Method for producing prebiotics material using enzymatic hydrolysis of apple juice residue

Abstract

The present invention relates to a method for manufacturing a prebiotic material, characterized by comprising the steps of: crushing pretreated apples and extracting juice to separate the pulp and the juice; and adding an enzyme to the separated pulp to induce an enzymatic reaction and then filtering; and to a prebiotic material manufactured by the said method and a processed food using said prebiotic material.

Description

Technology Field

[0001] The present invention relates to a method for manufacturing a prebiotic material characterized by including the step of adding an enzyme to apple juice residue and causing an enzymatic reaction, and to a prebiotic material manufactured by said method. The prebiotic material of the present invention has the advantage of excellent effects in improving intestinal function and promoting the growth of beneficial bacteria in the intestines. Background Technology

[0002] Numerous microorganisms inhabit the human gastrointestinal tract (GI) in colonies, collectively known as the enteric microbiota. The enteric microbiota can be divided into beneficial and harmful bacteria, and health status is regulated by their balance. To maintain health, the enteric microbiota must be maintained in a state where beneficial bacteria are abundant and harmful bacteria are scarce. Among the efforts to correct imbalances in the enteric microbiota and enhance the activity of beneficial microorganisms, the most popular and widespread method is the consumption of probiotic products.

[0003] Probiotics refer to microorganisms beneficial to the human body; they are live bacteria that reach the intestines, produce lactic acid, and change the environment to acidic conditions that prevent harmful bacteria from growing and allow beneficial bacteria to increase. However, because the probability of ingested probiotics surviving to the desired body parts—such as the small and large intestines—is low, prebiotics—organic substances that serve as food for probiotics—are usually consumed together with probiotics to enhance their activity and lifespan within the body.

[0004] Prebiotics are indigestible components that promote the growth of beneficial bacteria in the gut. They serve as nutrient sources for probiotics, thereby helping to improve the intestinal environment. Certain food components are utilized by beneficial microorganisms in the large intestine to promote their growth or activity, which in turn exerts beneficial effects on host health. Consequently, there is a growing need to explore and develop prebiotic materials derived from food components.

[0005] apologize( Malus pumila var. dulcissima Apples are perennial woody plants classified under the Rosaceae family, and their fruits are known for their excellent sensory characteristics. The first apple variety introduced to Korea was Neunggeum, but currently, varieties such as Hongok, Fuji, and Aori are mainly cultivated. Apples are not only rich in water-soluble solids, organic acids, and vitamins, but also contain organic substances such as carbohydrates and physiologically active polyphenol compounds such as caffeic acid, quercetin, chlorogenic acid, phlorizin, and epicatechin. As such, they are known to be effective in preventing degenerative diseases caused by oxidation, including arteriosclerosis, diabetes, and hypertension.

[0006] In Korea, most of the apple production is consumed as fresh fruit, but there are significant difficulties in maintaining marketability and extending shelf life due to physiological factors during the distribution process. Meanwhile, apple juice currently accounts for more than 90% of processed apple products other than fresh fruit, and the pulp remaining after juice production is mainly discarded.

[0007] Korean Patent Publication No. 2023-0051593 discloses a prebiotic food, and Korean Patent Publication No. 2021-0052051 discloses a complex biotic lactic acid bacteria fermented beverage containing prebiotics, probiotics, and postbiotics, but these are different from the method of manufacturing a prebiotic material by enzymatically hydrolyzing apple juice pomace according to the present invention. The problem to be solved

[0008] The present invention was devised in response to the above-mentioned needs, and the objective of the present invention is to provide a method for manufacturing a prebiotic material that is easy to consume with improved taste and flavor while exhibiting prebiotic functionality, by utilizing apple juice residue, which is mainly discarded during the process of processing and juicing apples, and optimizing manufacturing conditions such as material pretreatment and enzyme treatment. means of solving the problem

[0009] To solve the above problem, the present invention provides a method for manufacturing a prebiotic material, characterized by comprising the steps of: (1) crushing and extracting juice from an apple to separate it into pulp and juice; (2) mixing water with the separated pulp from step (1) and then crushing it to produce an apple pulp mixture; (3) sterilizing and cooling the apple pulp mixture produced in step (2); and (4) adding an enzyme to the cooled apple pulp mixture from step (3) to perform an enzymatic reaction and then filtering.

[0010] In addition, the present invention provides a prebiotic material prepared by the above method.

[0011] In addition, the present invention provides a method for producing ABC juice characterized by mixing the above-mentioned prebiotic material with apple juice, carrot puree, carrot juice, beet juice, water, and fructooligosaccharide. Effects of the invention

[0012] The prebiotic material produced according to the manufacturing method of one aspect of the present invention is commercially competitive because it uses the juice residue remaining after processing and juicing apples as the main ingredient, thereby increasing the utilization of the juice residue, which is usually discarded.

[0013] In addition, the prebiotic material produced according to the manufacturing method according to one aspect of the present invention has the advantage of being easy to consume with a soft flavor, as the characteristic off-flavor or grassy smell of the material is removed by pre-treating the apple juice residue under optimal conditions.

[0014] In addition, it is believed that the prebiotic material prepared according to the method of one aspect of the present invention can help improve intestinal health by increasing the production of short-chain fatty acids and promoting the growth of beneficial bacteria in the intestines.

