CN110959863A

CN110959863A - Stevia rebaudiana leaf mixed nutrient syrup and preparation method and application thereof

Info

Publication number: CN110959863A
Application number: CN201911288732.4A
Authority: CN
Inventors: 徐明德
Original assignee: Individual
Current assignee: Individual
Priority date: 2019-12-13
Filing date: 2019-12-13
Publication date: 2020-04-07

Abstract

The invention provides a preparation method of stevia rebaudiana leaf mixed nutrient syrup, which at least comprises stevia rebaudiana leaf extract, nutrient extract, sweetener, brewed vinegar, fruit juice and a flavoring agent, and the preparation method comprises the following steps: step (1), mixing stevia rebaudiana leaf extract, nutrient materials, sweetener, brewed vinegar, fruit juice and a regulator, adding water, and stirring uniformly to fully dissolve the mixture; standing for 24 hours, filtering, adding water to a specified amount, decocting with slow fire while stirring, decocting until the relative density is 3-5, and stopping heating; and (3) standing for 24 hours, filling and sterilizing to obtain the product. The active extract has obvious liver protection effect on chemical liver injury and alcoholic liver injury. The stevia rebaudiana leaf mixed nutrient syrup reduces the sugar content of the syrup, and the stevioside component can obviously improve the sweetness of the syrup without increasing the blood sugar content of eaters, so that the stevia rebaudiana leaf mixed nutrient syrup can be eaten by diabetics, essential hypertension patients, hyperglycemia patients and hyperlipidemia patients. The invention solves the problem that the common syrup has no health care value.

Description

Stevia rebaudiana leaf mixed nutrient syrup and preparation method and application thereof

Technical Field

The invention relates to the technical field of food processing, and particularly relates to stevia rebaudiana leaf mixed nutrient syrup and a preparation method and application thereof.

Background

Syrups are viscous, high-concentration sugar-containing solutions prepared by cooking or other techniques. The raw material for making syrup may be sugar water, sugar cane juice, fruit juice or other plant juice, etc. Since the sugar content of the syrup is very high, it can be preserved for a long time in a sealed state without refrigeration. The syrup can be used for preparing beverages or making sweets.

The method is based on syrup, and stevia rebaudiana leaves are selected as raw materials, so that the obtained stevia rebaudiana leaf mixed nutrient syrup has a good protection effect on liver injury after being eaten, a plurality of lung moistening syrups, cough relieving syrups, sore throat relieving syrups and the like are prepared in the prior art, but the preparation of the stevia rebaudiana leaf mixed nutrient syrup for preventing and treating liver injury is not reported at present. Meanwhile, the low-fat low-calorie syrup has the characteristics of low fat and low calorie, and is prepared by mixing nutrient materials such as vitamins, amino acids, animal proteins, plant proteins or traditional Chinese medicines and the like to supplement nutrient substances required by a human body, so that the healthy and low-calorie syrup is complementary to the deficiency of daily diet and is beneficial to the health of people after being eaten for a long time.

Stevia belongs to Compositae, also called stevia rebaudiana and sweet grass, is a perennial wild herb of Compositae. Stevia leaves contain a large amount of stevioside, the content of the stevioside accounts for about 10 percent, the stevioside is a natural sweetener, and the stevioside is internationally endowed with the names of 'plant sugar king', 'best natural sweetener' and the like. It can be used to replace various chemically synthesized sweeteners such as sugar and saccharin, and can be widely used in food, beverage, medicine, daily chemical industry, wine brewing, and cosmetics. The food beverage prepared from the edible oil can not make people fat after long-term eating, and is especially suitable for patients with obesity, diabetes, hypertension, arteriosclerosis, dental caries, etc. In recent years, the research on stevia rebaudiana leaves at home and abroad mainly focuses on the extraction and application of stevioside, and the research on the capability of the stevia rebaudiana leaves in protecting chemically damaged liver and preventing and treating liver damage is not reported yet.

