Method for rapid degradation of monosaccharide and disaccharide in litchi pulp by lactic acid bacteria

Abstract

The application discloses a method for quickly degrading monosaccharide and disaccharide in litchi pulp by lactic acid bacteria and retaining active ingredients. The method comprises the following steps: (1) cleaning fresh litchi, removing shells and kernels, beating into pulp, pasteurizing and cooling to obtain litchi pulp; (2) under sterile conditions, adding activated lactic acid bacteria suspension into the litchi pulp, uniformly mixing, and then fermenting at 37±2 DEG C to obtain fermented litchi pulp; wherein the lactic acid bacteria are at least one of Lactobacillus fermentum, Lactobacillus paracasei, Lactobacillus rhamnosus and Lactobacillus plantarum. The method can quickly degrade monosaccharide (fructose, glucose) and disaccharide (sucrose) in litchi pulp, solve the problem of high sugar content in litchi pulp, and well retain the effective active ingredients (polyphenols) in litchi pulp from being degraded, and improve the antioxidant activity of litchi pulp.

Description

Technical Field

[0001] This invention belongs to the field of food technology, and specifically relates to a method for the rapid degradation of monosaccharides and disaccharides in lychee pulp by lactic acid bacteria while retaining active ingredients. Background Technology

[0002] Lychee is a typical tropical and subtropical fruit. The abundant phenolic compounds in lychee pulp have anti-inflammatory, antioxidant, lipid metabolism-regulating, and gut microbiota-regulating effects. However, the production and market availability of fresh lychees are concentrated from April to July. After harvesting, they are difficult to preserve and spoil easily, causing significant losses to agricultural production. Therefore, it is imperative to deepen the intensive processing and utilization of lychees and increase the economic added value of the lychee industry.

[0003] However, lychees are a prime example of high-sugar fruits, with their pulp containing abundant sugars. Every 100g of lychee pulp contains 16g of sugar, mostly fructose. Excessive sugar intake can easily lead to obesity, accelerated cell aging, diabetes, cardiovascular disease, and other health problems. The World Health Organization recommends that adults and children limit their sugar intake to less than 10% of their total energy intake (equivalent to no more than 50 grams of sugar per day). Further reducing this to less than 5% of total energy intake (no more than 25 grams) would bring even greater health benefits. With increasing consumer health awareness, controlling sugar intake has become a trend in the food industry and represents a market with enormous development potential.

[0004] Lactic acid bacteria fermentation is a simple, low-cost, and sustainable process that can preserve or improve the nutritional and sensory properties of raw materials and extend the shelf life of fruits and vegetables under hygienic and safe conditions. Fermented foods derived from lactic acid bacteria fermentation have been produced for thousands of years due to their health benefits and are gradually being accepted by more and more consumers.

[0005] Previous studies have shown that lactic acid bacteria can utilize the sugars in fruit juice as a carbon source during fermentation, thereby reducing the content of reducing sugars (sucrose, fructose, and glucose) in the pulp and further enhancing its health value. Zeinab E. Mousavi et al. found that lactic acid bacteria fermentation significantly consumed fructose and glucose in pomegranate juice and increased antioxidant activity. Therefore, lactic acid bacteria fermentation for degrading sugars in litchi pulp has great application potential. However, while lactic acid bacteria fermentation may degrade polyphenolic active substances with potential health-promoting effects, research on degrading sugars in litchi pulp while preserving polyphenolic active substances has not yet been reported. Summary of the Invention

[0006] The primary objective of this invention is to overcome the shortcomings and deficiencies of the prior art and to provide a method for lactic acid bacteria to rapidly degrade monosaccharides and disaccharides in lychee pulp while retaining active ingredients.

[0007] Another object of the present invention is to provide the application of the method for rapidly degrading monosaccharides and disaccharides of lychee pulp by lactic acid bacteria while retaining active ingredients.

