Preparation method of corncob extract and its application in probiotics

Abstract

The present invention relates to a preparation method of corncob extract and its application in probiotics, belonging to the technical field of functional foods. The present invention provides a preparation method of corncob extract, firstly drying and grinding corncob, adding water for extraction, concentrating the liquid phase obtained by the first solid-liquid separation, adding anhydrous ethanol for mixing, performing a second solid-liquid separation to obtain a solid phase, and obtaining a water-soluble extract of corncob; then drying the solid phase obtained by the first solid-liquid separation, adding a mixed solution of an alkaline solution and hydrogen peroxide for heating, performing solid-liquid separation to obtain a liquid phase, concentrating the obtained liquid phase, adding anhydrous ethanol for mixing, performing solid-liquid separation to obtain a solid phase, and obtaining a water-insoluble extract of corncob; finally, mixing the water-soluble extract of corncob with the water-insoluble extract, and obtaining a corncob extract. The corncob extract obtained in the present invention can not only promote the growth of probiotics, have a greater influence on the acidification activity of probiotics, but also have a stronger promoting effect on the short-chain fatty acids of probiotics.

Description

Technical Field

[0001] The invention belongs to the technical field of functional foods, and particularly relates to a preparation method of a corncob extract and application of the corncob extract in probiotics. Background Art

[0002] Corn is an annual herbaceous plant of the genus Zea in the family Poaceae. It is an important food crop and feed crop, and is also the crop with the highest total output in the world. Its planting area and total output are second only to rice and wheat. Corncob is the cob of corn after the kernels are removed from the ear. It is mainly composed of 35%-40% hemicellulose, 32%-36% cellulose, 17%-20% lignin and 1.2%-1.8% ash.

[0003] At present, corn cobs in my country are mainly used for pig feed, cardboard, cement board, cement brick production, etc. The functional components in corn cobs are not fully utilized, which causes waste of the functional components in corn cobs. Therefore, by extracting and applying the functional components from corn cobs, the remaining corn cob residues can be used as usual for cardboard, pig feed, etc., thereby improving the utilization value of corn cobs.

[0004] Therefore, it is necessary to study the differences in the physicochemical properties and beneficial activities of corncob extracts from different varieties, so as to facilitate their widespread application in the food and pharmaceutical industries and generate good social and economic benefits. Summary of the invention

[0005] In order to solve the above technical problems, the present invention provides a preparation method of corn cob extract and its application in probiotics. By extracting corn cobs of different varieties, corn cob extracts that can promote the proliferation of probiotics, promote acidification activity and promote the increase of short-chain fatty acids are prepared.

[0006] The present invention is achieved through the following technical solutions:

[0007] The first object of the present invention is to provide a method for preparing a corncob extract, comprising the following steps:

[0008] (1) drying and grinding the corn cob, adding water for extraction, performing a first solid-liquid separation to obtain a solid phase and a liquid phase, concentrating the obtained liquid phase, adding anhydrous ethanol for mixing, performing a second solid-liquid separation to obtain a solid phase, drying the obtained solid phase, removing starch, and obtaining a water-soluble extract of the corn cob;

[0009] (2) drying the solid phase obtained by the first solid-liquid separation in step (1), adding a mixed solution of an alkaline solution and hydrogen peroxide to heat, performing solid-liquid separation to obtain a liquid phase, concentrating the obtained liquid phase, adding anhydrous ethanol to mix, performing solid-liquid separation to obtain a solid phase, drying the obtained solid phase, removing starch and protein, and obtaining a water-insoluble extract of corn cob;

[0010] (3) Mixing the corncob water-soluble extract obtained in step (1) and the water-insoluble extract obtained in step (2) to obtain a corncob extract.

[0011] In one embodiment of the present invention, in step (1), the corn cob is selected from glutinous corn cobs and / or sweet corn cobs.

[0012] In one embodiment of the present invention, the corn variety of the glutinous corn cob is selected from one or more of Zhongnuo 2, Shenkenuo 602 and Huhongnuo 1; the corn variety of the sweet corn cob is selected from one or more of Jinyin 208, Xuetian 7401 and Shenxuetian 1.

[0013] In one embodiment of the present invention, in step (1), water is added at a solid-liquid ratio of 1:15-1:20.