Disclosure of Invention

The invention provides a stevia rebaudiana leaf mixed nutrient syrup and a preparation method thereof, and aims to provide the stevia rebaudiana leaf mixed nutrient syrup, wherein an active extract of the stevia rebaudiana leaf mixed nutrient syrup is prepared by adopting water soaking and supercritical fluid extraction, and the extracted stevia rebaudiana leaf extract is mixed with nutrient materials such as traditional Chinese medicine extract, vitamins, animal protein, plant protein, amino acid and the like, so that the stevia rebaudiana leaf mixed nutrient syrup can be used for preparing health-care products or food for preventing and treating various types of liver protection, can obviously improve the sweetness of the syrup without increasing the blood sugar content of eaters, and can be eaten by diabetics, essential hypertension patients, hyperglycemia patients and hyperlipidemia patients. The invention solves the problem that the common syrup has no health care value.

The invention provides a preparation method of stevia rebaudiana leaf mixed nutrient syrup, which at least comprises stevia rebaudiana leaf extract, nutrient extract, sweetener, brewed vinegar, fruit juice and a flavoring agent, and the preparation method comprises the following steps:

step (1), mixing stevia rebaudiana leaf extract, nutrient materials, sweetener, brewed vinegar, fruit juice and a regulator, adding water, and stirring uniformly to fully dissolve the mixture;

standing for 24 hours, filtering, adding water to a specified amount, decocting with slow fire while stirring, decocting until the relative density is 3-5, and stopping heating;

and (3) standing for 24 hours, filling and sterilizing to obtain the product.

Preferably, after the step (3), the method further comprises: and (4a) mixing the stevia rebaudiana leaf mixed nutrient syrup into one of pure water, milk, fruit juice, almond juice, soybean milk, cream, light cream, sour cream, cheese, yoghourt, snow lamb, vinegar, fruit vinegar, black tea, milk tea, coffee, cocoa powder beverage, traditional Chinese medicine liquid, western medicine liquid, natural fruit juice, artificial fruit juice, assam black tea, Ceylon black tea, Burju black tea, Litton black tea, Pu' er tea, black tea powder and coarse tea and uniformly stirring.

Preferably, after the step (3), the method further comprises: and (4b) mixing the syrup with one of syrup jelly, milk jelly, almond jelly, soybean milk jelly, black tea jelly, milk tea jelly, coffee jelly, cocoa powder jelly and Chinese medicinal liquid jelly, and uniformly mixing.

Preferably, after the step (3), the method further comprises: and (4c) mixing the syrup with one of cake, biscuit, peanut butter, jam and liquor and uniformly mixing.

Preferably, the sterilization method is pasteurization, specifically heating the syrup to 62-65 deg.C, and maintaining for 30 min.

Preferably, the sweetening agent is selected from one or more of inulin, mogroside, erythritol, malt syrup, yacon syrup and sorbitol, the brewing vinegar is selected from one or more of glutinous rice vinegar, black vinegar, red vinegar, persimmon vinegar, apple vinegar, red date vinegar, rose vinegar, peach blossom vinegar and roselle vinegar, the fruit juice is selected from one or more of lemon juice, orange juice, apple juice, watermelon juice, grape juice, grapefruit juice, passion fruit juice and blueberry juice, and the blending agent is selected from one or more of Arabic gum, tamarind polysaccharide gum, sesbania gum, agar, xanthan gum, β -cyclodextrin and propylene glycol alginate.

As a further improvement of the invention, the stevia rebaudiana leaf extract is prepared by the following method:

s1, cleaning stevia rebaudiana leaves, cutting into pieces, adding the stevia rebaudiana leaves into pure water for soaking, performing ultrasonic-assisted extraction for 30min, filtering, reserving filter residues, and freeze-drying filtrate to obtain water extract freeze-dried powder;

s2, filling the filter residue obtained in the step S1 into an extraction kettle, and opening CO₂The steel cylinder is used for carrying out supercritical fluid extraction, after the extraction is finished, a valve of the extraction kettle is opened, the extract is collected and dried at low temperature, and the supercritical fluid extract is obtained;

the extraction conditions were:

extraction pressure: 21-24 MPa; the extraction temperature is as follows: 45-50 ℃; CO 2₂Flow rate: 6-9L/h; extraction time: 2 h; entrainer: water and ethanol in a volume ratio of 3: 1;

s3, uniformly mixing the water extraction freeze-dried powder obtained in the step S1 and the supercritical fluid extract obtained in the step S2 to obtain the stevia rebaudiana leaf extract.

As a further improvement of the invention, the ultrasonic power is 1500-2000W, the freeze drying condition is that the mixture is frozen at-15 ℃ for 30min and then cooled to-35 ℃ for 10h, and the low-temperature drying is drying at-5 ℃ for 1 h.

