CN115429813A

CN115429813A - Preparation method of Jerusalem artichoke oligosaccharide with blood lipid reducing effect and intestinal flora structure improving effect

Info

Publication number: CN115429813A
Application number: CN202211139322.5A
Authority: CN
Inventors: 姬妍茹; 张正海; 董艳; 杨庆丽; 关向军; 李国巍; 石杰; 魏连会; 高宇; 潘静; 高媛
Original assignee: Heilongjiang Academy of Sciences Daqing Branch
Current assignee: Heilongjiang Academy of Sciences Daqing Branch
Priority date: 2022-09-19
Filing date: 2022-09-19
Publication date: 2022-12-06

Abstract

A preparation method of jerusalem artichoke oligosaccharides with effects of reducing blood fat and improving intestinal flora structure belongs to the technical field of functional food preparation. The method comprises the following steps: slicing, dicing or dicing the black jerusalem artichoke according to the proportion of 1g: adding purified water in a proportion of 10-20 mL, homogenizing and homogenizing by adopting a dynamic ultrahigh-pressure micro-jet technology for auxiliary extraction, and filtering to obtain a crude polysaccharide extracting solution and filter residues; performing rotary evaporation and concentration on the crude polysaccharide extracting solution at the speed of 90-120 r/min, then freeze-drying, adding ethanol according to the amount which is 5 times of the mass-volume ratio (g: mL) of the freeze-dried powder, standing for 10-14 h, centrifuging to remove supernatant, adding 5 times of water for redissolution, removing protein by adopting a Sevage reagent, adding activated carbon according to the amount which is 2% of the mass of the solution for decoloring for 30-50 min, dialyzing by using a 300Da membrane to remove monosaccharide, and freeze-drying to obtain black jerusalem artichoke oligosaccharide powder. The jerusalem artichoke oligosaccharide prepared by the invention can adjust the blood fat level of a hyperlipemia model mouse, and increase the abundance of lactobacillus, bifidobacterium, lachnospiraceae, coprobacterium and uncultured Lachnospiraceae _ NKA136_ group flora.

Description

Preparation method of Helianthus tuberosus oligosaccharide with blood lipid reducing effect and intestinal flora improving effect

Technical Field

The invention belongs to the technical field of functional food preparation, and particularly relates to a preparation method of jerusalem artichoke oligosaccharides with the effects of reducing blood fat and improving intestinal flora structure.

Background

Hyperlipidaemia (HLP) is a condition in which the concentration of one or more lipid components in the plasma exceeds normal values due to disturbances in the lipid metabolism in the body. Hyperlipidemia is an important factor for inducing cardiovascular and cerebrovascular diseases, and the cardiovascular and cerebrovascular diseases have extremely high mortality rate and are generally regarded as important. With the improvement of the living standard of people, the change of diet structure disorder and life style, patients with hyperlipidemia tend to rise year by year. The treatment of the disease is mainly based on drug therapy at present, but the curative effect is not ideal, the common problems are that the blood fat is reduced during the taking process, the medicine is easy to rebound after stopping taking, and the harm to the body is large after the western medicine is taken for a long time. The mode of regulating blood fat by taking functional food is accepted by most patients, so the research and development of the functional food with the function of assisting in reducing blood fat becomes a research hotspot in the field of functional food.

The black jerusalem artichoke is a novel food which is processed by the process of the invention patent (201610124756.6) by taking jerusalem artichoke as a raw material, the components of the jerusalem artichoke are greatly changed after being processed into the black jerusalem artichoke, the fructan is degraded into monosaccharide, disaccharide and fructo oligosaccharide, the polyphenol content is obviously improved, and the melanoidin component is increased. The change of the nutrient components and the active substances endows the black jerusalem artichoke with various physiological functions. Earlier researches show that the black jerusalem artichoke polyphenol has good in-vitro antioxidant activity, has good effects of removing DPPH free radicals and hydroxyl free radicals, and has good reducing power on iron ions. The black jerusalem artichoke water extract can effectively promote intestinal peristalsis of constipation mice, regulate intestinal flora, and metabolize to generate acetic acid, propionic acid and n-butyric acid, thereby playing a role in relieving constipation. No research is carried out on the blood fat reducing effect of the jerusalem artichoke oligosaccharide.