[0014] In one embodiment of the present invention, in step (1), the extraction conditions are: extraction at 80°C-88°C for 2h-2.5h.

[0015] In one embodiment of the present invention, in step (1), anhydrous ethanol is added at a material-liquid ratio of 1:3-1:4.

[0016] In one embodiment of the present invention, the alkaline solution is selected from sodium hydroxide solution and / or potassium hydroxide solution.

[0017] In one embodiment of the present invention, in step (2), a mixed solution of alkaline solution and hydrogen peroxide is added at a material-liquid ratio of 1:15-1:20.

[0018] The second object of the present invention is to provide the corncob extract prepared by the preparation method.

[0019] The third object of the present invention is to provide the application of the corncob extract in probiotics.

[0020] Mechanism of the present invention:

[0021] (1) Extraction mechanism of corncob extract: Water-soluble extracts are soluble in water and can be easily extracted by hot water. Water-insoluble extracts are entangled with cellulose in corncobs and covalently linked to lignin, making them difficult to extract by water. Alkaline solutions can separate the extracts from cellulose, while hydrogen peroxide can oxidize lignin, allowing the water-insoluble extracts to be extracted.

[0022] (2) Mechanism in probiotics: Probiotics contain enzymes such as β-xylanase that can degrade corncob extracts. Probiotics can use the carbon source after the degradation of corncob extracts as nutrients to achieve their own growth. Corncob extracts contain water-soluble and water-insoluble extracts, which can produce synergistic effects when applied to probiotics. The water-soluble extracts and water-insoluble extracts can interact with each other to increase the activity of enzymes in probiotics, which is more conducive to promoting the growth of probiotics.

[0023] The above technical solution of the present invention has the following advantages compared with the prior art:

[0024] The present invention provides a method for preparing a corncob extract and application of the corncob extract in probiotics. The corncob extract is extracted from corncobs of different varieties, wherein both the water-soluble extract and the water-insoluble extract contain components such as arabinose, xylose, galactose, glucose, mannose and Ara / Xyl. The mixed corncob extract of the present invention can not only promote the growth of probiotics and have a greater influence on the acidification activity of probiotics, but also has a stronger promoting effect on the short-chain fatty acids of the probiotics. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] In order to make the content of the present invention more clearly understood, the present invention is further described in detail below according to specific embodiments of the present invention in conjunction with the accompanying drawings, wherein

[0026] Figure 1 is a scanning electron microscope image of the corncob extract in Test Example 3 of the present invention;

[0027] Figure 2 The effect of corncob extract on the growth of probiotics in Test Example 4 of the present invention; wherein, Figure 2 -A is the effect of water-soluble extract on the growth of Lactobacillus plantarum; Figure 2 -B is the effect of water-insoluble extract on the growth of Lactobacillus plantarum; Figure 2 -C is the effect of water-soluble extract on the growth of animal Bifidobacterium; Figure 2 -D is the effect of water-insoluble extract on the growth of animal Bifidobacterium;

[0028] Figure 3 is the effect of corncob extract on probiotic acidification activity in Test Example 4 of the present invention; wherein Figure 3 -A is the effect of water-soluble extract on the acidification activity of Lactobacillus plantarum; Figure 3 -B is the effect of water-insoluble extract on the acidification activity of Lactobacillus plantarum; Figure 3 -C is the effect of water-soluble extract on the acidification activity of animal Bifidobacterium; Figure 3 -D is the effect of water-insoluble extract on the acidification activity of animal Bifidobacterium;

[0029] Figure 4 The effect of the corncob extract on the promotion of acetic acid increase by Lactobacillus plantarum in Test Example 5 of the present invention;

[0030] Figure 5 It is the effect of corncob extract on the promotion of propionic acid increase by Lactobacillus plantarum in Test Example 5 of the present invention;

[0031] Figure 6 The effect of the corncob extract on the promotion of butyric acid increase by Lactobacillus plantarum in Test Example 5 of the present invention;

[0032] Figure 7 The effect of the corncob extract on the promotion of acetic acid increase by Lactobacillus plantarum in Test Example 5 of the present invention;

[0033] Figure 8 It is the effect of corncob extract on the promotion of propionic acid increase by Lactobacillus plantarum in Test Example 5 of the present invention;

[0034] Fig. 9 This is the effect of the corncob extract on the promotion of butyrate increase by Lactobacillus plantarum in Test Example 5 of the present invention. DETAILED DESCRIPTION

[0035] The present invention is further described below in conjunction with the accompanying drawings and specific embodiments so that those skilled in the art can better understand the present invention and implement it, but the embodiments are not intended to limit the present invention.