[0015] In addition, the prebiotic material prepared according to the method according to one aspect of the present invention possesses prebiotic functionality, has a small particle size, high solubility, and excellent shelf life, so it can be easily utilized in various materials such as food additives, processed foods, health functional foods, or pharmaceutical compositions. Brief explanation of the drawing

[0016] Figure 1 is a graph comparing the particle sizes of enzyme-treated apple pomace filtrate according to pH conditions. Figure 2 is a graph comparing the water solubility of apple juice residue filtrate according to the enzyme reaction time. Figure 3 shows the reaction time according to the enzyme reaction time. This is a graph comparing the relative molecular weights of apple pomace filtrate. Figure 4 is a graph comparing the effect of probiotic viable cell proliferation of enzyme-treated apple pomace filtrate (APH) and ABC juice (APHJ). Figure 5 is a graph comparing the short-chain fatty acid content of mouse feces treated with a normal diet, enzyme-free apple pomace filtrate (0 h sample), enzyme-treated apple pomace filtrate (1 h sample), and 1 h sample + fructooligosaccharide (1 h sample + FOS), respectively. Specific details for implementing the invention

[0017] To achieve the objective of the present invention, the present invention

[0018] (1) A step of crushing and juicing apples to separate them into pulp and juice;

[0019] (2) A step of preparing an apple juice pulp mixture by mixing water with the separated juice pulp from step (1) above and then grinding it;

[0020] (3) A step of sterilizing and cooling the apple juice mixture prepared in step (2) above; and

[0021] (4) A method for manufacturing a prebiotic material is provided, characterized by including the step of adding an enzyme to the cooled apple juice mixture of step (3) above, causing an enzymatic reaction, and then filtering.

[0022] In the method for manufacturing the prebiotic material of the present invention, the apple juice pulp mixture of step (2) can preferably be prepared by mixing water with the separated juice pulp in a ratio of 0.8 to 1.2:0.8 to 1.2 (w:v) and then grinding it, and more preferably by mixing water with the separated juice pulp in a ratio of 1:1 (w:v) and then grinding it to prepare the juice pulp having a concentration suitable for subsequent enzyme treatment.

[0023] In addition, in the method for manufacturing the prebiotic material of the present invention, step (3) can preferably be performed by sterilizing the apple juice pomace mixture at 70 to 90°C for 50 to 70 minutes and then cooling it to 35 to 45°C, and more preferably by sterilizing the apple juice pomace mixture at 80°C for 60 minutes and then cooling it to 40°C. By sterilizing and cooling as described above, the apple juice pomace mixture can be pretreated to improve the treatment efficiency during enzyme treatment.

[0024] In addition, in the method for manufacturing the prebiotic material of the present invention, the enzyme reaction of step (4) can preferably be carried out at 35 to 45°C for 30 to 90 minutes, and more preferably at 40°C for 60 minutes. By carrying out the enzyme reaction under such conditions, the discarded juice residue can be utilized, and the functionality of the prebiotic can be further enhanced.

[0025] The enzyme used in the above enzyme reaction may be a mixed enzyme consisting of xylanase, cellulase, pectinase, hemicellulase, and beta-glucanase.

[0026] The method for manufacturing a prebiotic material of the present invention, more specifically, is

[0027] (1) A step of crushing and juicing apples to separate them into pulp and juice;

[0028] (2) A step of preparing an apple juice pulp mixture by mixing water with the separated juice pulp from step (1) in a ratio of 0.8~1.2:0.8~1.2 (w:v) and then grinding it;

[0029] (3) A step of sterilizing the apple juice residue mixture prepared in step (2) at 70~90℃ for 50~70 minutes and then cooling it to 35~45℃; and

[0030] (4) The method may include a step of adding enzymes to the cooled apple juice mixture from step (3) above, performing an enzymatic reaction at 35 to 45°C for 30 to 90 minutes, and then filtering.

[0031] More specifically,

[0032] (1) A step of crushing and juicing apples to separate them into pulp and juice;

[0033] (2) A step of preparing an apple juice pulp mixture by mixing water in a 1:1 (w:v) ratio with the separated juice pulp from step (1) above, and then grinding it;

[0034] (3) A step of sterilizing the apple juice pulp mixture prepared in step (2) at 80°C for 60 minutes and then cooling it to 40°C; and

[0035] (4) The step of adding enzymes to the cooled apple juice mixture of step (3) above, reacting the enzymes at 40°C for 60 minutes, and then filtering may be included.

[0036] In addition, when manufacturing the prebiotic material of the present invention, the prebiotic material can be manufactured by additionally including a composition for pre-treating apples to impart a taste-enhancing effect in addition to the sterilization and browning inhibition effects of apples, specifically,

[0037] (1) Step of immersing the apple in apple mint extract, then taking it out and washing it;

[0038] (2) A step of preparing an immersion solution by mixing water, hibiscus extract, lemongrass extract, Brussels sprouts juice, citric acid and cyclodextrin;

[0039] (3) A step of immersing the washed apple from step (1) in the soaking solution prepared in step (2) and then taking it out to ripen;

[0040] (4) A step of crushing and extracting juice from the ripened apples of step (3) above to separate them into pulp and juice;

[0041] (5) A step of preparing an apple juice pulp mixture by mixing water with the separated juice pulp from step (4) above and then grinding it;

[0042] (6) A step of sterilizing and cooling the apple juice mixture prepared in step (5) above; and

[0043] (7) The method may include a step of adding an enzyme to the cooled apple juice mixture of step (6) above, performing an enzymatic reaction, and then filtering.

[0044] More specifically,

[0045] (1) A step of immersing an apple in apple mint extract at 18~20℃ for 30~90 seconds, then taking it out and washing it;

[0046] (2) A step of preparing an immersion solution by mixing 91-93% by weight of water, 2-4% by weight of hibiscus extract, 1-3% by weight of lemongrass extract, 1-3% by weight of Brussels sprouts juice, 0.2-0.8% by weight of citric acid and 0.2-0.8% by weight of cyclodextrin, based on the total weight of the immersion solution;

[0047] (3) A step of immersing the washed apples from step (1) in the soaking solution prepared in step (2) at a ratio of 0.8~1.2:1.3~1.7 (w / v) at 18~20℃ for 4~6 minutes, then removing them and aging them at 8~10℃ for 50~70 minutes;

[0048] (4) A step of crushing and extracting juice from the ripened apples of step (3) above to separate them into pulp and juice;

[0049] (5) A step of preparing an apple juice pulp mixture by mixing water with the separated juice pulp from step (4) in a ratio of 0.8~1.2:0.8~1.2 (w:v) and then grinding it;

[0050] (6) A step of sterilizing the apple juice residue mixture prepared in step (5) at 70~90℃ for 50~70 minutes and then cooling it to 35~45℃; and

[0051] (7) The method may include a step of adding enzymes to the cooled apple juice mixture of step (6) above, performing an enzymatic reaction at 35-45°C for 30-90 minutes, and then filtering.