As a further improvement of the invention, the mass ratio of the stevia rebaudiana leaf extract to the nutrient material to the sweetener to the brewed vinegar to the fruit juice to the blending agent is 100 (5-10): 2: (3-7): (10-20): (2-5).

As a further improvement of the invention, the nutrient material is selected from one or a mixture of several of traditional Chinese medicine extract, vitamins, animal protein, plant protein and amino acid.

Preferably, the vitamin is one or a mixture of several selected from vitamin A, vitamin C, vitamin E, B family vitamin, vitamin D and vitamin K.

Preferably, the amino acid is selected from one or more of glycine, alanine, valine, leucine, isoleucine, phenylalanine, proline, tryptophan, tyrosine, serine, cysteine, methionine, asparagine, glutamine, threonine, aspartic acid, glutamic acid, lysine, arginine and histidine.

Preferably, the animal protein is selected from one or more of whey protein, collagen, beef powder, pork powder and chicken powder. The pork powder and chicken powder are prepared by oven drying fresh pork or chicken until the water content is less than 5%, and pulverizing.

Preferably, the vegetable protein is selected from one or more of bean protein, pea protein, corn protein, hydrolyzed wheat protein and peanut protein.

As a further improvement of the invention, the traditional Chinese medicine extract comprises the following raw materials in parts by weight: 100-200 parts of liquorice, 50-120 parts of sugarcane and 10-20 parts of dendrobium officinale.

As a further improvement of the invention, the traditional Chinese medicine extract is prepared by the following method:

s1, pretreatment: weighing the raw medicinal materials according to a certain proportion, cleaning, drying and crushing to obtain traditional Chinese medicine powder for later use;

s2, extraction: adding the Chinese medicinal powder into pure water, adding cellulase, adjusting pH and temperature, performing enzymolysis for 0.5-1 hr, inactivating enzyme, adding sodium carbonate to adjust pH to neutral, performing ultrasonic-assisted extraction for 1-3 hr, centrifuging, collecting supernatant, concentrating, and freeze drying to obtain Chinese medicinal extract.

As a further improvement of the invention, the pH value is adjusted to 5, the temperature is adjusted to 45-60 ℃, the enzyme deactivation method is heating at 100 ℃ for 10min, the ultrasonic power is 1200-1500W, the centrifugation condition is 4 ℃, 10000-12000r/min centrifugation is 5min, the freeze drying condition is freezing at-10 ℃ for 15min and freezing at-30 ℃ for 12h, and the mass-volume ratio of the traditional Chinese medicine powder to the pure water is 1: (5-10); the addition amount of the cellulase is 1000-1200U/kg, the concentration method adopts ceramic membrane ultrafiltration, the aperture of the ceramic membrane is 0.05-0.10 mu m, and the operation conditions of the ceramic membrane ultrafiltration are as follows: the transmembrane pressure difference is 0.17-0.30MPa, the filtration temperature is 52-57 ℃, and the membrane surface flow rate is 7-10 m/s.

The invention further protects the application of the stevia rebaudiana leaf mixed nutrient syrup in preparing health-care food or food for preventing and treating liver injury.

In a further improvement of the present invention, the liver injury is chemical liver injury or alcoholic liver injury.

Preferably, the health food is prepared into tablets, injections, capsules, granules, pills, decoctions, syrups and mixtures.

The invention has the following beneficial effects: on the basis of the research on the original chemical components and the traditional efficacy of the stevia rebaudiana leaves, the active extract is prepared by simultaneously adopting water soaking and supercritical fluid extraction, and compared with the prior art, the extracted stevia rebaudiana leaf extract can obviously reduce CCl₄Or ALT and AST activity in serum of a model mouse with alcohol-induced liver injury and liver tissue MDA content, and liver SOD activity are enhanced, which shows that the active extract has obvious liver protection effect on chemical liver injury and alcoholic liver injury. Therefore, the stevia rebaudiana leaf extract is expected to be developed into a new generation of safe and effective medicine, health-care product or food for preventing and treating various types of liver protection.