[0008] The objective of this invention is achieved through the following technical solution:

[0009] A method for rapidly degrading monosaccharides and disaccharides in litchi pulp using lactic acid bacteria while retaining active ingredients includes the following steps:

[0010] (1) After washing the fresh lychees, removing the shells and pits, they are mashed into a pulp, pasteurized and cooled to obtain lychee pulp.

[0011] (2) Under aseptic conditions, the activated lactic acid bacteria suspension is added to the litchi pulp and pulp, mixed evenly, and then fermented at a constant temperature of 37±2℃ to obtain the fermented litchi pulp and pulp; wherein the lactic acid bacteria is at least one of Lactobacillus fermentum, Lactobacillus casei, Lactobacillus rhamnosus and Lactobacillus plantarum.

[0012] The pulping process described in step (1) is preferably carried out using a juicer; it is also preferred to use a conventional juicer to stir for 0.5 to 5 minutes.

[0013] The pasteurization conditions described in step (1) are: temperature 60-90℃, time 20-60min; preferably: temperature 80℃, time 40min.

[0014] The cooling described in step (1) is to cool to about 25-50°C using an ice water bath.

[0015] The lactic acid bacteria mentioned in step (2) are Lactobacillus fermentum, or Lactobacillus casei, or Lactobacillus rhamnosus, or Lactobacillus plantarum, or a mixture of Lactobacillus fermentum, Lactobacillus casei, Lactobacillus rhamnosus and Lactobacillus plantarum; more preferably, Lactobacillus fermentum, Lactobacillus casei, Lactobacillus rhamnosus and Lactobacillus plantarum; even more preferably, Lactobacillus fermentum, Lactobacillus casei, Lactobacillus rhamnosus and Lactobacillus plantarum in a volume ratio of 1:1:1:1.

[0016] The preferred fermenting lactobacillus is Limosilactobacillus fermentum GDMCC1.985.

[0017] The preferred Lactobacillus paracasei is Lactobacillus paracasei GDMCC 1.1799.

[0018] The preferred Lactobacillus rhamnosus is Lactobacillus rhamnosus GDMCC 1.1798.

[0019] The preferred Lactobacillus plantarum is Lactobacillus plantarum GDMCC1.140.

[0020] The concentration of the lactic acid bacteria suspension in step (2) is 6.0 to 9.0 Log (CFU / mL); preferably 9.0 Log (CFU / mL).

[0021] The amount of lactic acid bacteria suspension added in step (2) is calculated as 1-5% of its volume percentage in the fermentation system; preferably, it is calculated as 5% of its volume percentage in the system (i.e., the volume ratio of lactic acid bacteria suspension to lychee pulp is 5:95).

[0022] The activated lactic acid bacteria suspension in step (2) is preferably prepared by the following method: lactic acid bacteria are inoculated into a culture medium for activation. After activation, the bacteria are centrifuged, the supernatant is discarded, the precipitate is collected, and then washed by centrifugation with sterile PBS buffer. The collected microbial cell precipitate is then dissolved in sterile PBS buffer to obtain the lactic acid bacteria suspension.

[0023] The inoculation amount of the lactic acid bacteria is calculated based on its volume percentage of 1-5% in the system; preferably, it is calculated based on its volume percentage of 1% in the system.

[0024] The culture medium can be selected according to the strain to be activated; preferably, it is MRS broth medium.

[0025] The activation time is 20-24 hours; preferably 22 hours.

[0026] The centrifugation conditions are: 5000-9000 rpm / min, centrifugation for 5-10 min; preferably: 6000 rpm / min, centrifugation for 5 min.

[0027] The preferred number of centrifugal washing cycles is 2 to 3.

[0028] The fermentation time mentioned in step (2) is 1 to 4 days; preferably 4 days.

[0029] The method described above for rapidly degrading monosaccharides and disaccharides in litchi pulp while retaining active ingredients is applied to the fermentation of litchi pulp.