[0052] Most specifically,

[0053] (1) A step of immersing an apple in apple mint extract at 18~20℃ for 60 seconds, then taking it out and washing it;

[0054] (2) A step of preparing an immersion solution by mixing 92% by weight of water, 3% by weight of hibiscus extract, 2% by weight of lemongrass extract, 2% by weight of Brussels sprouts juice, 0.5% by weight of citric acid, and 0.5% by weight of cyclodextrin, based on the total weight of the immersion solution;

[0055] (3) A step of immersing the washed apples from step (1) in the soaking solution prepared in step (2) at a ratio of 1:1.5 (w / v) at 18~20℃ for 5 minutes, then removing them and aging them at 8~10℃ for 60 minutes;

[0056] (4) A step of crushing and extracting juice from the ripened apples of step (3) above to separate them into pulp and juice;

[0057] (5) A step of preparing an apple juice pulp mixture by mixing water in a 1:1 (w:v) ratio with the separated juice pulp from step (4) above, and then grinding it;

[0058] (6) A step of sterilizing the apple juice pulp mixture prepared in step (5) at 80°C for 60 minutes and then cooling it to 40°C; and

[0059] (7) The step of adding enzymes to the cooled apple juice mixture from step (6) above, reacting the enzymes at 40°C for 60 minutes, and then filtering may be included.

[0060] In the method for preparing the prebiotic material of the present invention, the apple mint extract of step (1) can preferably be prepared by adding 8 to 12 times (v / w) of water to apple mint and extracting at 50 to 60°C for 50 to 70 minutes, and more preferably, the apple mint extract can be prepared by adding 10 times (v / w) of water to apple mint and extracting at 50 to 60°C for 1 hour. Pre-treating the apple under the same conditions as in step (1) allows the taste and aroma of apple mint to be imbued, thereby removing the characteristic grassy smell of the material and further enhancing the flavor.

[0061] In addition, in the method for manufacturing the prebiotic material of the present invention, preparing the soaking solution with the material type and mixing ratio of step (2) above could further enhance the taste and flavor while blending well with the apple.

[0062] The hibiscus extract of step (2) above may preferably be prepared by adding 12 to 18 times (v / w) of water to hibiscus and extracting at 85 to 95°C for 5 to 15 minutes, and more preferably by adding 15 times (v / w) of water to hibiscus and extracting at 90°C for 10 minutes.

[0063] In addition, the lemongrass extract of step (2) above may preferably be prepared by adding 12 to 18 times (v / w) of water to lemongrass and extracting at 85 to 95°C for 5 to 15 minutes, and more preferably by adding 15 times (v / w) of water to lemongrass and extracting at 90°C for 10 minutes.

[0064] In addition, the Brussels sprout juice of step (2) above may preferably be prepared by extracting juice from Brussels sprouts blanched at 90 to 110°C for 30 to 90 seconds and then filtering, and more preferably by extracting juice from Brussels sprouts blanched at 100°C for 60 seconds and then filtering.

[0065] In addition, in the method for manufacturing the prebiotic material of the present invention, step (3) can preferably be performed by immersing an apple in an immersion solution at a ratio of 0.8~1.2:1.3~1.7 (w / v) at 18~20°C for 4~6 minutes, then removing it and aging it at 8~10°C for 50~70 minutes, and more preferably by immersing an apple in an immersion solution at a ratio of 1:1.5 (w / v) at 18~20°C for 5 minutes, then removing it and aging it at 8~10°C for 60 minutes. Immersion and aging under the above conditions could provide a taste enhancement effect in addition to the sterilization and browning inhibition effects of the apple.

[0066] The present invention also provides a prebiotic material prepared by the above method.

[0067] The present invention also provides a method for producing ABC juice characterized by mixing the above-mentioned prebiotic material with apple juice, carrot puree, carrot juice, beet juice, water, and fructooligosaccharide.

[0068] In the above ABC juice, ABC stands for the first letters of the main ingredients: Apple, Beet, and Carrot.

[0069] In the method for preparing ABC juice according to the present invention, the mixture may preferably be a mixture of 20 to 30 parts by weight of prebiotic material, 180 to 220 parts by weight of apple juice, 80 to 120 parts by weight of carrot puree, 80 to 120 parts by weight of carrot juice, 80 to 120 parts by weight of beet juice, 80 to 120 parts by weight of water, and 40 to 70 parts by weight of fructooligosaccharide, and more preferably a mixture of 25 parts by weight of prebiotic material, 200 parts by weight of apple juice, 100 parts by weight of carrot puree, 100 parts by weight of carrot juice, 100 parts by weight of beet juice, 100 parts by weight of water, and 53.95 parts by weight of fructooligosaccharide.

[0071] The present invention will be described in detail below with reference to manufacturing examples and embodiments. However, the following manufacturing examples and embodiments are merely illustrative of the present invention, and the content of the present invention is not limited to the following manufacturing examples and embodiments.

[0073] Preparation Example 1. Prebiotic material (enzyme-treated apple pomace filtrate)

[0074] (1) The inedible parts (skin, seeds, etc.) were removed from the washed and sorted apples, and they were divided into 8 pieces.