Disclosure of Invention

The invention aims to solve the problems that the taking of western medicines for treating hyperlipemia causes medicine dependence and damages to the body, and provides a preparation method of jerusalem artichoke oligosaccharide with the effects of reducing blood fat and improving the intestinal flora structure.

In order to realize the purpose, the technical scheme adopted by the invention is as follows:

a preparation method of Helianthus tuberosus oligosaccharides with blood lipid reducing effect and intestinal flora structure improving effect comprises the following steps:

the method comprises the following steps: preparing the oligosaccharide of black jerusalem artichoke: slicing (1.0-2.0 cm), dicing or slicing (as small as possible, beneficial to extraction) black jerusalem artichoke according to the weight ratio of 1g: adding purified water in a proportion of 10-20 mL, homogenizing and homogenizing by adopting a dynamic ultrahigh pressure microjet technology for auxiliary extraction, and filtering by adopting six layers of gauze to obtain a crude polysaccharide extracting solution and filter residues;

step two: performing rotary evaporation and concentration on the crude polysaccharide extracting solution at 90-120 r/min, performing freeze-drying to obtain crude polysaccharide freeze-dried powder, adding 80-95 vol.% of ethanol, standing for 10-14 h, centrifuging to remove supernatant, adding 5 times of purified water for redissolution, removing protein by adopting a Sevage reagent, adding activated carbon according to the amount of 2% of the solution by mass for decoloring for 30-50 min, performing membrane dialysis with 300Da to remove monosaccharide, and performing freeze-drying to obtain black jerusalem artichoke oligosaccharide powder.

Further, in the first step, the microjet pressure is 120-150 Mpa, the extraction temperature is 70-80 ℃, the extraction time is 60-120 min, and the extraction times are 1-2.

Further, in the second step, the mass-to-volume ratio of the crude polysaccharide freeze-dried powder to the ethanol is 1g:5 to 6mL.

Further, in the second step, the volume ratio of chloroform to n-butanol in the Sevage reagent is 5:1.

further, in the second step, the oligosaccharide content is 80% -88%, the components include kestose, nystose and nystose, and the proportion of the components is 1: 3.0-3.5: 2.0 to 2.5:1.0 to 1.5.

Compared with the prior art, the invention has the beneficial effects that: animal experiments prove that the jerusalem artichoke oligosaccharide prepared by the invention can adjust the blood fat level of a hyperlipemia model mouse, and increase the abundance of Lachnospiraceae-NKA 136_ group flora, such as lactobacillus, bifidobacterium, lachnospiraceae, coprinus and uncultured Lachnospiraceae. The lactobacillus and the bifidobacterium have the effects of inhibiting the propagation of putrefying bacteria, eliminating toxin generated by the putrefying bacteria, clearing away intestinal garbage, inhibiting the absorption of cholesterol, reducing blood fat and lowering blood pressure.

Drawings

FIG. 1 is a diagram of morphological observation (HE staining) of liver tissues of mice in a blank control group;

FIG. 2 is a diagram of morphological observation (HE staining) of liver tissues of a mouse in a model control group;

FIG. 3 is a diagram showing morphological observation (HE staining) of liver tissues of mice in a positive control group;

FIG. 4 is the liver histomorphism observation (HE staining) chart of the mice in the low dosage group of the oligosaccharide of black jerusalem artichoke;

FIG. 5 is the liver tissue morphology observation (HE staining) of mice in the medium dosage group of Helianthus tuberosus oligosaccharides;

FIG. 6 is a diagram of the morphology observation (HE staining) of the liver tissues of mice in the high-dose group of Helianthus tuberosus oligosaccharides;

FIG. 7 is a diagram showing the morphological observation (oil red O staining) of the liver tissue of the blank control group mouse;

FIG. 8 is a diagram showing the morphological observation (oil red O staining) of the liver tissue of the mouse in the model control group;

FIG. 9 is a diagram showing a morphological observation (oil red O staining) of liver tissues of mice in a positive control group;

FIG. 10 is the liver tissue morphology observation (oil red O staining) of mice in the low-dose group of Helianthus tuberosus oligosaccharides;

FIG. 11 is a diagram of the morphology observation (oil red O staining) of the liver tissues of mice in the medium dosage group of Helianthus tuberosus oligosaccharides;

FIG. 12 is a diagram of the observation of the liver tissue morphology (oil red O staining) of mice in the high-dose group of Helianthus tuberosus oligosaccharides;

FIG. 13 is a graph of the number of OTUs from different groups of mice;

FIG. 14 is a horizontal bar graph of gut flora of different groups;

FIG. 15 is a horizontal bar graph of different groups of intestinal flora.