[0036] Example 1

[0037] This embodiment provides a method for preparing a corncob extract, which specifically comprises the following steps:

[0038] (1) Waxy corn cobs (Zhongnuo No. 2 ZW, Shenkenuo No. 602SW, Huhongnuo No. 1 HW) were dried and ground into powder. The mixture was extracted with hot water at 80°C for 2 h at a solid-liquid ratio of 1:15 (w / v), and the supernatant was obtained by centrifugation. The supernatant was concentrated, and four times the volume of anhydrous ethanol was added to precipitate polysaccharides. The polysaccharides were precipitated and freeze-dried after centrifugation. The starch was then removed by repeated freeze-thaw method to obtain water-soluble extracts, which were labeled as ZW E , SW E and HW E .

[0039] (2) The residue after the first centrifugation in step (1) was dried to constant weight, added to a solution of 0.16 mol / L NaOH and 1.5% H2O2 (solid-liquid ratio 1:15), heated at 80°C for 2 h, and centrifuged to obtain the supernatant. After the supernatant was concentrated, four times the volume of anhydrous ethanol was added to precipitate the polysaccharide, centrifuged and freeze-dried, and then the starch was removed by repeated freeze-thaw method, and the protein was removed by alkaline protease (Alcalase 2.4 L) to obtain water-insoluble extracts, which were labeled as ZW, respectively. U , SWU and HW U .

[0040] (3) Mixing the water-soluble extract obtained in step (1) with the water-insoluble extract obtained in step (2) to obtain a corncob extract.

[0041] Example 2

[0042] This embodiment provides a method for preparing a corncob extract, which specifically comprises the following steps:

[0043] (1) Sweet corn cobs (Jinyin 208JS, Xuetian 7401XS, Shenxuetian No. 1 SS) were dried and ground into powder. They were then extracted with hot water at 80°C for 2 h at a solid-liquid ratio of 1:15 (w / v), and the supernatant was collected by centrifugation. The supernatant was concentrated, and four times the volume of anhydrous ethanol was added to precipitate polysaccharides. The polysaccharides were precipitated and freeze-dried after centrifugation. The starch was then removed by repeated freeze-thaw method to obtain water-soluble extracts, which were labeled as JS and JS. E 、XS E and SS E .

[0044] (2) The residue after the first centrifugation in step (1) was dried to constant weight, added to a solution of 0.16 mol / L NaOH and 1.5% H2O2 (solid-liquid ratio 1:15), heated at 80°C for 2 h, and centrifuged to obtain the supernatant. After the supernatant was concentrated, four times the volume of anhydrous ethanol was added to precipitate the polysaccharide, centrifuged and freeze-dried, and then the starch was removed by repeated freeze-thaw method, and the protein was removed by alkaline protease (Alcalase 2.4 L) to obtain a water-insoluble extract, which was labeled as JS. U 、XS U and SS U .

[0045] (3) Mixing the water-soluble extract obtained in step (1) with the water-insoluble extract obtained in step (2) to obtain a corncob extract.

[0046] Comparative Example

[0047] Treat corn silk powder with 95% ethanol and air dry. Weigh 1 kg of corn silk powder in an extraction container, heat with distilled water (1:20, w / v) at 80°C for 2h, extract twice, filter and centrifuge to obtain the supernatant, concentrate to 1 / 3 of the original volume, add 4 times the volume of anhydrous ethanol to the concentrate, let stand at 4°C for 12h, centrifuge to obtain the precipitate, wash twice with anhydrous ethanol, collect the precipitate, vacuum freeze-dry, weigh and record, and calculate the polysaccharide yield.

[0048] Prepare 1% corn silk polysaccharide solution, add Sevag reagent, rotate and stir for 30 minutes at room temperature, let stand for 30 minutes, centrifuge to remove protein, and repeat the above steps 3 times. Add anhydrous ethanol to 80% of the polysaccharide solution after removing protein, let stand for 12 hours at 4°C, centrifuge and repeatedly freeze and thaw to remove starch to obtain corn silk polysaccharide (CSP).