[0075] (2) The apple prepared in step (1) above was immersed in an apple mint extract at 18-20°C for 1 minute, then removed and washed. The apple mint extract was prepared by adding 10 times (v / w) of purified water to apple mint and steeping it at 50-60°C for 1 hour.

[0076] (3) Based on the total weight of the steeping solution, 92% by weight of purified water, 3% by weight of hibiscus extract, 2% by weight of lemongrass extract, 2% by weight of Brussels sprouts juice, 0.5% by weight of citric acid, and 0.5% by weight of cyclodextrin were mixed to prepare the steeping solution. The hibiscus extract was prepared by adding 15 times (v / w) of purified water to hibiscus tea, extracting at 85–95°C for 10 minutes, and then filtering. Additionally, the lemongrass extract was prepared by adding 15 times (v / w) of purified water to lemongrass tea, extracting at 85–95°C for 10 minutes, and then filtering. Additionally, the Brussels sprouts juice was prepared by extracting juice from Brussels sprouts blanched at 100°C for 1 minute and then filtering.

[0077] (4) The washed apples from step (2) above were immersed in the soaking solution prepared in step (3) above at a ratio of 1:1.5 (w / v) at 18~20℃ for 5 minutes, then removed and aged (left) at 8~10℃ for 1 hour.

[0078] (5) The ripened apples from step (4) above were ground to an average particle size of 2 mm using a wet grinder, and then juiced using a juicer to separate them into apple juice pulp and apple juice.

[0079] (6) The separated apple juice residue from step (5) above was mixed with water in a 1:1 (w:v) ratio and then ground (grinde mill) to produce an apple juice residue mixture.

[0080] (7) The apple juice mixture prepared in step (6) above was sterilized at 80°C for 1 hour and then cooled down to 40°C.

[0081] (8) Enzymes were added to the cooled apple juice mixture from step (7) above and the mixture was reacted at 40°C for 1 hour at 30 rpm. The mixture was then filtered through a juicer (100 mesh) and concentrated with a concentrator until it reached 50% (v / v) to produce an enzyme-treated apple juice filtrate. The enzymes added were Pectinex Ultra SP-L and Ultimase MFC (Table 1) at 24.11 µl / g and 4.88 µl / g, respectively.

[0082] Enzyme types and activity Product Enzyme Dosage(mL / 1,000 kg) Activity Ultimase MFC β-GlucanaseXylanaseCellulase 40~60 300 AGU / mL Pectinex Ultra SP-L PectinaseHemicellulaseB-Glucanase 300 3,300 PGNU / g

[0084] Preparation Example 2. ABC juice

[0085] (1) After removing the inedible parts (peeling, etc.) from washed and sorted carrots, carrot puree was prepared by grinding them to an average particle size of 2 mm using a wet grinder.

[0086] (2) The carrot puree prepared in step (1) above was juiced using a juicer to produce carrot juice.

[0087] (3) After removing the inedible parts (skin, seeds, etc.) from the washed and sorted beets, the beets were ground to an average particle size of 2 mm using a wet grinder, and then juiced using a juicer to produce beet juice.

[0088] (4) 25 g of the enzyme-treated apple juice residue filter of Preparation Example 1, 200 g of the separated apple juice from step (5) of Preparation Example 1, 100 g of the carrot puree prepared in step (1) of Preparation Example 2, 100 g of the carrot juice prepared in step (2) of Preparation Example 2, 100 g of the beet juice prepared in step (3) of Preparation Example 2, 100 g of distilled water, and 53.95 g of fructooligosaccharide were mixed, stirred at 30 rpm, and sterilized at 85°C for 10 minutes and packaged.

[0090] Comparative Example 1. Apple pulp filtrate

[0091] (1) The inedible parts (skin, seeds, etc.) were removed from the washed and sorted apples, and they were divided into 8 pieces.

[0092] (2) The apple prepared in step (1) above was ground to an average particle size of 2 mm using a wet grinder, and then juiced using a juicer to separate it into apple juice pulp and apple juice.

[0093] (3) The separated apple juice residue from step (2) above was mixed with water in a 1:1 (w:v) ratio and then ground (grinde mill) to produce an apple juice residue mixture.

[0094] (4) The apple juice mixture prepared in step (3) above was sterilized at 80°C for 1 hour, then cooled to 40°C, filtered through a juicer (100 mesh), and concentrated with a concentrator until it reached 50% (v / v).

[0096] Comparative Example 2. Enzyme-treated apple pulp filtrate

[0097] (1) The inedible parts (skin, seeds, etc.) were removed from the washed and sorted apples, and they were divided into 8 pieces.

[0098] (2) The apple prepared in step (1) above was ground to an average particle size of 2 mm using a wet grinder, and then juiced using a juicer to separate it into apple juice pulp and apple juice.

[0099] (3) Using the separated apple juice residue from step (2) above, an enzyme-treated apple juice residue filter was prepared by processing it in the same way as steps (6) to (8) of Preparation Example 1.

[0101] Comparative Example 3. Enzyme-treated apple pulp filtrate

[0102] (1) The inedible parts (skin, seeds, etc.) were removed from the washed and sorted apples, and they were divided into 8 pieces.

[0103] (2) The apple prepared in step (1) above was immersed in apple mint extract at 18~20℃ for 1 minute, then removed and washed.

[0104] (3) The washed apples from step (2) above were ground to an average particle size of 2 mm using a wet grinder, and then juiced using a juicer to separate them into apple juice pulp and apple juice.

[0105] (4) Using the separated apple juice residue from step (3) above, an enzyme-treated apple juice residue filter was prepared by processing it in the same way as steps (6) to (8) of Preparation Example 1.

[0107] Comparative Example 4. Enzyme-treated apple pulp filtrate

[0108] (1) The inedible parts (skin, seeds, etc.) were removed from the washed and sorted apples, and they were divided into 8 pieces.

