A coupling method for producing fructo-oligosaccharides and feed yeast by using moringa oleifera water extract

Abstract

The application discloses a coupling method for producing fructo-oligosaccharide and feed yeast by using water extract of moringa oleifera branch, and belongs to the field of high-value utilization of agricultural and forestry by-products and biological processing technology. The method comprises the following steps: water extraction is performed on the moringa oleifera branch to obtain a water extract of the moringa oleifera branch containing fermentable sugar; then, a fructosyltransferase is used for enzymatic reaction to obtain an enzyme cut solution containing fructo-oligosaccharide; then, the enzyme cut solution is inoculated with saccharomyces cerevisiae for fermentation; after the fermentation is completed, the fermentation liquid is separated into solid and liquid, the supernatant is a fructo-oligosaccharide solution, and the part of the bacteria is centrifuged, washed and dried to obtain feed yeast. The method realizes efficient utilization of natural sugar components in the moringa oleifera branch and the synergistic preparation of fructo-oligosaccharide and feed yeast, and has the advantages of wide raw material sources, mild process, high added value and high resource utilization rate.

Description

Technical Field

[0001] This invention belongs to the field of high-value utilization and bioprocessing technology of agricultural and forestry by-products, specifically involving a method for producing fructooligosaccharides and feed yeast by coupling the natural sugar components in the water extract of Moringa branches through fructosyltransferase catalysis and fermentation with Saccharomyces cerevisiae. Background Technology

[0002] Fructose oligosaccharides (FOS) are an important class of functional oligosaccharides widely used in food, feed, and functional products. Most existing FOS production processes use sucrose or molasses as raw materials, employing a transglycosylation reaction catalyzed by fructosyltransferases. Some processes also introduce yeast to consume residual glucose, fructose, and sucrose in the system, thereby increasing the proportion of the target component in the FOS product. However, existing technologies mainly focus on refined sucrose, molasses, or direct microbial fermentation, with public disclosures primarily emphasizing enzyme selection, substrate purification, and residual sugar removal.

[0003] Moringa is an abundant resource, containing rich sugar alcohols, with a high sucrose content. Since sucrose is a good substrate for fructosyltransferases, the aqueous extract of Moringa branches has the potential to be used as a raw material for the preparation of fructooligosaccharides (FOS). However, the aqueous extract of Moringa branches is not a single sucrose system; it also contains fermentable sugars such as glucose and fructose. These sugars remain in the system after enzymatic conversion, affecting the relative proportion of FOS and the purity of subsequent applications. Therefore, developing a coupled process that can both utilize the natural sugar components of Moringa branches to prepare FOS and further remove residual fermentable sugars while simultaneously obtaining high-value-added byproducts has significant application value. Summary of the Invention

[0004] The technical problem solved by this invention is to provide a coupled method for producing fructooligosaccharides and feed yeast using Moringa water extract. First, a high proportion of sucrose in the extract is used for fructosyltransferase catalysis, and then the subsequent residual sugar removal process is coupled with feed yeast production. This approach takes into account raw material substitution, process coupling, and by-product value enhancement, representing a route more suitable for utilizing agricultural and forestry by-products than existing conventional sucrose processes. This addresses the problems in existing technologies where fructooligosaccharide production typically relies on refined sucrose or molasses, has low utilization of agricultural and forestry by-products, is difficult to efficiently remove residual monosaccharides and disaccharides, and suffers from insufficient development of by-product value.

[0005] Technical Solution: To solve the above-mentioned technical problems, the present invention adopts the following technical solution:

[0006] A coupled method for producing fructooligosaccharides and feed yeast using Moringa water extract involves water extraction of Moringa branches to obtain a Moringa branch water extract containing fermentable sugars; then, an enzymatic reaction is carried out using fructosyltransferase to obtain an enzymatic digest containing fructooligosaccharides; the enzymatic digest is then inoculated with Saccharomyces cerevisiae for fermentation; after fermentation, the fermentation broth is separated into solid and liquid components, the supernatant is an fructooligosaccharide solution, and the bacterial cells are centrifuged, washed, and dried to obtain feed yeast.

[0007] The coupling method for producing fructooligosaccharides from Moringa water extract and feed yeast involves adding 10-100 U / mL of fructosyltransferase, reacting at 45-55℃, shaking at 300-900 rpm, and for 0.5-2 h.

[0008] The method for producing oligofructose using Moringa water extract and feed yeast involves a fermentation culture medium comprising the following components: yeast extract 0.5-2 g / L, ammonium sulfate 3-8 g / L, potassium dihydrogen phosphate 2-5 g / L, magnesium sulfate heptahydrate 0.2-1.0 g / L; brewer's yeast inoculation amount 0.5-2 g / L; and fermentation at 25-35 ℃ and 100-140 rpm in a shaker with ventilation for 12-36 h.