The stevia rebaudiana leaf mixed nutrient syrup provided by the invention reduces the sugar content of the syrup, the stevioside component can obviously improve the sweetness of the syrup without increasing the blood sugar content of eaters, can be eaten by diabetics, essential hypertension patients, hyperglycemia patients and hyperlipidemia patients, further has the effects of reducing blood pressure, blood sugar, blood fat and the like, and has wide application prospect.

Drawings

In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, and it is obvious that the drawings in the following description are only some embodiments of the present invention, and for those skilled in the art, other drawings can be obtained according to these drawings without creative efforts.

FIG. 1 is a graph showing a comparison of blood glucose levels at different times in each group of mice in test example 2 of the present invention.

Detailed Description

The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.

Test methods in which specific conditions are not specified in the following examples are generally carried out under conventional conditions or under conditions recommended by the manufacturer.

The comminution operation was carried out with a TRM series mechanical impact mill, supplied by Weifang environmental protection science and technology Limited. Drying was carried out by an SZG double-cone rotary vacuum dryer supplied by Henzhou remote drying apparatus Co. Freeze-drying operation was performed with a vacuum freeze-drying agent provided by Shanghai Toulong Fulong science and technology Co., Ltd; the centrifugal operation is carried out by a PZ flat plate direct-connected centrifuge provided by Toxico Longtai chemical mechanical equipment Co., Ltd; the ultrasonic treatment was carried out by an ultrasonic extraction tank provided by Shandong Baihe Biotechnology Co. The supercritical fluid extraction operation is carried out by supercritical fluid extraction equipment provided by Hainan Hua supercritical equipment limited, and the concentration operation is carried out by ceramic membrane equipment provided by Nanjing Pengheng Membrane technology limited.

Stevia rebaudiana leaves are provided by Jining Ooxing stevia products, Inc.

Cellulase, CAS No. 9012-54-8, was supplied by Shanghai Allantin Biotech Co., Ltd.

Sorbitol, CAS number 50-70-4; inulin, CAS No. 9005-80-5; erythritol, CAS number 149-32-6; maltose syrup, CAS number 69-79-4.

Whey protein, CAS No. 84082-51-9; collagen, CAS number 9064-67-9; pea protein, CAS No. 222400-29-5; beef powder, CAS number 68990-09-0; hydrolyzed wheat protein, CAS number 70084-87-6.

Gum arabic, CAS No. 9000-01-5; tamarind polysaccharide gum CAS No. 39386-78-2; sesbania gum, CAS number 12493-90; agar, CAS number 9002-18-0; carrageenan, CAS number 11114-20-8.

Various vitamins and various amino acids are provided by Zhejiang Kangpu Biotechnology GmbH.

Various brewed vinegars and various fruit juices are provided by fuzhou kangda food ltd.

Sugarcane is sweet in taste and neutral in nature, and has the effects of removing heat and quenching thirst, regulating the middle warmer, and widely separating and draining water. Mainly treats fever, dry mouth, cough due to lung dryness, sore throat, dysphoria with smothery sensation in heart and chest, regurgitation, emesis, and edema during pregnancy. Belongs to a material with homology of medicine and food, and is provided by a traditional Chinese medicine drinking tablet factory of Guangdong province medicinal material company.

The liquorice is sweet in taste and neutral in nature, has the functions of tonifying spleen and qi, clearing away heat and toxic materials, eliminating phlegm and stopping cough, relieving spasm and pain and harmonizing the drugs, and enters heart, lung, spleen and stomach channels. Can be used for treating weakness of spleen and stomach, asthenia, palpitation, short breath, cough, excessive phlegm, abdominal and limb spasm, pain, carbuncle, suppurative sore, and relieving drug toxicity and pungency. Belongs to a material with homology of medicine and food, and is provided by a traditional Chinese medicine drinking tablet factory of Guangdong province medicinal material company.