[0030] The present invention has the following advantages and effects compared with the prior art:

[0031] This invention utilizes a suspension of lactic acid bacteria, either single-strain or combined-strain (Lactobacillus fermentum, Lactobacillus casei, Lactobacillus rhamnosus, Lactobacillus plantarum), to ferment lychee pulp. This rapidly degrades monosaccharides (fructose, glucose) and disaccharides (sucrose) in lychee pulp, addressing the issue of high sugar content in lychee pulp and the risk of obesity and cardiovascular diseases from excessive consumption. Furthermore, the fermentation process using lactic acid bacteria or their combinations effectively preserves the effective active ingredient—polyphenols—in lychee pulp, preventing degradation and enhancing the antioxidant activity of the pulp. This has commercial value in the food fermentation industry and meets consumers' demands for high-quality consumption and a healthy lifestyle. In addition, the lactic acid bacteria used in this invention are common and safe in the market, and the cost of preparing fermented lychee pulp is low, making it highly valuable for the development and utilization of functional foods. Attached Figure Description

[0032] Figure 1 This is a graph showing the changes in polyphenol content in litchi pulp during lactic acid bacteria fermentation (in the graph, BLK is the control group without added lactic acid bacteria, 0d is fermentation day 0, 4d is fermentation day 4, Lb is Lactobacillus bulgaricus, Lf is Limosacchari fermentum, Lc is Lacticaseibacillus casei, L.pc is Lacticaseibacillus paracasei, Lr is Lacticaseibacillus rhamnosus, La is Lactobacillus acidophilus, and Lp is Lactobacillus plantarum)).

[0033] Figure 2 This is a diagram showing the degradation effect of lactic acid bacteria on monosaccharides (fructose and glucose) in lychee pulp; where A is the degradation effect of fructose; and B is the degradation effect of glucose and fructose.

[0034] Figure 3 This is a graph showing the changes in the iron reducing power (FRAP) of lychee pulp polyphenols during fermentation with different combinations of lactic acid bacteria (in the graph, combination 1 is a combination of Lactobacillus fermentum, Lactobacillus casei, Lactobacillus rhamnosus, and Lactobacillus plantarum; combination 2 is a combination of Lactobacillus fermentum and Lactobacillus casei; combination 3 is a combination of Lactobacillus rhamnosus and Lactobacillus plantarum; combination 4 is a combination of Lactobacillus fermentum, Lactobacillus casei, and Lactobacillus rhamnosus; combination 5 is a combination of Lactobacillus fermentum, Lactobacillus casei, and Lactobacillus plantarum). Detailed Implementation

[0035] The present invention will be further described in detail below with reference to embodiments, but the implementation of the present invention is not limited thereto. Unless otherwise specified, the reagents, methods, and equipment used in the present invention are conventional reagents, methods, and equipment in this technical field. Test methods in the following embodiments that do not specify specific experimental conditions are generally performed according to conventional experimental conditions or experimental conditions recommended by the manufacturer. Unless otherwise specified, the reagents and raw materials used in the present invention are commercially available.

[0036] 1. The strain information involved in the embodiments and comparative examples of this invention is as follows:

[0037] (1) The fermenting lactobacillus is Lactobacillus fermentum GDMCC 1.985 (according to the National Health Commission Announcement No. 4 of 2022 "List of microorganisms that can be used in food", the updated name of the microorganism is: Lactobacillus fermentum).

[0038] (2) Lactobacillus paracasei is Lactobacillus paracasei GDMCC1.1799 (According to the National Health Commission Announcement No. 4 of 2022, "List of Microbial Strains that Can Be Used in Food", the updated name of the microbial strain is: Lactobacillus paracasei).

[0039] (3) Lactobacillus rhamnosus is Lactobacillus rhamnosus GDMCC1.1798 (according to the National Health Commission Announcement No. 4 of 2022, "List of Strains that can be Used in Food", the updated name of the strain is: Lactobacillus rhamnosus).

