slow fermentation stage, and the increase in organic acid concentration caused the pH to drop[45]. The growth and metabolism of the bacteria produced a large amount of carbon dioxide, which appeared in the water in the form of bubbles. Due to the instability of the bubbles, the pH of the enzyme solution was also unstable, and it fell and rose at the same time. Compared with the case without adding bacteria, the pH of the case with added bacteria dropped by 5% on the 90th day of fermentation.
6.3 Summary of this chapter
This chapter studies the changes in metabolites and pH during enzyme fermentation. Studies have shown that within the first 15 days of enzyme fermentation, microorganisms grow and reproduce fastest, and total acid, total sugar, alcohol content, and pH change the most. Within 30 days of fermentation, the total acid content of the enzyme solution shows an upward trend, but after 30 days, the total acid content decreases and finally stabilizes. Within the first 15 days of fermentation, the total acid content accumulates rapidly, the total sugar content decreases significantly, a large amount of alcohol is produced, and the pH value drops sharply.
After 15 days, it enters the slow fermentation stage. Compared with the case without adding strains, on the 90th day of fermentation, the total acid content of the case with added strains was 24.14% higher, the total sugar content was 12.5% lower, the alcohol content was 16.67% higher, and the pH was 5% lower.