Example 1Preparation of stevia leaf extract

A method for preparing stevia rebaudiana leaf extract comprises the following steps:

s1, pure water extraction: cleaning stevia rebaudiana leaves, cutting into pieces, adding into pure water, soaking, performing ultrasonic-assisted extraction for 30min at an ultrasonic power of 1500W, filtering, retaining filter residues, freeze-drying the filtrate, and cooling to-35 ℃ for 10h after freezing for 30min under the freeze-drying condition of-15 ℃ to obtain water extract freeze-dried powder;

s2, supercritical fluid extraction: filling the filter residue obtained in the step S1 into an extraction kettle, and opening CO₂Performing supercritical fluid extraction in a steel cylinder, opening a valve of an extraction kettle after extraction is finished, collecting extract, and drying at low temperatureDrying at low temperature of-5 deg.C for 1h to obtain supercritical fluid extract;

the extraction conditions were:

extraction pressure: 21 MPa; the extraction temperature is as follows: 45 ℃; CO 2₂Flow rate: 6L/h; extraction time: 2 h; entrainer: water and ethanol in a volume ratio of 3: 1;

s3, uniformly mixing the water extraction freeze-dried powder obtained in the step S1 and the supercritical fluid extract obtained in the step S2 to obtain the stevia rebaudiana leaf extract, wherein the yield is 82%.

Example 2Preparation of stevia leaf extract

s2, supercritical fluid extraction: filling the filter residue obtained in the step S1 into an extraction kettle, and opening CO₂Performing supercritical fluid extraction in a steel cylinder, opening a valve of an extraction kettle after extraction is finished, collecting an extract, and drying at low temperature, wherein the drying at low temperature is-5 ℃ for 1h to obtain a supercritical fluid extract;

the extraction conditions were:

extraction pressure: 24 MPa; the extraction temperature is as follows: 50 ℃; CO 2₂Flow rate: 9L/h; extraction time: 2 h; entrainer: water and ethanol in a volume ratio of 3: 1;

s3, uniformly mixing the water extraction freeze-dried powder obtained in the step S1 and the supercritical fluid extract obtained in the step S2 to obtain the stevia rebaudiana leaf extract, wherein the yield is 85%.

Example 3Preparation of stevia leaf extract

s1, pure water extraction: cleaning stevia rebaudiana leaves, cutting into pieces, adding into pure water, soaking, performing ultrasonic-assisted extraction for 30min at the ultrasonic power of 1700W, filtering, retaining filter residues, freeze-drying the filtrate, and cooling to-35 ℃ for 10h after freezing for 30min under the freeze-drying condition of-15 ℃ to obtain water extract freeze-dried powder;

the extraction conditions were:

extraction pressure: 22 MPa; the extraction temperature is as follows: 47 ℃; CO 2₂Flow rate: 7L/h; extraction time: 2 h; entrainer: water and ethanol in a volume ratio of 3: 1;

s3, uniformly mixing the water extraction freeze-dried powder obtained in the step S1 and the supercritical fluid extract obtained in the step S2 to obtain the stevia rebaudiana leaf extract, wherein the yield is 90%.

Comparative example 1

The experimental process parameters were different compared to example 3.

s1, pure water extraction: cleaning stevia rebaudiana leaves, cutting into pieces, adding into pure water, soaking, performing ultrasonic-assisted extraction for 10min at the ultrasonic power of 500W, filtering, retaining filter residues, freeze-drying the filtrate, and cooling to-35 ℃ for 10h after freezing for 10min under the freeze-drying condition of-15 ℃ to obtain water extract freeze-dried powder;

s2, supercritical fluid extraction: filling the filter residue obtained in the step S1 into an extraction kettle, and opening CO₂Performing supercritical fluid extraction in a steel cylinder, opening a valve of an extraction kettle after extraction is finished, collecting an extract, drying at low temperature, and drying at the low temperature of-5 ℃ for 30min to obtain a supercritical fluid extract;

the extraction conditions were:

extraction pressure: 12 MPa; the extraction temperature is as follows: 27 ℃; CO 2₂Flow rate: 3L/h; extraction time: 1 h; entrainer: water and ethanol in a volume ratio of 1: 1;

s3, uniformly mixing the water extraction freeze-dried powder obtained in the step S1 and the supercritical fluid extract obtained in the step S2 to obtain the stevia rebaudiana leaf extract, wherein the yield is 20%.

Example 4

Step (1), mixing 100g of stevia rebaudiana extract prepared in example 1, 5g of nutrient, 2g of inulin, 3g of red vinegar, 10g of lemon juice and 2g of carrageenan, adding water, and uniformly stirring to fully dissolve the mixture;

the nutrient material comprises vitamin A, whey protein, pea protein, glycine and alanine, and the mass ratio is 1: 50:30:1:2.