[0109] (2) Based on the total weight of the soaking solution, 92% by weight of purified water, 3% by weight of hibiscus extract, 2% by weight of lemongrass extract, 2% by weight of Brussels sprouts juice, 0.5% by weight of citric acid and 0.5% by weight of cyclodextrin were mixed to prepare the soaking solution.

[0110] (3) The apple prepared in step (1) above was immersed in the soaking solution prepared in step (2) above at a ratio of 1:1.5 (w / v) at 18~20℃ for 5 minutes, then removed and aged (left) at 8~10℃ for 1 hour.

[0111] (4) The ripened apples from step (3) above were ground to an average particle size of 2 mm using a wet grinder, and then juiced using a juicer to separate them into apple juice pulp and apple juice.

[0112] (5) Using the separated apple juice residue from step (4) above, an enzyme-treated apple juice residue filter was prepared by processing it in the same way as steps (6) to (8) of Preparation Example 1.

[0114] Comparative Example 5. Apple pulp filtrate

[0115] (1) The inedible parts (skin, seeds, etc.) were removed from the washed and sorted apples, and they were divided into 8 pieces.

[0116] (2) The apple prepared in step (1) above was immersed in apple mint extract at 18~20℃ for 1 minute, then removed and washed.

[0117] (3) Based on the total weight of the soaking solution, 92% by weight of purified water, 3% by weight of hibiscus extract, 2% by weight of lemongrass extract, 2% by weight of Brussels sprouts juice, 0.5% by weight of citric acid and 0.5% by weight of cyclodextrin were mixed to prepare the soaking solution.

[0118] (4) The washed apples from step (2) above were immersed in the soaking solution prepared in step (3) above at a ratio of 1:1.5 (w / v) at 18~20℃ for 5 minutes, then removed and aged (left) at 8~10℃ for 1 hour.

[0119] (5) The ripened apples from step (4) above were ground to an average particle size of 2 mm using a wet grinder, and then juiced using a juicer to separate them into apple juice pulp and apple juice.

[0120] (6) The separated apple juice residue from step (5) above was mixed with water in a 1:1 (w:v) ratio and then ground (grinde mill) to produce an apple juice residue mixture.

[0121] (7) The apple juice mixture prepared in step (6) above was sterilized at 80°C for 1 hour, then placed in a juicer (100 mesh) and filtered, and then concentrated with a concentrator until it reached 50% (v / v).

[0122] Comparison of Apple Juice Paste Processing Processes Processing process Preparation Example 1 Comparative Example 1 Comparative Example 2 Comparative Example 3 Comparative Example 4 Comparative Example 5 Apple mint soak ○ × × ○ × ○ Immersion treatment ○ × × × ○ ○ Enzyme treatment ○ × ○ ○ ○ ×

[0124] Experimental method

[0125] (1) Particle size analysis

[0126] The supernatant obtained by centrifuging enzyme-treated apple pomace filtrate at 3000×g for 20 minutes using a centrifuge (LaboGene 1580R, GUROZEN Co. Ltd., Seoul, Korea) was diluted 8-fold with distilled water and stirred for 5 minutes using a vortex mixer to prepare the sample. 2 mL of the prepared sample was transferred to a polystyrene cuvette, and the hydrodynamic diameter of the particles was measured using a DLS (SZ-100, HORIBA Instruments Inc., Japan).

[0128] (2) Acceptance analysis

[0129] For the measurement of water solubility, the sample was freeze-dried using a freeze-dryer (LP-100, Ilshinbiobase Co., Ltd, Dongducheon, Korea). 7.5 g of the freeze-dried sample and 92.5 mL of distilled water were mixed with a vortex mixer for 10 minutes and stored at room temperature for 24 hours. Subsequently, 20 mL of the supernatant was collected, dried at 105°C for 24 hours, and its weight was measured. The formula for calculating water solubility is as follows.

[0130] Solubility (%) = (Initial solution concentration / Supernatant concentration after 24 hours) × 100

[0132] (3) Relative molecular weight analysis

[0133] Relative molecular weight was measured using a Gel Permeation Chromatography (GPC, Waters 1515, Waters, Milford, USA) with two columns, TSKgel G5000PWXL and TSKgel G3000PWXL (Tosoh Bioscience, Tokyo, Japan), connected in series. Samples were prepared by diluting the solution 10-fold with distilled water, filtering it through a 0.45 µm filter (PVDF, Whatman Co. Ltd., Maidstone, UK), and then analyzing the solution. Pullulan (Pullulan standard set, Sigma Aldrich, Burlington, USA) was used as the standard. The mobile phase used was a solvent mixture of 0.1 M sodium nitrate and 0.05% sodium azide, the flow rate was measured at 0.7 mL / min, and detection was performed using an RID (Refractive Index Detector, Waters 2414, Waters, Milford, USA).

[0135] (4) Analysis of the growth rate of live probiotics

[0136] The increase rate of probiotic viable cell count was prepared by drying the sample with a freeze dryer and replacing 20 g / L glucose, the carbon source of the MBS medium, with an equal amount of the sample (enzyme-treated apple juice pulp) and a prebiotic prototype (Chong Kun Dang), respectively, to prepare the MRS medium.

[0137] In addition, ABC juices were prepared with or without the addition of enzyme-treated apple pulp, and then mixed with the juices in a 9:1 ratio in MRS medium. In each of the MRS media prepared above, Lactobacillus casei, a beneficial intestinal bacterium ( Lactobacillus casei After inoculating and culturing at 37°C for 24 hours, viable cells were counted and the average was expressed as CFU / mL.