[0009] The method for producing oligofructose and feed yeast using Moringa water extract involves a solid-liquid ratio of 1:10-1:20, an extraction temperature of 40-90℃, and an extraction time of 10-60 min.

[0010] The method for producing fructooligosaccharides from Moringa water extract and feed yeast includes the following steps:

[0011] S1, after crushing the Moringa branch raw material, mix it with water, perform water extraction, and filter to obtain Moringa branch water extract;

[0012] S2, the water extract of Moringa branches is concentrated under reduced pressure to obtain a sugar concentrate containing sucrose, glucose and fructose.

[0013] S3, add fructosyltransferase to the sugar concentrate to carry out enzymatic transglycosylation reaction to obtain an enzymatic digest containing oligofructose;

[0014] S4, inoculate the enzyme digestion solution with Saccharomyces cerevisiae, and add nitrogen source and inorganic salt nutrients for fermentation, so that the remaining sucrose, glucose and fructose in the enzyme digestion solution are consumed, and yeast cells are generated at the same time.

[0015] S5, the fermentation broth is subjected to solid-liquid separation, the resulting supernatant is the oligofructose product liquid or its concentrate, and the obtained yeast cells are recovered to produce feed yeast.

[0016] The method for producing oligofructose and feed yeast using Moringa water extract involves a solid-liquid ratio of 1:15, a temperature of 80℃, and a time of 30 min.

[0017] The coupling method for producing fructooligosaccharides from Moringa water extract and feed yeast results in a sugar concentrate with a sucrose mass fraction of 8%-15%, a glucose mass fraction of 4%-10%, and a fructose mass fraction of 5%-10%.

[0018] The coupling method for producing fructooligosaccharides from Moringa water extract and feed yeast yields a sugar concentrate with a sucrose mass fraction of 10.5%, a glucose mass fraction of 6.5%, and a fructose mass fraction of 7.5%.

[0019] The method for producing oligofructose and feed yeast using Moringa water extract involves adding 50 U / mL of fructosyltransferase, reacting at 50 °C, shaking at 700 rpm, and for 1 h. Under these conditions, enzymatic digestion resulted in an 88% decrease in sucrose, a 33% increase in glucose, a 4% increase in fructose, a 19% increase in fructotriose, a 49% increase in fructotetraose, and a 27% increase in fructopentose.

[0020] The method for producing oligofructose using Moringa water extract and feed yeast involves a fermentation culture medium comprising the following components: yeast extract 1 g / L, ammonium sulfate 5 g / L, potassium dihydrogen phosphate 3 g / L, magnesium sulfate heptahydrate 0.5 g / L; brewer's yeast inoculation amount 1 g / L; fermentation is carried out in a shaker at 30 ℃ and 120 rpm for 24 h with ventilation; after fermentation, the remaining glucose content is 0, and the fructose and sucrose contents are both less than 1 g / L.

[0021] Beneficial effects: Compared with the prior art, the present invention has the following advantages:

[0022] (1) This invention converts the naturally occurring sucrose, glucose, and fructose in Moringa branches in a stepwise manner. The first stage involves hot water extraction and concentration to enrich the sugar components; the second stage involves the formation of fructooligosaccharides through fructosyltransferase catalysis; and the third stage involves fermentation with Saccharomyces cerevisiae to remove residual sucrose, glucose, and fructose, while simultaneously generating yeast cells. This process realizes the conversion of the sugar components of Moringa branches into two types of high-value-added products: sugar solution products rich in fructooligosaccharides and feed yeast products that can be further processed. It reduces sugar loss and waste emissions during the process and has good prospects for industrial application.

[0023] (2) Using low-value agricultural and forestry by-products such as Moringa branches as raw materials to replace refined sucrose as a substrate for fructooligosaccharide preparation improves resource utilization. Hot water extraction is used as a pretreatment method, which is simple, mild, and low-cost, and easy to scale up. Enzymatic conversion and yeast fermentation are linked together, which removes residual fermentable sugars in the system and obtains yeast cells simultaneously, realizing the coupled production of fructooligosaccharides and feed yeast.

[0024] (3) The remaining glucose in the fermentation system was 0, and the fructose and sucrose contents were both below 1 g / L. The feed yeast product obtained after fermentation had a crude protein content of 41.8%, a total amino acid content of 33.5%, a total cell count of 1.6 × 10^10 cells / g, a viable count of 8.7 × 10^8 CFU / g, and a survival rate of 68.4%. The above indicators indicate that the feed yeast product has a relatively balanced nutritional composition and moderate activity level, and has the potential to be used as a feed product. Attached Figure Description

[0025] Figure 1 Figure 1 shows the effect of different extraction temperatures on the yield of fermentable sugars in the aqueous extract of Moringa branches.