[0040] (4) The Lactobacillus plantarum is Lactiplantibacillus plantarum GDMCC 1.140 (according to the National Health Commission Announcement No. 4 of 2022, "List of Microbial Strains that Can Be Used in Food", the updated name of the strain is: Lactiplantibacillus plantarum);

[0041] (5) Lactobacillus bulgaricus is Lactobacillus bulgaricus GDMCC1.189 (i.e. Lactobacillus delbrueckii subsp. bulgaricus GDMCC 1.189);

[0042] (6) Lactobacillus casei is Lactobacillus casei GDMCC 1.410;

[0043] (7) The Lactobacillus acidophilus is Lactobacillus acidophilus GDMCC 1.731;

[0044] All of the above strains were purchased from the Guangdong Provincial Microbial Culture Collection Center.

[0045] 2. The lactic acid bacteria suspension involved in the embodiments of the present invention is obtained by the following method:

[0046] (1) Lactic acid bacteria (Lactobacillus fermentum, Lactobacillus casei, Lactobacillus rhamnosus, Lactobacillus plantarum, Lactobacillus bulgaricus, Lactobacillus casei, Lactobacillus acidophilus) were inoculated into 40 mL of MRS broth medium at an inoculation rate of 1% (v / v) under aseptic conditions and activated at 37°C for 22 h.

[0047] (2) Centrifuge the activated lactic acid bacteria solution from step (1) at 6000 rpm / min for 5 min and remove the upper liquid culture medium, and collect the microbial cell precipitate;

[0048] (3) Wash the microbial cell precipitate collected in step (2) with sterile PBS buffer, centrifuge at 6000 rpm / min for 5 min and collect the microbial cell precipitate. Repeat the operation twice and dissolve the microbial cells in sterile PBS buffer to obtain a lactic acid bacteria suspension with a concentration of 9.0 Log (CFU / mL).

[0049] 3. The methods for measuring monosaccharides and disaccharides involved in the embodiments and comparative examples of this invention are as follows:

[0050] (1) Add 0.5 ml each of 0.5 mol / L 1-phenyl-3-methyl-5-pyrazolone (PMP) reagent dissolved in anhydrous methanol and 0.3 M NaOH solution to 100 μL of litchi pulp fermentation broth (fermentation supernatant). Mix thoroughly and react in a water bath at 70 °C for 30 min. Cool to room temperature, add 0.5 ml of 0.3 M HCl solution, mix thoroughly, add 1.6 ml of chloroform, shake thoroughly to extract, centrifuge (5000 rpm / min, 5 min) to remove the chloroform layer, and repeat the extraction three times. Filter the aqueous layer (not less than 0.4 ml) through a 0.22 μm filter membrane for HPLC analysis.

[0051] (2) Glucose, fructose, and sucrose in the samples were detected by high-performance liquid chromatography (HPLC, Shimadzu LC-20A) with a differential detector (DAD), and quantified using the external standard method. An amino column (250 mm × 4.6 mm, 5 μm) was used, with a mobile phase of acetonitrile (mobile phase A):water (mobile phase B) = 7:3 (V:V); a flow rate of 1.0 mL / min; a column oven and colorimetric detector temperature of 40 °C; an injection volume of 20 μL; and a detection wavelength of 245 nm. The substances corresponding to the chromatographic peaks were determined based on the retention times of the standards, and the content of each substance was expressed as mg / mL. The experiment was performed in triplicate.

[0052] 4. The methods for measuring the polyphenol content of litchi pulp involved in the embodiments and comparative examples of the present invention are as follows:

[0053] The monomeric phenols in the supernatant of lactic acid bacteria-fermented litchi pulp were quantified using a high-performance liquid chromatography (HPLC) system equipped with a diode array detector (DAD). A Zorbax SB-C18 column (250 mm × 4.6 mm, 5 μm) was used; the column temperature was 40 °C; the flow rate was set to 1.0 mL / min; and the injection volume was 20 μL. The mobile phase consisted of 0.4% glacial acetic acid (solvent A) and acetonitrile (solvent B) by volume percentage. The gradient elution program was as follows: 0–40 min, 5%–25% solvent B; 40–45 min, 25%–35% solvent B; 45–50 min, 35%–50% solvent B. The substances corresponding to the collected chromatographic peaks at a wavelength of 280 nm were determined based on the retention times of the standards. The experiment was conducted in triplicate.