Standing for 24 hours, filtering, adding water to a specified amount, decocting with slow fire while stirring, decocting until the relative density is 3, and stopping heating;

and (3) standing for 24 hours, filling, and sterilizing, wherein the sterilization method is pasteurization, and specifically the syrup is heated to 62 ℃ and kept for 30min to obtain the syrup.

Example 5

Step (1), mixing 100g of stevia rebaudiana extract prepared in example 2, 10g of nutrient materials, 2g of mogroside, 7g of apple vinegar, 20g of apple juice and 5g of agar, adding water, and uniformly stirring to fully dissolve the mixture;

the nutritional material comprises vitamin C, vitamin E, collagen, beef powder, hydrolyzed wheat protein, pea protein, tyrosine and serine in a mass ratio of 3:1:10:20:15:22:1: 2.

Standing for 24 hours, filtering, adding water to a specified amount, decocting with slow fire while stirring, decocting until the relative density is 5, and stopping heating;

and (3) standing for 24 hours, filling, and sterilizing, wherein the sterilization method is pasteurization, and specifically the syrup is heated to 65 ℃ and kept for 30min to obtain the syrup.

And (4) mixing the stevia rebaudiana leaf mixed nutrient syrup into milk and uniformly stirring.

Example 6

Step (1), mixing 100g of stevia leaf extract prepared in example 3, 6g of nutrient material, 2g of erythritol, 4g of persimmon vinegar, 12g of grape juice and 3g of sesbania gum, adding water, and stirring uniformly to fully dissolve the components;

the nutrient material comprises traditional Chinese medicine extract, vitamin C, lactalbumin, tryptophan and tyrosine, and the mass ratio is 10:2:12:1: 2.

The traditional Chinese medicine extract comprises the following raw materials in parts by weight: 100 parts of liquorice, 50 parts of sugarcane and 10 parts of dendrobium officinale.

The traditional Chinese medicine extract is prepared by the following method:

s2, extraction: adding 50mL of traditional Chinese medicine powder 10g into pure water, adding cellulase with the addition of 1000U/kg, adjusting the pH value to 5 by using 1mol/L hydrochloric acid, adjusting the temperature to 45 ℃, performing enzymolysis for 0.5h, performing enzyme deactivation, wherein the enzyme deactivation method comprises the steps of heating at 100 ℃ for 10min, adding sodium carbonate to adjust the pH value to be neutral, performing ultrasonic assisted extraction at 1200W for 1h, centrifuging under the centrifugal condition of 4 ℃ and 10000r/min for 5min, collecting supernatant and concentrating, wherein the concentration method comprises the steps of performing ultrafiltration by adopting a ceramic membrane, the aperture of the ceramic membrane is 0.10 mu m, and the operation condition of the ceramic membrane ultrafiltration is as follows: transmembrane pressure difference of 0.30MPa, filtration temperature of 57 deg.C, membrane surface flow rate of 10m/s, and freeze drying at-10 deg.C for 15min and-30 deg.C for 12 hr to obtain Chinese medicinal extract.

Standing for 24 hours, filtering, adding water to a specified amount, decocting with slow fire while stirring, decocting until the relative density is 3.5, and stopping heating;

and (3) standing for 24 hours, filling, and sterilizing, wherein the sterilization method is pasteurization, and specifically the syrup is heated to 63 ℃ and kept for 30min to obtain the syrup.

And (4) mixing the syrup with the black tea jelly uniformly.

Example 7

Step (1), mixing 100g of stevia rebaudiana extract prepared in example 3, 9g of nutrient material, 2g of malt syrup, 6g of black vinegar, 18g of passion fruit juice and 4g of tamarind polysaccharide gum, adding water, and uniformly stirring to fully dissolve the components;

the nutrient material comprises traditional Chinese medicine extract, B vitamins, whey protein, pea protein and proline in a mass ratio of 12:1:15:5:1:2, wherein the B vitamins comprise vitamin B1, vitamin B12 and folic acid in a mass ratio of 5:1: 1.

The traditional Chinese medicine extract comprises the following raw materials in parts by weight: 200 parts of liquorice, 120 parts of sugarcane and 20 parts of dendrobium officinale.