Detailed Description

The technical solutions of the present invention are further described below with reference to the drawings and the embodiments, but the present invention is not limited thereto, and modifications or equivalent substitutions may be made to the technical solutions of the present invention without departing from the spirit of the technical solutions of the present invention.

Example 1:

1. and (3) preparing the jerusalem artichoke oligosaccharide. Weighing a proper amount of black jerusalem artichoke, cutting into slices of 1.0-2.0 cm, adding purified water according to a material-water ratio (g/mL) of 1. The microjet pressure is 120-150 Mpa, the extraction temperature is 70-80 ℃, the extraction time is 60-120 min, and the extraction times are 1-2.

2. Carrying out rotary evaporation and concentration on the black jerusalem artichoke polysaccharide solution at the speed of 90-120 r/min, freeze-drying to obtain freeze-dried powder, adding 80-95% ethanol according to the amount 5 times of the mass-volume ratio (g: mL) of the freeze-dried powder, standing for 10-14 h, and centrifuging to remove supernatant. Adding purified water of which the volume is 5 times that of the solution for redissolution, adopting Sevage reagent with the volume ratio of chloroform to n-butyl alcohol being 5 to remove protein, and adding activated carbon according to the amount of 2 percent of the solution mass for decolorization for 30-50 min. Dialyzing with a 300Da membrane to remove monosaccharide, and freeze-drying to obtain the jerusalem artichoke oligosaccharide powder, wherein the oligosaccharide content is 80-88%. The candy comprises kestose, nystose and nystose, and the proportion of the components is 1: 3.0-3.5: 2.0 to 2.5:1.0 to 1.5.

Research on effect of Helianthus tuberosus oligosaccharides prepared in example 1 on reducing blood lipid of hyperlipidemic mice

Materials and instruments

1. Experimental animal SPF-level male health C57BL/6 mice vinpocetine experimental animal technology ltd, experimental animal production license number SCXK- (gis) 2020-0002; the indoor environment conditions of the feeding are 20-22 ℃ and 28-50% of relative humidity, and the experiment accords with the national regulations on the management of experimental animals.

2. The feed and reagent mouse basic feed and the high-fat feed are prepared by Beijing Australian cooperative feed Co., ltd, the formula of the high-fat feed is executed according to the requirement of the national food and drug administration No. 2012 document No. 107, and the formula of the high-fat feed is prepared by adding 20.0% of cane sugar, 15% of lard, 1.2% of cholesterol and 0.2% of sodium cholate into the basic feed; atorvastatin, beijing Jialin, pharmaceutical Co., ltd; the blood lipid level detection kit and the antioxidant index detection kit Nanjing are built into a biotechnology limited company;

3. instrument 3BT9610 PCR Instrument, select Cycler II, USA; CFX96 Real time PCR instrument, bio-Rad, inc; 5200 automated chemiluminescent image analysis System, shanghai Tanon technologies, inc.; multiskan FC microplate reader Thermo corporation; TANKPE0600 pure Water System MILLIPORE, USA; VFD-4500 model lyophilizer, beijing Bo Yi kang laboratory instruments, inc.; XMTD-204 digital display type electric heating constant temperature water bath Tianjin Euro instrument Meter Co., ltd; sorvall ST8/8R cryocentrifuge, thermo corporation, USA.

(II) an experimental method.

1. Establishment of hyperlipemia model

Mice were acclimatized for 7 days, during which period ordinary feed was given, and free access to water. The 8d group was randomly divided into two groups, a high fat diet group (42) and a blank group (10). And 35d, taking blood to detect the content of TC, TG, HDL-C and LDL-C in the serum, and judging that the model is successfully established.

2. Grouping animals

The mice were divided into 6 groups, the original blank group was changed to a blank control group (CK), the original high fat diet group was randomly divided into 5 groups, which were respectively a model control group (MC), a positive control group (AC), a low dosage group (OSL) of oligosaccharide from black jerusalem artichoke, a medium dosage group (OSM) and a high dosage group (OSH), and 8 mice were each group. The gavage amount of each mouse is 0.2mL per day, and the gavage is performed for 1 time per day and for 46 days. Group CK and MC were given physiological saline; atorvastatin 1.0 mg/(kg bw) was administered to group AC; the low, medium and high dosage components of the oligosaccharide of the black jerusalem artichoke are respectively given with 0.25 g/(kg bw), 1.25 g/(kg bw) and 2.50 g/(kg bw) of oligosaccharide solution of the black jerusalem artichoke. During the period, CK group was fed normal diet, the rest groups were fed high fat diet, and each group of mice had free access to drinking water. The experiment period is 82d, and the quality of the mouse body is periodically detected.