[0049] Test Example 1

[0050] The yield and chemical composition of the water-soluble extract and water-insoluble extract prepared from different varieties of corn cobs in the examples and comparative examples were tested. The results are shown in Tables 1 and 2:

[0051] Table 1 Yield and chemical composition of water-soluble extracts and water-insoluble extracts

[0052]

[0053] Table 2

[0054]

[0055] na means not determined.

[0056] As can be seen from Tables 1 and 2, the neutral sugar content of corn cob extract is high, the uronic acid content is below 10%, and the protein content is less than 4%. In the comparison between water-soluble extracts and water-insoluble extracts, the neutral sugar content of water-soluble extracts is between 54.94%-77.77%, while the neutral sugar content of water-insoluble extracts is between 28.13%-51.32%. It is obvious that the neutral sugar content of water-soluble extracts is higher than that of water-insoluble extracts. The uronic acid content of water-insoluble extracts is higher than that of water-soluble extracts. In terms of monosaccharide composition, mannose is found in most water-soluble extracts. In Ara / Xyl, the Ara / Xyl of sweet corn cob extract is higher than that of waxy corn cob, indicating that the branching degree of arabinoxylan in the extract of sweet corn cob is higher than that of waxy corn. It can be seen that the polysaccharides in sweet corn extracts may be different from those in waxy corn in terms of structural characteristics.

[0057] Test Example 2

[0058] The average molecular weights of the water-soluble extracts and water-insoluble extracts prepared from different varieties of corn cobs in Example 1 were tested, and the results are shown in Table 3:

[0059] Table 3 Average molecular weight of corncob extract

[0060]

[0061] in, Indicates weight average molecular weight It represents the number average molecular weight.

[0062] As can be seen from Table 3, the polydispersity index (PDI, ) is small, indicating that the molecular weight of the corncob extract samples is homogeneous. On the other hand, among the water-soluble extracts, the water-soluble extract of sweet corn cob Greater than waxy corncob water-soluble extract.

[0063] Test Example 3

[0064] The microstructures of the water-soluble extracts and water-insoluble extracts prepared from different varieties of corn cobs in the examples were tested, and the results were as follows: Figure 1 As shown:

[0065] Figure 1 This is a 500× scanning electron micrograph of corncob extract. Figure 1 It can be seen that the structure of the water-soluble extract of corn cob is sponge-network-like, and its surface morphology is like that formed by countless sponges, while the water-insoluble extract is in sheet form and arranged in disorder.

[0066] Test Example 4

[0067] The probiotic activity of the water-soluble extracts and water-insoluble extracts prepared from different varieties of corn cobs and the corn cob extracts in the examples was tested. The results are as follows Figure 2 and Figure 3 As shown:

[0068] Figure 2 The growth curves of Lactobacillus plantarum and Bifidobacterium animalis were plotted. MRS medium (purchased from Beijing Solebow Technology Co., Ltd., model M8540) was used as a blank control, MRS medium containing 1% xylooligosaccharide was used as a positive control, and the experimental groups were MRS medium containing 1% corncob extract. After inoculation, anaerobic fermentation was performed, and OD was measured every 2 hours. 600 .Depend on Figure 2 It can be seen that the OD of all samples 600 The values ​​were higher than those of the blank group. Corncob extract significantly promoted the growth of animal Bifidobacterium and plantarum Lactobacillus. The water-insoluble extract had a stronger growth-promoting effect on probiotics (plantarum Lactobacillus and animal Bifidobacterium) than the water-soluble extract. The effect of sweet corncob extract on promoting the growth of probiotics was weaker than that of glutinous corncob extract. In the mixed group of water-soluble extract and water-insoluble extract (ZW E +ZW U ), the number of the two bacteria is the largest, indicating that they have the best effect in promoting the growth of probiotics.

[0069] Figure 3 Effect of corncob extract on the acidification activity of Lactobacillus plantarum and Bifidobacterium animalis. Anaerobic culture, pH was measured every 2 hours. Figure 3 The results showed that the pH of the fermentation broth of all samples was lower than that of the blank group, and the pH of the fermentation broth of the glutinous corncob extract was lower than that of the sweet corncob extract, indicating that the glutinous corncob extract had a greater effect on the acidification activity of the strain. The pH of the fermentation broth in the mixed group of water-soluble extract and water-insoluble extract was the lowest, indicating that the probiotics were promoted to produce a large amount of organic acids.