[0054] 5. The method for measuring the antioxidant activity of litchi pulp polyphenols involved in the embodiments and comparative examples of the present invention is as follows:

[0055] (1) Preparation of FRAP stock solution:

[0056] Solution A, i.e., 40 mmol / L hydrochloric acid solution: Take 0.834 mL of concentrated hydrochloric acid (12 mol / L) and add water to 250 mL. Store at 4℃ for later use.

[0057] Solution B, i.e., 300 mmol / L sodium acetate buffer solution (pH = 3.6): Weigh 0.7750 g of sodium acetate trihydrate (C2H3NaO2·3H2O), add 4 mL of glacial acetic acid, dilute with water to a final volume of 250 mL, and store at 4°C for later use.

[0058] Solution C, i.e. 10 mmol / L TPTZ solution: Weigh 0.3122 g of 2,4,6-tripyridyltriazine (TPTZ) sample, dilute to 100 mL with 40 mmol / L hydrochloric acid (solution A), and store in a 4°C refrigerator protected from light using aluminum foil for later use.

[0059] Solution D, i.e. 20 mmol / L FeCl3·6H2O: Weigh 0.5406 g of ferric chloride hexahydrate, and dilute to 100 mL with 40 mmol / L hydrochloric acid (solution A). Store in a tin foil container at 4°C for later use.

[0060] (2) Preparation of FRAP working solution: Mix 25 mL solution B + 2.5 mL solution C + 2.5 mL solution D thoroughly and incubate in a water bath at 37°C until ready for use.

[0061] (3) The iron reducing power (FRAP) of polyphenols during the 0-4 day fermentation of litchi pulp by lactic acid bacteria, i.e., the antioxidant activity, was measured using a UV spectrophotometer. 300 μL of a polyphenol extract sample diluted to a certain factor was taken, and then FRAP working solution preheated to 37℃ was added. After mixing with a vortex mixer, the mixture was allowed to stand at room temperature in the dark for 5 min. The absorbance at 593 nm was measured using a UV spectrophotometer, with the unit being mmol / L FeSO4. The experiment was repeated three times.

[0062] Example 1

[0063] The lychee pulp and pulp were fermented using lactic acid bacteria. By volume percentage, the raw materials included: 5% of a 9.0 Log (CFU / mL) lactobacillus suspension and 95% lychee pulp and pulp.

[0064] The specific steps for making it are as follows:

[0065] S1: Wash fresh lychees (purchased from Guangzhou Fruit Wholesale Market, variety: Huaizhi) with distilled water, remove the shells and pits, weigh 120g of lychee pulp and blend it in a juicer (brand: Philips, model: HR2095) for 30s, dispense it into 3 tubes of 38mL lychee pulp under aseptic conditions, pasteurize at 80℃ for 40min, and immediately cool the sterilized pulp in an ice water bath to about 25-50℃ for inoculation.

[0066] S2: Under aseptic conditions, 2 mL of 9.0 Log (CFU / mL) Lactobacillus fermentation suspension was added to 38 mL of pasteurized lychee pulp and mixed evenly to start fermentation. The fermentation temperature was 37℃ and the fermentation time was 4 days to obtain lychee pulp and pulp fermented with Lactobacillus fermentation.

[0067] S3: Centrifuge the litchi pulp and pulp fermented with Lactobacillus fermentation for 0, 1, 2, 3 and 4 days respectively (centrifugation conditions: 9000 rpm / min, 10 min) to obtain the supernatant of the lactic acid bacteria fermented litchi pulp and pulp.

[0068] Example 2

[0069] The litchi pulp and juice were fermented using lactic acid bacteria, expressed as a volume percentage, and consisted of the following ingredients: 5% of a 9.0 Log (CFU / mL) suspension of Lactobacillus casei and 95% litchi pulp and juice.