[0139] (5) Analysis of short-chain fatty acid content

[0140] 1 mL of meta-phosphoric acid was added to 50 mg of fecal matter on day 6 of mice (BALB / C, female, 5 weeks old) orally administered a normal diet (untreated, control), enzyme-untreated apple pomace filtrate (0 h), enzyme-treated apple pomace filtrate (1 h), and 1 h + fructooligosaccharide, respectively. The mixture was stirred using a vortex mixer for 5 minutes, left overnight at room temperature, and sonicated for 10 minutes. The extracted samples were centrifuged at 1,350 rpm for 30 minutes (LaboGene 1580R, GUROZEN Co. Ltd., Seoul, Korea), and only the supernatant was collected for analysis. The supernatant was filtered using a 0.45 µm syringe filter (PTFE-H, HYUNDAI MICRO Co. Ltd, Seoul, Korea), transferred to a GC vial, and injected 1 µl into a GC / FID (FID-7890B, Agilent Technologies Inc., Santa Clara, USA) system. An HP-INNOWAX column (30 m × 250 µm × 0.25 µm, 19091N-133, Agilent Technologies Inc., Santa Clara, USA) was used, and the analysis was performed with an inlet temperature of 260°C, a flow rate of 1 mL / min of nitrogen gas, an inlet temperature of 80°C for 3 minutes and increased to 230°C at a rate of 10°C / min, a detector temperature of 230°C, and a split ratio of 10:1. As standard substances, acetic acid, propionic acid, and butyric acid (99%, Sigma Aldrich Co. Ltd, Seoul, Korea) were diluted to meta-phosphoric acid and analyzed at different concentrations.

[0142] (6) Changes over time

[0143] Changes over time were determined by periodically analyzing sensory evaluation, bacterial count, coliforms, lead, and cadmium for 72 days while storing samples in incubators at 15 and 25°C.

[0144] Sensory evaluation was conducted in accordance with Section 8. General Test Methods, 1. General Test Methods for Food, 1.1. Appearance (Sensory Evaluation) of the "Standards and Specifications for Food." Color, flavor, texture, and appearance were scored using a 5-point scale according to their respective scoring criteria (Good: 5 points; generally good: 4 or 3 points depending on the degree; poor: 2 points; significantly poor: 1 point).

[0145] The bacterial count was determined according to Section 8. General Test Methods, 4. Microbiological Test Methods, 4.5. Bacterial Count, and 4.5.1. General Bacterial Count, Subsection A. Standard Plate Method of the *Standards and Specifications for Food*. A test solution was prepared by placing 25 g of the sample into a sterile filter bag, adding 9 times the amount of sterile physiological saline, and shaking. 1 mL of the test solution and 1 mL of each dilution step were placed into a sterile Petri dish. Standard agar medium maintained at approximately 43–45°C was dispensed, and the sample and medium were thoroughly mixed and allowed to solidify. To inhibit the formation of spreading colonies, an additional 3–5 mL of standard agar medium was added and overlaid. The solidified Petri dish was then inverted and incubated at 35 ± 1°C for 48 ± 2 hours. For the calculation of the colony count, plates with no spreading colonies and 15–300 colonies per plate were selected and counted. The sterility of the test procedure was confirmed by using 1 mL of the same diluted solution without the test solution as a control solution.

[0146] The coliform bacteria were tested according to Section 8. General Test Methods, 4. Microbiological Test Methods, 4.7. Coliform bacteria, 4.7.2. Quantitative Tests, c. Dry Film Method of the "Standards and Specifications for Food." 25 g of the sample was placed in a sterile filter bag, and 9 times the amount of sterile physiological saline was added to prepare the test solution. 1 mL of the test solution and 1 mL of each step dilution were inoculated onto the coliform bacteria dry film medium and allowed to absorb well. After incubation at 35±1℃ for 24±2 hours, the number of red colonies that formed bubbles around them was counted, and the number of coliform bacteria was calculated by multiplying the average number of colonies by the dilution factor.

[0147] Lead was tested according to Section 8. General Test Methods, 9. Hazardous Substances in Food, 9.1.2. Lead (Pb) of the Standards and Specifications for Food, and cadmium was tested according to Section 8. General Test Methods, 9. Hazardous Substances in Food, 9.1.3. Cadmium (Cd) of the Standards and Specifications for Food.

[0149] (7) Analysis of general components and dietary fiber content

[0150] General component analysis was performed using Section 8. General Test Methods, 2. Test Methods for Food Components, and 2.1 General Component Test Methods of the Standards and Specifications for Food. Moisture was measured using Sections 2.1.1. Moisture and 2.1.1.1. Loss on Drying, and ash was measured using Section 2.1.2. Ash. Crude protein was measured using Sections 2.1.3. Nitrogen Compounds and 2.1.3.1. Total Nitrogen and Crude Protein, and crude fiber was measured using Sections 2.1.4. Carbohydrates and 2.1.4.2. Crude Fiber. Crude fat was measured using Sections 2.1.5. Lipids and 2.1.5.1. Crude Fat.

[0151] Dietary fiber content was analyzed by referring to Section 8. General Test Methods, 2. Test Methods for Food Components, 2.1. General Test Methods for Components, 2.1.4. Carbohydrates, 2.1.4.3. B. Quantification of Soluble Dietary Fiber by Liquid Chromatography of Dietary Fiber in the Standards and Specifications for Food.

[0153] Example 1. Particle size of enzyme-treated apple pulp filtrate

[0154] The particle size of the apple pomace filtrate treated with enzymes for 1 hour according to pH conditions is as shown in Fig. 1. As a result, the apple pomace filtrate of the present invention exhibited a relatively small particle size of 194.52 nm at pH 5.

[0156] Example 2. Water solubility of apple pulp filtrate according to enzyme treatment time

[0157] Adding enzymes to an apple pulp mixture according to the enzyme reaction time The results of comparing the water solubility of apple pomace filtrate are shown in Figure 2. As a result, compared to the apple pomace filtrate not treated with enzymes, the water solubility of the enzyme-treated apple pomace filtrate increased with a significant difference, and there was no significant difference depending on the enzyme treatment time.