3. Sample collection

After the test object drying is finished, the mice are fasted for 12h, the weight is weighed, blood is taken out in an enzyme-free tube, the tube is stood for 2h at room temperature and centrifuged for 10min at 5000r/min, and serum is taken out and frozen at the temperature of minus 80 ℃. Dissecting and taking liver and cecum, quickly freezing with liquid nitrogen, and freezing at-80 deg.C for later use.

4. Detection of serum markers

Total Cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), superoxide dismutase (SOD), glutathione (GSH-Px) and Malondialdehyde (MDA) in the blood serum of the mice are detected according to the requirements of different kits, and each sample is provided with 3 times of technical repetition.

5. Hematoxylin, eosin (HE) and oil red O staining for observing morphological change of liver tissue of hyperlipidemic mouse

Take 1cm ³ Fixing the liver tissue with 10% formaldehyde, dehydrating with alcohol, embedding with paraffin, making pathological section, and performing HE or oil red O staining. The specific steps are operated according to the kit instructions.

6. Mouse intestinal flora diversity and short-chain fatty acid content detection

The cecum of the mouse is taken as a sample, and the flora diversity and the content of short-chain fatty acid in the content of the cecum are detected. This step is accomplished by Biotech, inc., baimaike, beijing.

(III) data analysis

DPS v7.05 software is adopted for data statistical analysis, single-factor analysis of variance (One-way ANOVA) is adopted for significance analysis of differences among groups, turkey post-test is carried out simultaneously, data results are expressed by 'mean value plus or minus standard deviation', and the difference when P is less than 0.05 is considered to have statistical significance.

(IV) results of the experiment

1. Influence of Jerusalem artichoke oligosaccharide on quality change of mice

TABLE 1 body Mass Change during the test in groups of mice

Time

CK

MC

AC

OSL

OSM

OSH

7d

21.00±1.04

21.06±1.31

20.56±1.34

20.88±0.65

20.88±1.07

20.06±0.54

21d

21.87±1.05

23.24±0.92

22.91±0.85

23.41±0.88

23.55±1.12

23.01±0.72

35d

22.53±1.11

25.37±1.23 ^#

25.74±0.79 ^#

25.74±1.25 ^#

25.77±0.79 ^#

25.87±0.49 ^#

49d

23.20±0.89

27.35±0.54 ^#

27.67±0.53 ^#

27.85±0.58 ^#

28.23±0.83 ^#

28.03±0.86 ^#

63d

23.83±0.67

29.63±0.78 ^##

30.23±1.12 ^##

30.33±0.73 ^##

30.41±0.66 ^##

30.22±0.77 ^##

82d

25.02±1.26

33.34±0.82 ^##

35.12±0.93 ^##

34.14±1.15 ^##

34.53±1.06 ^##

33.08±0.57 ^##

Note: # (P is less than 0.01), and the difference is very obvious compared with CK; the # P is less than 0.05, and the difference is obvious compared with CK;

as can be seen from Table 1, the body mass of each group of mice shows a gradual increase trend in the whole experimental process, and when the model building is successful for 35d, the body mass of the mice in the high-fat feed group is obviously higher than that of the mice in the CK group (P is less than 0.05). At the end of feeding, the body mass of each group of mice is still obviously higher than that of CK group (P is less than 0.01), and compared with MC group, the difference of the body mass of each treatment group of mice is not obvious (P is more than 0.05). The result shows that the mice body mass can be increased by feeding high-fat feed, and the stomach-perfused black jerusalem artichoke oligosaccharide can not obviously inhibit the body mass increase, but can not further fatten the mice.