[0070] The remaining production steps are the same as in Example 1.

[0071] Example 3

[0072] The lychee pulp and pulp were fermented using lactic acid bacteria, and the composition, expressed as a volume percentage, included the following ingredients: 5% Lactobacillus rhamnosus suspension at 9.0 Log (CFU / mL) and 95% lychee pulp and pulp.

[0073] The remaining production steps are the same as in Example 1.

[0074] Example 4

[0075] The litchi pulp and pulp were fermented using lactic acid bacteria, and the composition, expressed as a volume percentage, included the following ingredients: 5% of a 9.0 Log (CFU / mL) Lactobacillus plantarum suspension and 95% litchi pulp and pulp.

[0076] The remaining production steps are the same as in Example 1.

[0077] Example 5

[0078] The litchi pulp and pulp were fermented using a variety of lactic acid bacteria, expressed as a volume percentage, and included the following raw materials: 1.25% Lactobacillus fermentum (9.0 Log (CFU / mL), 1.25% Lactobacillus casei, 1.25% Lactobacillus rhamnosus, 1.25% Lactobacillus plantarum, and 95% litchi pulp and pulp (the various bacteria were added to the litchi pulp and pulp in proportion, the same below).

[0079] The remaining production steps are the same as in Example 1.

[0080] Comparative Example 1

[0081] The litchi pulp and pulp were fermented using lactic acid bacteria, and the composition, expressed as a percentage by volume, consisted of the following ingredients: 5% of a 9.0 Log (CFU / mL) Lactobacillus bulgaricus suspension and 95% litchi pulp and pulp.

[0082] The remaining production steps are the same as in Example 1.

[0083] Comparative Example 2

[0084] The lychee pulp and pulp were fermented using lactic acid bacteria, and the composition, expressed as a volume percentage, included the following ingredients: 5% Lactobacillus casei suspension at 9.0 Log (CFU / mL) and 95% lychee pulp and pulp.

[0085] The remaining production steps are the same as in Example 1.

[0086] Comparative Example 3

[0087] The lychee pulp and pulp were fermented using lactic acid bacteria, and the composition, expressed as a volume percentage, included the following ingredients: 5% Lactobacillus acidophilus suspension at 9.0 Log (CFU / mL) and 95% lychee pulp and pulp.

[0088] The remaining production steps are the same as in Example 1.

[0089] Comparative Example 4

[0090] The litchi pulp and pulp were fermented using a variety of lactic acid bacteria. The fermentation process, expressed as a volume percentage, included the following ingredients: 2.5% Lactobacillus fermentum (9.0 Log (CFU / mL), 2.5% Lactobacillus casei, and 95% litchi pulp and pulp.

[0091] The remaining production steps are the same as in Example 1.

[0092] Comparative Example 5

[0093] The litchi pulp and pulp were fermented using a variety of lactic acid bacteria, expressed as a volume percentage, and included the following raw materials: 2.5% Lactobacillus rhamnosus (9.0 Log (CFU / mL), 2.5% Lactobacillus plantarum, and 95% litchi pulp and pulp.

[0094] The remaining production steps are the same as in Example 1.

[0095] Comparative Example 6

[0096] The litchi pulp and pulp were fermented using a variety of lactic acid bacteria. The fermentation process, expressed as a percentage by volume, included the following ingredients: 1.67% Lactobacillus fermentum (9.0 Log (CFU / mL), 1.67% Lactobacillus casei, 1.66% Lactobacillus rhamnosus, and 95% litchi pulp and pulp.

[0097] The remaining production steps are the same as in Example 1.

[0098] Comparative Example 7

[0099] The litchi pulp and pulp were fermented using a variety of lactic acid bacteria. The fermentation process, expressed as a percentage by volume, included the following ingredients: 1.67% Lactobacillus fermentum (9.0 Log (CFU / mL), 1.67% Lactobacillus casei, 1.66% Lactobacillus plantarum, and 95% litchi pulp and pulp.