The traditional Chinese medicine extract is prepared by the following method:

s2, extraction: adding 10g of traditional Chinese medicine powder into 100mL of pure water, adding cellulase with the addition of 1200U/kg, adjusting the pH value to 5 by using 1mol/L hydrochloric acid, adjusting the temperature to 60 ℃, performing enzymolysis for 1h, and then performing enzyme deactivation, wherein the enzyme deactivation method comprises the steps of heating at 100 ℃ for 10min, adding sodium carbonate to adjust the pH value to be neutral, performing 1500W ultrasonic-assisted extraction for 3h, centrifuging at the centrifugal condition of 4 ℃ and 12000r/min for 5min, collecting supernatant and concentrating, wherein the concentration method comprises the steps of performing ultrafiltration by adopting a ceramic membrane, the aperture of the ceramic membrane is 0.05 mu m, and the operation condition of the ceramic membrane ultrafiltration is as follows: transmembrane pressure difference is 0.17MPa, filtering temperature is 52 ℃, membrane surface flow rate is 7m/sm/s, and freeze drying is carried out under the conditions of freezing at-10 ℃ for 15min and freezing at-30 ℃ for 12h to obtain the traditional Chinese medicine extract.

Standing for 24 hours, filtering, adding water to a specified amount, decocting with slow fire while stirring, decocting until the relative density is 4.5, and stopping heating;

and (3) standing for 24 hours, filling, and sterilizing, wherein the sterilization method is pasteurization, and specifically the syrup is heated to 64 ℃ and kept for 30min to obtain the syrup.

And (4) mixing the syrup into peanut butter and uniformly mixing.

Example 8

Step (1), mixing 100g of stevia rebaudiana extract prepared in example 3, 7g of nutrient material, 2g of sorbitol, 5g of glutinous rice vinegar, 15g of blueberry juice and 3g of acacia, adding water, and stirring uniformly to fully dissolve the mixture;

the nutrient material comprises traditional Chinese medicine extract, vitamin A, vitamin C, lactalbumin and cysteine in a mass ratio of 10:1:2:10: 1.

The traditional Chinese medicine extract comprises the following raw materials in parts by weight: 150 parts of liquorice, 70 parts of sugarcane and 15 parts of dendrobium officinale.

The traditional Chinese medicine extract is prepared by the following method:

s2, extraction: adding 70mL of traditional Chinese medicine powder 10g into pure water, adding cellulase with the addition of 1100U/kg, adjusting the pH value to 5 by using 1mol/L hydrochloric acid, adjusting the temperature to 55 ℃, performing enzymolysis for 1h, and then performing enzyme deactivation, wherein the enzyme deactivation method comprises the steps of heating at 100 ℃ for 10min, adding sodium carbonate to adjust the pH value to be neutral, performing ultrasonic-assisted extraction for 2h under 1350W, centrifuging at the centrifugal condition of 4 ℃ and 11000r/min for 5min, collecting supernatant and concentrating, wherein the concentration method comprises the steps of performing ultrafiltration by adopting a ceramic membrane, the aperture of the ceramic membrane is 0.07 mu m, and the operating conditions of the ceramic membrane ultrafiltration are as follows: transmembrane pressure difference of 0.22MPa, filtration temperature of 55 deg.C, membrane surface flow rate of 8m/s, and freeze drying at-10 deg.C for 15min and-30 deg.C for 12 hr to obtain Chinese medicinal extract.

Standing for 24 hours, filtering, adding water to a specified amount, decocting with slow fire while stirring, decocting until the relative density is 4, and stopping heating;

And (4) uniformly mixing the syrup and auxiliary materials, and tabletting to obtain the stevia rebaudiana leaf mixed nutrient syrup tablets.

Comparative example 2

In comparison with example 8, the stevia leaf extract prepared in example 3 was replaced with the stevia leaf extract prepared in comparative example 1, and other materials and preparation methods were the same as example 8.

Comparative example 3

In comparison with example 8, the stevia leaf extract prepared in example 3 was replaced with a general stevia leaf extract, and other materials and preparation methods were the same as example 8. The extract of common stevia rebaudiana leaves is provided by Shaanxi Fuzheng Biotech limited.

Comparative example 4

Compared with the example 8, the nutrient material is not added with the traditional Chinese medicine extract, and other raw materials and the preparation method are the same as the example 8.

The nutrient material comprises vitamin A, vitamin C, lactalbumin and cysteine in a mass ratio of 1:2:20: 1.

Comparative example 5

Compared with the example 8, the nutrient material is not added with whey protein, and other raw materials and the preparation method are the same as the example 8.