[0159] Example 3. Relative molecular weight of apple pulp filtrate according to enzyme treatment time

[0160] Adding enzymes to an apple pulp mixture according to the enzyme reaction time The results of comparing the molecular weight of the apple pomace filtrate are shown in Figure 3 and Table 3. As a result, peak differences were observed between the filtrate without enzyme treatment (0 h) and the filtrate with enzyme treatment, and no significant difference was observed depending on the enzyme treatment time. In addition, peaks were observed at 2,465 and 485 Da in all filtrates, and a substance peak at 1,074,368 Da was found in the filtrate without enzyme treatment. Furthermore, in the filtrate with enzyme treatment, peaks around 3,878 and 1,576 Da, which were not found in the filtrate without enzyme treatment, were confirmed, and the peak around 2,465 Da increased. This suggests that the substance around 1,074,368 Da, which was observed in the filtrate without enzyme treatment, was generated through hydrolysis by the enzyme.

[0161] Relative molecular weight of apple pulp filtrate Peak No. Peak time(min) Molecular weight (Da) 1 17.62 1,074,368 2 27.30 3,878 3 28.08 2,465 4 28.85 1,576 5 30.88 485

[0163] Example 4. Analysis of the growth rate of viable probiotic cells in enzyme-treated apple pomace filtrate

[0164] Lactobacillus casei, known as a beneficial gut bacterium Lactobacillus casei The proliferation effect of the enzyme-treated apple pomace filtrate (APH) and APH-added ABC juice (APHJ) of the present invention on beneficial bacteria was confirmed using the strain. First, as compared in Figure 4 (A), the viable cell counts of the prebiotic prototype (Control) and APH were 8.0 × 10⁶, respectively. 8 and 9.7×10 8 In terms of CFU / mL, the enzyme-treated apple juice residue filtrate of the present invention was found to be about 21.3% higher.

[0165] In addition, as compared in (B) of Fig. 4, the ABC juice with added enzyme-treated apple pomace (APHJ) had 4.0 × 10, respectively, compared to the ABC juice without added enzyme-treated apple pomace (Control). 8 and 5.9×10 8 It increased by 47.5%. This indicates that the hydrolysate produced by enzymatically treating apple pomace acts as a prebiotic to promote the growth of beneficial bacteria in the intestines.

[0167] Example 5. Concentration of short-chain fatty acids in enzyme-treated apple pomace filtrate

[0168] Short-chain fatty acids (SCFAs) produced by intestinal microorganisms are absorbed into colon tissue and act as anions, serving various purposes such as host energy sources and immune modulation, and are reported to be effective in anti-inflammatory, intestinal motility, and anticancer effects. The content of short-chain fatty acids in the feces of mice fed a normal diet (control), an enzyme-free apple pomace filtrate (0 h), an enzyme-treated apple pomace filtrate (1 h), and a diet administered orally with 1 h plus fructooligosaccharide on day 6 was analyzed using gas chromatography. The concentrations of the generated short-chain fatty acids, namely acetic acid, butyric acid, and propionic acid, and the sum of their concentrations were calculated as the total short-chain fatty acid concentration.

[0169] The acetic acid content showed a significant difference in the feces of mice fed a normal diet, mice orally administered untreated apple juice (0 h), mice orally administered enzyme-treated apple juice (1 h), and mice orally administered enzyme-treated apple juice + fructooligosaccharide (1 h + FOS), with the highest content observed in the feces of mice orally administered enzyme-treated apple juice + fructooligosaccharide. In the case of propionic acid, a significant difference was also observed in the feces of mice orally administered enzyme-treated apple juice (1 h) and mice orally administered enzyme-treated apple juice + fructooligosaccharide (1 h + FOS), with the highest content observed in the feces of mice orally administered enzyme-treated apple juice + fructooligosaccharide (1 h + FOS). The butyric acid content showed a tendency to increase with changes in diet, but statistical analysis results showed no significant difference. The total short-chain fatty acid content was found to be high in the feces of mice orally administered enzyme-treated apple juice (1 h) and enzyme-treated apple juice + fructooligosaccharide (1 h + FOS) together (Fig. 5). As a result of this, it is determined that the enzyme-treated apple juice filter of the present invention can help improve intestinal health by increasing beneficial intestinal bacteria that promote short-chain fatty acid metabolism.

[0171] Example 6. Changes in ABC juice over time

[0172] Using enzyme-treated apple pomace added ABC juice (APHJ), changes over time were measured using established quality indicators during a 72-day actual measurement period at 15℃ and 25℃.

[0173] Korean quality limit standards for setting expiration dates Quality parameters Quality Limits Sensory evaluation (5-point scale) ≥3, cannot because there is 1 Total bacterial count (CFU / g) ≤100 E. coli count (CFU / g) ≤10 per 1 g Lead content (mg / kg) ≤0.05 Cadmium content (mg / Kg) ≤0.1

[0175] As a result, since no items reached the quality limit for 72 days at 15℃ and 25℃, the shelf life can be set within the experimental period. Considering external factors during the distribution process, the actual shelf life may be shortened, so the experimental period (72 days) × safety factor (0.85) = 61.2 days. Therefore, it was determined that the shelf life of APHJ is at least 2 months at room temperature.