[0070] Test Example 5

[0071] Acetic acid, propionic acid, and butyric acid are the main short-chain fatty acids formed by in vitro fermentation and are typical representative products of in vitro fermentation of polysaccharides. Among them, the bacteria will secrete more acetic acid. In this test example, the content of short-chain fatty acids was determined by anaerobically fermenting Lactobacillus plantarum and Bifidobacterium animalis in the culture medium for 48 hours. The results are as follows Figure 4-9 As shown. Figure 4-9 It can be concluded that the content of acetic acid, propionic acid and butyric acid produced by the two probiotics in the experimental group and the positive control group (XOS, MRS medium with 1% xylo-oligosaccharide added) was higher than that in the blank control group, and corn cob extracts (ZW, SW, HW, JS, XS and SS) promoted the production of more acetic acid, propionic acid and butyric acid by the strains relative to corn silk polysaccharides. Among them, the content of acetic acid, propionic acid and butyric acid in the mixed group of water-soluble extract and water-insoluble extract was the highest.

[0072] Among the short-chain fatty acids produced by the strains, acetic acid had the highest content and propionic acid had the lowest content. The acetic acid produced by the waxy corn cob extract was higher than that produced by the sweet corn cob extract. Similarly, the acetic acid produced by the water-soluble extract was lower than that produced by the water-insoluble extract. There was no significant difference between the fermentation broth of the waxy corn cob extract and the fermentation broth of the sweet corn cob extract, and similarly, there was no significant difference between most of the water-soluble extracts and the water-insoluble extracts.

[0073] Water-soluble extracts may interact with water-insoluble extracts to increase the enzyme activity in probiotics, thereby producing more carbon sources for probiotics to utilize.

[0074] Obviously, the above embodiments are merely examples for clear explanation and are not intended to limit the implementation methods. For those skilled in the art, other different forms of changes or modifications can be made based on the above description. It is not necessary and impossible to list all the implementation methods here. The obvious changes or modifications derived from these are still within the protection scope of the invention.

Claims

1. An application of corncob extract in promoting the growth of animal bifidobacteria, characterized in that: The preparation method of the corncob extract comprises the following steps: (1) drying and grinding the corn cob, adding water for extraction, performing a first solid-liquid separation to obtain a solid phase and a liquid phase, concentrating the obtained liquid phase, adding anhydrous ethanol for mixing, performing a second solid-liquid separation to obtain a solid phase, drying the obtained solid phase, removing starch, and obtaining a water-soluble extract of the corn cob; (2) drying the solid phase obtained by the first solid-liquid separation in step (1), adding a mixed solution of an alkaline solution and hydrogen peroxide to heat, separating the solid and liquid to obtain a liquid phase, concentrating the obtained liquid phase, adding anhydrous ethanol to mix, separating the solid and liquid to obtain a solid phase, drying the obtained solid phase, removing starch and protein, and obtaining a water-insoluble extract of corn cob; (3) mixing the corncob water-soluble extract obtained in step (1) and the water-insoluble extract obtained in step (2) to obtain a corncob extract; In step (1), the extraction conditions are: 80°C-88°C for 2 h-2.5 h.

2. The use according to claim 1, characterized in that: In step (1), the corn cob is selected from glutinous corn cob and / or sweet corn cob.

3. The use according to claim 2, characterized in that: The corn variety of the glutinous corn cob is selected from one or more of Zhongnuo 2, Shenkenuo 602 and Huhongnuo 1; the corn variety of the sweet corn cob is selected from one or more of Jinyin 208, Xuetian 7401 and Shenxuetian 1.

4. The use according to claim 1, characterized in that: In step (1), water is added at a material-liquid ratio of 1:15-1:

20.

5. The use according to claim 1, characterized in that: In step (1), anhydrous ethanol is added according to a material-liquid ratio of 1:3-1:

4.

6. The use according to claim 1, characterized in that: The alkaline solution is selected from sodium hydroxide solution and / or potassium hydroxide solution.

7. The use according to claim 1, characterized in that: In step (2), a mixed solution of alkaline solution and hydrogen peroxide is added at a material-liquid ratio of 1:15-1:20.

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