2. Influence of jerusalem artichoke oligosaccharide intervention on blood lipid level of hyperlipidemic mouse

TABLE 2 blood lipid levels in mice of different groups

Group of	TG(mmol/L)	TC(mmol/L)	HDL-C(mmol/L)	LDL-C(mmol/L)
					CK	0.59±0.09	3.41±0.36	8.01±0.70	1.26±0.31
MC	1.32±0.35 ^##	9.82±1.56 ^##	5.28±0.95 ^##	6.26±1.52 ^##
					AC	1.09±0.27	3.83±0.53 ^**	8.66±1.06 ^**	1.59±0.63 ^**
OSL	0.82±0.30	9.32±1.37 ^##	9.71±2.60 ^#**	5.61±0.35 ^##
					OSM	0.71±0.26 ^*	8.32±0.79 ^##	9.64±1.04 ^#**	3.71±0.41 ^#*
OSH	0.69±0.38 ^*	7.50±1.01 ^##	10.30±2.13 ^##**	3.57±0.87 ^#*

Note: # P is less than 0.01, and has obvious difference compared with CK, and # P is less than 0.05, and has obvious difference compared with CK;

* P < 0.01, which is significantly different from MC, and P < 0.01, which is significantly different from MC.

As can be seen from Table 2, the serum contents of TG, TC and LDL-C in the mice in the MC group are obviously higher than those in the CK group (P < 0.01), and the content of HDL-C is obviously lower than those in the CK group (P < 0.01), which indicates that the mixed type hyperlipemia model is successfully established. The level of HDL-C in serum of mice in each group of Helianthus tuberosus oligosaccharides is obviously higher than that in MC group (P is less than 0.01 or P is less than 0.05), the levels of TG, TC and LDL-C are all lower than that in MC group, the reduction levels of TG and LDL-C in mice in medium and high dose group have statistical significance (P is less than 0.05), and the level of TG is even lower than that in AC group. TG levels in mice in the medium-high dose group are not obviously different from CK groups, and TC, LDL-C and HDL-C levels are obviously increased (P < 0.01 or P < 0.05).

The intervention of a proper amount of jerusalem artichoke oligosaccharide can obviously reduce TG and LDL-C values of hyperlipidemic mice, the TG value can approach to a normal level, but the LDL-C is still higher than the normal level; the TC value can be reduced but still above the normal level.

3. Influence of Helianthus tuberosus oligosaccharide intervention on oxidative stress level of hyperlipidemic mice

TABLE 3 antioxidant index levels in different groups of mice

Group of	SOD(U/mgprot)	MDA(nmol/mgprot)	GSH-Px(U/mgprot)
				CK	289.63±51.17	2.23±0.57	61.49±8.65
MC	226.95±23.63 ^##	8.57±3.12 ^##	35.54±5.84 ^##
				AC	302.57±53.45 ^**	2.88±2.08 ^**	57.05±12.32 ^**
OSL	230.56±65.76 ^##	8.02±2.38 ^##	37.40±6.73 ^##
				OSM	260.09±21.00 ^**	7.58±5.79 ^##**	52.08±5.86 ^#**
OSH	279.82±32.53 ^**	5.01±2.26 ^##**	58.23±8.56 ^**

Note: # P is less than 0.01, has extremely obvious difference compared with CK, and # P is less than 0.05, has obvious difference compared with CK;

* P < 0.01 with very significant differences compared to MC and P < 0.01 with significant differences compared to MC.

As can be seen from Table 3, compared with the CK group, the serum levels of Malondialdehyde (MDA) in mice in the MC group are obviously increased (P is less than 0.01), the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) are obviously reduced (P is less than 0.01), the MDA levels in the Jerusalem artichoke oligosaccharide and in the high-dose group are obviously increased (P is less than 0.01), and the enzyme activities of SOD are not obviously different (P is more than 0.05); the GSH-Px enzyme activity of the OSM group is obviously lower than that of the CK group (P is less than 0.05), and the GSH-Px enzyme activity of the OSH group is not obviously different from that of the CK group (P is more than 0.05). Compared with the MC group, the MDA level of the mice in the medium and high dose of the Jerusalem artichoke oligosaccharide group is obviously reduced (P is less than 0.01), and the levels of SOD and GSH-Px are obviously increased (P is less than 0.01). Therefore, a proper amount of jerusalem artichoke oligosaccharide has a certain inhibiting effect on oxidative stress caused by high-fat diet, and can improve the activities of SOD and GSH-Px enzyme in the serum of a hyperlipidemic mouse to enable the activity to be recovered to be close to the CK group level; can reduce the content of MDA, but has certain difference compared with CK group.