[0026] Figure 2 Figure showing the effect of different extraction times on the yield of fermentable sugars in the aqueous extract of Moringa branches;

[0027] Figure 3 Figure showing the changes in sugar composition before and after 5 hours of treatment with fructosyltransferase;

[0028] Figure 4 Figure showing the changes in sugar composition before and after 1 hour of treatment with fructosyltransferase.

[0029] Figure 5 This graph shows the changes in the residual sucrose, glucose, and fructose content in the fermentation system at different fermentation times. Detailed Implementation

[0030] The present invention will be further illustrated below with reference to specific embodiments. These embodiments are implemented based on the technical solutions of the present invention, and it should be understood that these embodiments are only used to illustrate the present invention and are not intended to limit the scope of the present invention.

[0031] Example 1

[0032] Extraction of fermentable sugars from Moringa branch aqueous extract

[0033] Extraction effect of fermentable sugars at different extraction temperatures: Moringa branches were pulverized to 60-80 mesh, and water was added at a solid-liquid ratio of 1:15. Hot water extraction was carried out at 40, 50, 60, 70, 80, and 90℃ for 40 min at each temperature. The content of fermentable sugars in the extract was measured, and the results are as follows: Figure 1(The letters in the figure indicate significant differences, p < 0.05).

[0034] The results showed that as the extraction temperature increased, the amount of fermentable sugars initially increased and then decreased, reaching a maximum of 8.18 g / L at 80℃. At this temperature, the concentrations of sucrose, glucose, and fructose were 5.39 g / L, 1.96 g / L, and 0.83 g / L, respectively. With further increases in temperature, the total amount of fermentable sugars did not increase significantly and even decreased slightly, suggesting that some glucose and fructose underwent degradation.

[0035] Extraction effect of fermentable sugars at different extraction times: After determining the extraction temperature to be 80℃, extraction times of 10, 20, 30, 40, 50, and 60 min were set respectively, and the extraction effect was tested. The results are as follows: Figure 2 As shown.

[0036] The results showed that the concentration of fermentable sugars continuously increased in the first 30 min and then stabilized after 30 min. At 30 min, the concentration of fermentable sugars was 8.12 g / L, of which the concentrations of sucrose, glucose, and fructose were 5.41 g / L, 1.86 g / L, and 0.85 g / L, respectively.

[0037] Therefore, the optimal extraction conditions for fermentable sugars from Moringa branches were determined to be a solid-liquid ratio of 1:15, an extraction temperature of 80℃, and an extraction time of 30 min. Under these conditions, the extraction rate of fermentable sugars was 121.8 mg / g DW, including 81.15 mg / g DW of sucrose, 27.9 mg / g DW of glucose, and 12.75 mg / g DW of fructose.

[0038] Example 2

[0039] A coupled method for producing fructooligosaccharides using Moringa water extract and feed yeast includes the following steps:

[0040] (1) The water extract of Moringa branches obtained in Example 1 (extraction conditions: solid-liquid ratio 1:15, extraction temperature 80℃, extraction time 30 min) was concentrated under reduced pressure using a rotary evaporator. The mass fractions of sucrose, glucose and fructose in the concentrated liquid were 10.5%, 6.5% and 7.5%, respectively.

[0041] (2) Fructosyltransferase was added to the concentrate to a concentration of 50 U / mL, and the mixture was incubated at 50℃ and 700 rpm for 5 h and 1 h, respectively. After the reaction, the changes in sugar composition were detected, and the results are as follows: Figure 3 and Figure 4 As shown.

[0042] The results showed that after 5 hours of incubation, compared with before the reaction, sucrose decreased by 90%, glucose increased by 36%, and fructose increased by 17%. However, when the incubation was shortened to 1 hour, compared with before the reaction, sucrose decreased by 88%, glucose increased by 33%, fructose increased by 4%, fructosaccharides increased by 19%, fructotetrasaccharides increased by 49%, and fructopentose increased by 27%, indicating that the sucrose in the concentrate could be effectively converted into fructooligosaccharide components by fructosyltransferase.

[0043] (3) Using the enzyme digestion solution in step (2) after 1 hour of incubation as the fermentation substrate, nutrients were added to make the yeast extract 1 g / L, ammonium sulfate 5 g / L, potassium dihydrogen phosphate 3 g / L, and magnesium sulfate heptahydrate 0.5 g / L in the fermentation medium. The inoculum of Saccharomyces cerevisiae was 1 g / L, and fermentation was carried out in a shaker at 30 ℃ and 120 rpm for 24 hours with ventilation. After fermentation, the sugar composition was tested, and the results are as follows. Figure 5 As shown.