[0176] Changes in quality parameters of ABC juice at 15℃ (±1℃) Storage period (days) Sensory evaluation Total bacterial count (CFU / g) E. coli count (CFU / g) Lead (mg / Kg) Cadmium (mg / Kg) 0 5 0 0 0.05 0 3 5 0 0 6 5 0 0 10 5 0 0 13 5 0 0 17 5 0 0 19 5 0 0 21 5 0 0 24 5 0 0 27 5 0 0 31 5 0 0 34 5 0 0 38 5 0 0 40 5 0 0 42 5 0 0 45 5 0 0 48 5 0 0 52 5 0 0 55 5 0 0 59 5 0 0 61 5 0 0 63 5 0 0 66 5 0 0 69 5 0 0 72 5 0 0

[0178] Changes in quality parameters of ABC juice at 25℃ (±1℃) Storage period (days) Sensory evaluation Total bacterial count (CFU / g) E. coli count (CFU / g) Lead (mg / Kg) Cadmium (mg / Kg) 0 5 0 0 0.05 0 3 5 0 0 6 5 0 0 10 5 0 0 13 5 0 0 17 5 0 0 19 5 0 0 21 5 0 0 24 5 0 0 27 5 0 0 31 5 0 0 34 5 0 0 38 5 0 0 40 5 0 0 42 5 0 0 45 5 0 0 48 5 0 0 52 5 0 0 55 5 0 0 59 5 0 0 61 5 0 0 63 5 0 0 66 5 0 0 69 5 0 0 72 5 0 0

[0180] Example 7. General components and dietary fiber content of ABC juice

[0181] As a result of analyzing the general composition and dietary fiber content of enzyme-treated apple pulp added ABC juice, it was found to contain 87.35% moisture, 0.44% crude protein, 0.41% crude fat, 0.35% crude ash, and 0.28% crude fiber, and the dietary fiber content was analyzed to be 2.49%.

[0182] General ingredients and dietary fiber content of ABC juice (g / 100 mL) division Content (%) moisture content 87.35±0.03 crude fat 0.41±0.01 crude protein 0.44±0.01 Views 0.35±0.01 crude fiber 0.28±0.01 Dietary fiber content (%) 2.49±0.01

[0184] Example 8. Sensory evaluation of apple pomace filtrate prepared by different processing methods

[0185] Sensory evaluation was conducted using the apple pomace filtrate of Preparation Example 1 and Comparative Examples 1 to 5. The sensory evaluation was performed on 45 trained sensory evaluators who consumed each filtrate. The evaluation items included aroma, taste, and overall preference. Using a 5-point scale with a maximum score of 5, subjects recorded their scores according to the following evaluation criteria, and the average value was calculated and recorded. 5: Very good, 4: Good, 3: Average, 2: Poor, 1: Very bad.

[0186] Sensory evaluation results of apple pulp filtrate type incense taste Overall preference Preparation Example 1 4.4 4.5 4.4 Comparative Example 1 3.6 3.6 3.6 Comparative Example 2 3.7 3.7 3.7 Comparative Example 3 3.9 3.9 4.0 Comparative Example 4 3.8 4.0 3.9 Comparative Example 5 3.9 3.8 3.8

[0187] As a result, as can be seen in Table 8 above, Preparation Example 1 showed the highest score in all categories compared to the apple pomace filtrate of the comparative examples. Therefore, it was confirmed that preparing apple pomace filtrate by pre-treating the materials and then enzymatically treating them, as in Preparation Example 1, results in prebiotic functionality while further improving aroma, taste, and palatability.

Claims

Claim 1 delete Claim 2 delete Claim 3 (1) a step of immersing an apple in an apple mint extract, removing it, and washing it; (2) a step of preparing an immersion solution by mixing water, hibiscus extract, lemongrass extract, Brussels sprout juice, citric acid, and cyclodextrin; (3) a step of immersing the washed apple from step (1) in the immersion solution prepared in step (2), removing it, and aging it; (4) a step of crushing and extracting juice from the aged apple from step (3) to separate the extracted pulp and the extracted liquid; (5) a step of mixing water with the separated extracted pulp from step (4) and crushing it to prepare an apple extracted pulp mixture; (6) a step of sterilizing and cooling the apple extracted pulp mixture prepared in step (5); and (7) a step of adding an enzyme to the cooled apple extracted pulp mixture from step (6) to cause an enzymatic reaction and then filtering it. Claim 4 A method for manufacturing a prebiotic material according to claim 3, wherein the soaking solution of step (2) is prepared by mixing 91-93% by weight of water, 2-4% by weight of hibiscus extract, 1-3% by weight of lemongrass extract, 1-3% by weight of Brussels sprouts juice, 0.2-0.8% by weight of citric acid and 0.2-0.8% by weight of cyclodextrin, based on the total weight of the soaking solution. Claim 5 In claim 3, (1) a step of immersing an apple in an apple mint extract at 18–20°C for 30–90 seconds, then removing it and washing it; (2) a step of preparing an immersion solution by mixing, based on the total weight of the immersion solution, 91–93% by weight of water, 2–4% by weight of hibiscus extract, 1–3% by weight of lemongrass extract, 1–3% by weight of Brussels sprouts juice, 0.2–0.8% by weight of citric acid, and 0.2–0.8% by weight of cyclodextrin; (3) a step of immersing the washed apple from step (1) in the immersion solution prepared in step (2) at a ratio of 0.8–1.2:1.3–1.7 (w / v) at 18–20°C for 4–6 minutes, then removing it and aging it at 8–10°C for 50–70 minutes; (4) crushing and juicing the aged apple from step (3). A method for manufacturing a prebiotic material characterized by comprising the steps of: (5) separating the extracted pulp and the extracted liquid; (6) mixing water with the separated extracted pulp from step (4) in a ratio of 0.8~1.2:0.8~1.2 (w:v) and then grinding to produce an apple extracted pulp mixture; (6) sterilizing the apple extracted pulp mixture produced in step (5) at 70~90℃ for 50~70 minutes and then cooling it to 35~45℃; and (7) adding an enzyme to the cooled apple extracted pulp mixture from step (6) and performing an enzymatic reaction at 35~45℃ for 30~90 minutes and then filtering. Claim 6 A prebiotic material manufactured by the method of any one of paragraphs 3 to 5. Claim 7 A method for preparing ABC juice characterized by mixing the prebiotic material of claim 6 with apple juice, carrot puree, carrot juice, beet juice, water, and fructooligosaccharide.

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