4. Influence of jerusalem artichoke oligosaccharide intervention on liver histopathological changes of hyperlipidemic mice

As can be seen from FIGS. 1 to 6, the liver cells of the CK group mice are arranged regularly and regularly, the structure is normal, and the cytoplasm has no fat drop; the liver cells of the mice in the MC group are abnormal, and a large number of lipid droplets appear in cytoplasm; the medium-high dose of the Jerusalem artichoke oligosaccharide group has a good effect of protecting liver cells, the number of fat drops is obviously reduced, the liver cells are arranged relatively neatly, particularly, the high dose of the Jerusalem artichoke Oligosaccharide (OSH) group has the best liver tissue state, no obvious fatty degeneration liver cells are seen, and the effect is close to that of a positive control atorvastatin group.

After the liver tissue is stained by oil red O, lipid droplets in the tissue cells are red, and the nucleus is blue. As can be seen from FIGS. 7-12, liver tissue from CK mice was stained a uniform blue color, indicating the absence of fat droplets in the cytoplasm; the liver tissue section of the MC group mouse is obviously red, which indicates that a large amount of lipid drops exist in cytoplasm; the red color in the slices of each group of the jerusalem artichoke oligosaccharide gradually decreases and the blue color gradually increases along with the increase of the administration concentration, and the group with high dosage has the best performance and is close to the effect of a positive control group.

5. Diversity analysis of intestinal flora in mice

As shown in FIG. 13, the total number of OTUs in each group is 464, the difference between each group and the total number of OTUs is very significant (P < 0.05), and the difference between each group is not significant (P > 0.05), but still shows a certain regularity. The average number of OTU in MC group is 445, and the number of OTU in 451,OSL, OSM and OSH group is 447, 448 and 450 respectively, which are less than that in CK group. The intestinal flora diversity of the hyperlipidemic mice is reduced, and after the oligosaccharide dry prognosis of the jerusalem artichoke, the flora diversity is not obviously improved but is better than that of a positive control group.

Studies have shown that obese people have 20% more firmicutes in their bodies than lean people, and 90% less bacteroides in their bodies than lean people. From fig. 14, at the phylum level, all the groups except the group AC showed a law of increasing the firmicutes abundance and decreasing the bacteroidetes abundance compared to the group CK, and in terms of the firmicutes/bacteroidetes ratio, the group CK was the lowest, 1.01, the group MC was 1.17, slightly higher than the group CK, and the other groups were slightly higher than the group MC, but the difference was not significant, which coincided with the increase of body weight slightly after the jerusalem artichoke oligosaccharide intervention.

The abundance of actinomycetomycota of the high-fat diet mice is higher than that of the CK group, the AC group is obviously higher than that of the CK group, and the difference among other groups is not obvious. The actinomycetemcomia includes bifidobacterium and beneficial bacteria capable of producing streptomycin, terramycin, tetracycline, gentamicin, etc.

The abundance of the bacterial groups of the deironics of each group of mice is higher than that of the CK group, the MC group is 3.45 times of that of the CK group, the oligosaccharide of the Helianthus tuberosus is 1.70, 1.86 and 1.32 times of that of the MC group, and the AC group is 0.50 times of that of the MC group. Li Yanmei et al report that the abundance of deferribacterium in mice of an intestinal cancer model group is reduced from 1.9% to 0.9% and the abundance of codonopsis pilosula is restored to 1.2% after treatment compared with normal mice, and the trend indicates that the black jerusalem artichoke can reduce the occurrence of intestinal cancer by improving the abundance of the deferribacterium, and the effect of the black jerusalem artichoke oligosaccharide is better than that of the positive control drug atorvastatin.

The abundance of the microbial flora of verrucomicrobia of the high-fat diet mice is higher than that of the CK group, and the abundance of the microbial flora is reduced to a certain extent and gradually approaches to the normal level after the prognosis of the medium-high dose Jerusalem artichoke oligosaccharide.

The proteobacteria contains more pathogenic bacteria, and compared with the CK group, the proteobacteria content of each treatment group is reduced except for the AC group and the OSL group.