[0044] The results showed that the residual glucose in the system was 0, and the contents of fructose and sucrose were both below 1 g / L. This indicates that yeast can effectively remove residual fermentable sugars after enzymatic digestion. The yeast cells obtained after fermentation can be recovered by centrifugation and further dried to prepare feed yeast products.

[0045] Quality analysis of the feed yeast product, on a dry basis, showed a crude protein content of 41.8%, a moisture content of 7.6%, and a crude ash content of 8.9%, with relatively stable quality after processing. Amino acid analysis revealed a total amino acid content of 33.5%, with lysine, methionine, and threonine content of 2.45%, 0.62%, and 1.78%, respectively, indicating a relatively complete protein composition. Regarding cellular indicators, the total cell count was 1.6 × 10^10 cells / g, the viable cell count was 8.7 × 10^8 CFU / g, and the survival rate was 68.4%, indicating that the product possesses certain biological activity and can meet general feed development needs. Based on the comprehensive analysis of conventional nutritional components, amino acid composition, and cell activity, this feed yeast product exhibits a relatively balanced nutritional composition and moderate activity level, demonstrating potential as a feed product.

[0046] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A coupling method for producing fructooligosaccharides from Moringa water extract and feed yeast, characterized in that, Moringa branch raw material was extracted with water to obtain Moringa branch water extract; then, fructosyltransferase was used to carry out an enzymatic reaction to obtain an enzyme digest; the enzyme digest was then inoculated with Saccharomyces cerevisiae for fermentation. After fermentation, the fermentation liquid was separated into solid and liquid, the supernatant was an oligofructose solution, and the bacterial cells were recovered to obtain feed yeast.

2. The method according to claim 1, characterized in that, The amount of fructosyltransferase added was 10-100 U / mL, the reaction temperature was 45-55 ℃, the shaking speed was 300-900 rpm, and the reaction time was 0.5-2 h.

3. The method according to claim 1, characterized in that, The fermentation medium comprises the following components: yeast extract 0.5-2 g / L, ammonium sulfate 3-8 g / L, potassium dihydrogen phosphate 2-5 g / L, magnesium sulfate heptahydrate 0.2-1.0 g / L; brewing yeast inoculation amount 0.5-2 g / L; fermentation is carried out in a shaker at 25-35 ℃ and 100-140 rpm with ventilation for 12-36 h.

4. The method according to claim 1, characterized in that, The solid-liquid ratio of the water extraction is 1:10-1:20, the extraction temperature is 40-90℃, and the extraction time is 10-60 min.

5. The method according to claim 1, characterized in that, Includes the following steps: S1, after crushing the Moringa branch raw material, mix it with water, perform water extraction, and filter to obtain Moringa branch water extract; S2, the water extract of Moringa branches is concentrated under reduced pressure to obtain a sugar concentrate; S3, add fructosyltransferase to the sugar concentrate to carry out an enzymatic reaction and obtain the enzyme digestion solution; S4, inoculate the enzyme digestion solution with Saccharomyces cerevisiae, and add nitrogen source and inorganic salt for fermentation, so that the remaining sucrose, glucose and fructose in the enzyme digestion solution are consumed, and yeast cells are generated at the same time. S5, the fermentation broth is subjected to solid-liquid separation, and the resulting supernatant is an oligofructose product solution or its concentrate. The yeast cells are recovered to obtain feed yeast.

6. The method according to claim 1 or 5, characterized in that, The solid-liquid ratio for water extraction was 1:15, the temperature was 80℃, and the extraction time was 30 min.

7. The method according to claim 5, characterized in that, The sugar concentrate contains 8%-15% sucrose, 4%-10% glucose, and 5%-10% fructose.

8. The method according to claim 5, characterized in that, The sugar concentrate contains 10.5% sucrose, 6.5% glucose, and 7.5% fructose by mass.

9. The method according to claim 1 or 5, characterized in that, The fructosyltransferase was added at a rate of 50 U / mL, the reaction temperature was 50 °C, the shaking speed was 700 rpm, and the reaction time was 1 h.

10. The method according to claim 1 or 5, characterized in that, The fermentation medium comprises the following components: yeast extract 1 g / L, ammonium sulfate 5 g / L, potassium dihydrogen phosphate 3 g / L, magnesium sulfate heptahydrate 0.5 g / L; brewer's yeast inoculation amount 1 g / L; fermentation time 24 h; fermentation is carried out in a shaker at 30 ℃ and 120 rpm with ventilation for 24 h.

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