[0100] The remaining production steps are the same as in Example 1.

[0101] Example 1: Effect of Lactic Acid Bacteria Fermentation on Polyphenol Content in Lychee Pulp

[0102] The changes in polyphenol content in the supernatant of litchi pulp after lactic acid bacteria fermentation (4 days) in Examples 1-4 and Comparative Examples 1-3 were determined, with litchi pulp without lactic acid bacteria fermentation (4 days) serving as a control (BLK-4d). The changes in polyphenol content in litchi pulp are shown below. Figure 1 The results showed that the polyphenol content in litchi pulp decreased after fermentation with Lactobacillus bulgaricus (Comparative Example 1), Lactobacillus casei (Comparative Example 2), and Lactobacillus acidophilus (Comparative Example 3). However, the polyphenols in litchi pulp were not significantly degraded after fermentation with Lactobacillus fermentum (Example 1), Lactobacillus casei-like bacteria (Example 2), Lactobacillus rhamnosus (Example 3), and Lactobacillus plantarum (Example 4), and some new phenolic substances were generated. This indicates that fermentation with Lactobacillus fermentum, Lactobacillus casei-like bacteria, Lactobacillus rhamnosus, and Lactobacillus plantarum can preserve polyphenols, an active ingredient in litchi pulp, which is beneficial to human health.

[0103] Example 2: Effect of Lactic Acid Bacteria Fermentation on the Content of Monosaccharides (Fructose, Glucose) and Disaccharides (Sucrose) in Lychee Pulp

[0104] The content of monosaccharides (fructose, glucose) and disaccharides (sucrose) in the supernatant of litchi pulp and pulp fermented (0-4 days) by Example 5 (Lactobacillus fermentum, Lactobacillus casei, Lactobacillus rhamnosus and Lactobacillus plantarum; abbreviated as: combination 1) and Comparative Example 1 (Lactobacillus bulgaricus), Comparative Example 2 (Lactobacillus casei) and Comparative Example 3 (Lactobacillus acidophilus) was determined.

[0105] The changes in monosaccharide (fructose, glucose) content in lychee pulp are as follows: Figure 2 As shown in Table 1, the changes in disaccharide (sucrose) content are as follows.

[0106] Table 1. Effects of different lactic acid bacteria on the sucrose content in litchi pulp.

[0107] Fermentation time / day 0 1 2 3 4 Lactobacillus bulgaricus (Lb) 28.31 0.97 0.97 0.26 - Lactobacillus casei (Lc) 28.31 0.33 0.03 - - Lactobacillus acidophilus (La) 28.31 0.06 - - - Combination 1 28.31 - - - -

[0108] Note: "-" indicates that the sucrose content in the lychee pulp was not detected.

[0109] The results showed that different lactic acid bacteria fermented with these bacteria had varying effects on degrading monosaccharides and disaccharides in litchi pulp. After one day of fermentation, the combined fermentation of four lactic acid bacteria—*Lactobacillus fermentum*, *Lactobacillus casei*, *Lactobacillus rhamnosus*, and *Lactobacillus plantarum* (combination 1)—involved the rapid degradation of fructose and glucose in litchi pulp. Combination 1 showed the most significant effect on sucrose degradation, with complete degradation occurring after one day. This indicates that the combined fermentation of *Lactobacillus fermentum*, *Lactobacillus casei*, *Lactobacillus rhamnosus*, and *Lactobacillus plantarum* can rapidly degrade monosaccharides and disaccharides in litchi pulp without affecting polyphenol content, effectively preserving the active polyphenolic components.