As can be seen from fig. 15, at the genus level, high-fat diet reduced the abundance of uncultured Murbiaculaceae bacteria in mice, increased the abundance of uncultured lacosacceae bacteria, and the oligosaccharide stem prognosis of black jerusalem artichoke was not changed, but was not significantly different from the MC group; the high-fat diet can increase the abundance of lactic acid bacteria, bifidobacteria, undeplemented bacteria of the rhinaceae of the rhinacobacteriaceae of actinomycetemcomita and helicobacter in mice, the effect is enhanced by the intervention of the jerusalem artichoke oligosaccharide, and the abundance of the lactic acid bacteria in the OSH group is obviously higher than that in the MC group and the CK group (P is less than 0.05). High-fat diet can reduce the abundance of Lachnospiraceae NKA136 group flora in mice, and medium-dose Jerusalem artichoke oligosaccharide intervention can increase the abundance slightly higher than those of MC group and CK group. As is well known, lactic acid bacteria, bifidobacteria and lachnospiraceae bacteria are intestinal probiotics, and the lactic acid bacteria and the bifidobacteria can beneficially change the composition of intestinal flora, improve the function of the gastrointestinal tract of a human body, restore the flora balance in the intestinal tract of the human body, form an antibacterial biological barrier and maintain the health of the human body. Can inhibit the propagation of putrefying bacteria, eliminate toxin produced by putrefying bacteria, and remove intestinal rubbish, and has the effects of inhibiting cholesterol absorption, reducing blood lipid and lowering blood pressure; the lachnospiraceae bacteria can participate in the metabolism of human short-chain fatty acids, the short-chain fatty acids are main sources of intestinal epithelial cell energy, and the decrease of the abundance of the lachnospiraceae bacteria can cause the permeability change of the intestinal epithelial cells to further influence various physiological functions of organisms.

High-fat diet can increase the abundance of bacillus in intestinal tract of mice, the abundance of coprobacterium in each group of the oligosaccharide of the black jerusalem artichoke is higher than that of the MC group and the CK group, and the abundance of the oligosaccharide of the black jerusalem artichoke is 82 times and 273 times of that of the MC group and the CK group respectively according to the highest abundance of the OSL group. Zhouzhi research shows that the coprobacter elongatus has the efficacy of preventing and treating diabetes, and provides a composition containing the coprobacter elongatus.

Claims

1. A preparation method of jerusalem artichoke oligosaccharides with blood lipid reducing effect and intestinal flora structure improving effect is characterized by comprising the following steps: the method comprises the following steps:

the method comprises the following steps: preparing the oligosaccharide of black jerusalem artichoke: slicing, dicing or dicing the black jerusalem artichoke according to the proportion of 1g: adding purified water in a proportion of 10-20 mL, homogenizing and homogenizing by adopting a dynamic ultrahigh pressure microjet technology for auxiliary extraction, and filtering by adopting six layers of gauze to obtain a crude polysaccharide extracting solution and filter residues;

step two: performing rotary evaporation concentration on the crude polysaccharide extracting solution at the speed of 90-120 r/min, performing freeze-drying to obtain crude polysaccharide freeze-dried powder, adding 80-95 vol.% of ethanol, standing for 10-14 h, centrifuging to remove supernatant, adding 5 times of purified water for redissolution, removing protein by adopting a Sevage reagent, adding activated carbon according to the amount of 2% of the solution by mass for decolorization for 30-50 min, performing dialysis by using a 300Da membrane to remove monosaccharide, and performing freeze-drying to obtain black jerusalem artichoke oligosaccharide powder.

2. The method for preparing Jerusalem artichoke oligosaccharides with effects of reducing blood lipid and improving intestinal flora according to claim 1, which is characterized by comprising the following steps: in the first step, the microjet pressure is 120-150 Mpa, the extraction temperature is 70-80 ℃, the extraction time is 60-120 min, and the extraction times are 1-2.

3. The method for preparing jerusalem artichoke oligosaccharides with hypolipidemic effect and improved intestinal flora structure according to claim 1, wherein the method comprises the following steps: in the second step, the mass-to-volume ratio of the crude polysaccharide freeze-dried powder to the ethanol is 1g:5 to 6mL.

4. The method for preparing jerusalem artichoke oligosaccharides with hypolipidemic effect and improved intestinal flora structure according to claim 1, wherein the method comprises the following steps: in the second step, the volume ratio of chloroform to n-butanol in the Sevage reagent is 5:1.

5. the method for preparing jerusalem artichoke oligosaccharides with hypolipidemic effect and improved intestinal flora structure according to claim 1, wherein the method comprises the following steps: in the second step, the oligosaccharide content is 80% -88%, the components comprise kestose, nystose and nystose, and the proportion of the components is 1:3.0 to 3.5:2.0 to 2.5:1.0 to 1.5.