[0110] Effect Example 3: Effect of Lactic Acid Bacteria Fermentation on the Antioxidant Activity of Polyphenols in Litchi Pulp

[0111] The antioxidant activity of polyphenols in the supernatant of fermented litchi pulp was determined in Example 5 (Lactobacillus fermentum, Lactobacillus casei, Lactobacillus rhamnosus, and Lactobacillus plantarum; combination 1) and Comparative Examples 4 (Lactobacillus fermentum, Lactobacillus casei; combination 2), 5 (Lactobacillus rhamnosus, Lactobacillus plantarum; combination 3), 6 (Lactobacillus fermentum, Lactobacillus casei, Lactobacillus rhamnosus; combination 4), and 7 (Lactobacillus fermentum, Lactobacillus casei, and Lactobacillus plantarum; combination 5).

[0112] The results are as follows Figure 3 As shown, the results indicate that different combinations of lactic acid bacteria fermentation have varying effects on the antioxidant activity of polyphenols in litchi pulp. The litchi pulp fermented with the combination of four lactic acid bacteria—*Lactobacillus fermentatus*, *Lactobacillus casei*, *Lactobacillus rhamnosus*, and *Lactobacillus plantarum* (combination 1)—showed the highest antioxidant activity of polyphenols, significantly higher than other combinations. This suggests that combined fermentation with *Lactobacillus fermentatus*, *Lactobacillus casei*, *Lactobacillus rhamnosus*, and *Lactobacillus plantarum* is beneficial for enhancing the antioxidant capacity of polyphenolic active components in litchi pulp.

[0113] In summary, the combined fermentation of *Lactobacillus fermentum*, *Lactobacillus casei*, *Lactobacillus rhamnosus*, and *Lactobacillus plantarum* in this invention not only rapidly degrades monosaccharides (fructose, glucose) and disaccharides (sucrose) in lychee pulp but also effectively preserves polyphenols. Furthermore, it may generate other antioxidant substances, thereby enhancing antioxidant activity. Therefore, functional products made from lychee pulp fermented with this combined lactic acid bacteria possess significant commercial value.

[0114] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Any changes, modifications, substitutions, combinations, or simplifications made without departing from the spirit and principle of the present invention shall be considered equivalent substitutions and shall be included within the protection scope of the present invention.

Claims

1. A method for rapidly degrading monosaccharides and disaccharides in litchi pulp using lactic acid bacteria while retaining active ingredients, characterized in that... Includes the following steps: (1) After washing the fresh lychees, removing the shells and pits, they are mashed into a pulp, pasteurized and cooled to obtain lychee pulp. (2) Under aseptic conditions, the activated lactic acid bacteria suspension is added to the litchi pulp and pulp, mixed evenly, and then fermented at a constant temperature of 37±2℃ to obtain the fermented litchi pulp and pulp; wherein, the lactic acid bacteria are a mixture of Lactobacillus fermentum, Lactobacillus casei, Lactobacillus rhamnosus and Lactobacillus plantarum, with a volume ratio of 1:1:1:1; The pasteurization conditions described in step (1) are: temperature 80℃, time 40 min; The concentration of the lactic acid bacteria suspension mentioned in step (2) is 9.0 Log CFU / mL; The inoculation amount of the lactic acid bacteria suspension mentioned in step (2) is calculated based on its volume percentage of 5% in the fermentation system; The fermenting lactobacillus mentioned is Lactobacillus fermentum ( Limosilactobacillus fermentum GDMCC 1.985; The aforementioned *Lactobacillus casei* is *Lactobacillus casei* ( Lacticaseibacillus paracasei GDMCC1.1799; The Lactobacillus rhamnosus mentioned is Lactobacillus rhamnosus ( Lacticaseibacillus rhamnosus GDMCC1.1798; The aforementioned *Lactobacillus plantarum* is *Lactobacillus plantarum* (… Lactiplantibacillus plantarum GDMCC 1.

140.

2. The method according to claim 1, characterized in that: The fermentation time mentioned in step (2) is 1 to 4 days.

3. The method according to claim 1, characterized in that: The cooling described in step (1) is to cool to 25-50 °C using an ice water bath.

4. The method for rapid degradation of monosaccharides and disaccharides in litchi pulp by lactic acid bacteria and preservation of active ingredients as described in any one of claims 1 to 3, applied in the fermentation of litchi pulp.

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