The nutrient material is selected from traditional Chinese medicine extract, vitamin A, vitamin C and cysteine, and the mass ratio is 20:1:2: 1.

Test example 1Experimental study for anti-hepatic injury

Experimental materials, instruments and reagents

1. Experimental Material

Stevia rebaudiana leaves are collected from a New Cao farm in east Tai city of Jiangsu province, and are prepared according to the methods of examples 1-3 and comparative example 1 to obtain the stevia rebaudiana leaf extract. Stevia rebaudiana leaf mixed nutrient syrup prepared by the methods of examples 4-8 and comparative examples 2-5.

2. Laboratory apparatus

TG16-WS desk-top high-speed centrifuge, Hunan instrument laboratory Instrument development Co., Ltd; DK-98-1 model electric heating constant temperature water bath, Tianjin Tester instruments, Inc.; continuous Spectroscopy scanning microplate reader SpectraMaxPlus384 USA molecular biology Instrument company.

3. Experimental reagent

CCl₄Absolute ethanol, shanghai alatin Biotechnology, ltd; edible soybean oil, alanine Aminotransferase (ALT) kit and aspartate Aminotransferase (AST) kit, SOD kit, MDA kit, Coomassie brilliant blue kit, and Nanjing to build a bioengineering institute.

4. Laboratory animal

Male ICR mice of clean grade, body weight (20. + -. 2) g.

Second, Experimental methods

1. For CCl₄Influence of acute liver injury in mice

Mice were acclimatized for one week during which they had free access to water. After one week, the animals were randomly divided into 14 groups of 10 animals by weight, which were: is normalControl group, CCl₄Model groups, the stevia rebaudiana leaf extract low dose group (50mg/kg/d) prepared in examples 1-3, the middle dose group (100mg/kg/d), the high dose group (200mg/kg/d), the stevia rebaudiana leaf extract group (100mg/kg/d) prepared in comparative example 1, the stevia rebaudiana leaf mixed nutrient syrup group (100mg/kg/d) prepared in example 4, the stevia rebaudiana leaf mixed nutrient syrup group (100mg/kg/d) prepared in example 5, the stevia rebaudiana leaf mixed nutrient syrup group (100mg/kg/d) prepared in example 6, the stevia rebaudiana leaf mixed nutrient syrup group (100mg/kg/d) prepared in example 7, the stevia rebaudiana leaf mixed nutrient syrup group (100mg/kg/d) prepared in example 8, the stevia rebaudiana leaf mixed nutrient syrup group (100mg/kg/d) prepared in comparative example 2, the stevia rebaudiana leaf mixed nutrient syrup group prepared in comparative example 3 (100mg/kg/d), the stevia rebaudiana leaf mixed nutrient syrup group prepared in comparative example 4 (100mg/kg/d), the stevia rebaudiana leaf mixed nutrient syrup group prepared in comparative example 5 (100mg/kg/d), and the bifendate group (positive control group, 150mg/kg/d), all the drugs were dissolved in normal saline and the dissolution was assisted by ultrasound. Except normal control group and model group, the same amount of normal saline is used for intragastric administration according to body weight, and other administration groups are used for intragastric administration according to body weight. Gavage is performed 1 time per day for 7 days. In the 6 th day, fasting is performed for 12h without water prohibition, and after the normal administration is performed for 1h in the next day, 10% CCl is injected into abdominal cavity of each group except the normal control group₄Soybean oil solution to cause acute CCl₄Liver damage. After 6h, blood was taken from the eyeball, serum was centrifuged at 4200rpm for 10min, and the supernatant was stored in a refrigerator at 4 ℃. After blood sampling, mice are killed by cervical spondylolysis immediately, livers and spleens are dissected and taken out, after weighing, blood stains on the surfaces of the livers are washed off by normal saline, after the normal saline on the surfaces is wiped off, a part of the livers are soaked and fixed by 10% formalin, and the other part of the livers are wrapped by tinfoil and then are immediately placed into a refrigerator at the temperature of minus 80 ℃ for storage.

Serology and liver index detection: and (3) adding the centrifuged serum into a 96-well plate according to the instructions of ALT and AST kits, measuring the OD value by using an enzyme-labeling instrument, and calculating the activity of ALT and AST in the mouse serum.

Weighing a certain weight of frozen liver, adding cold normal saline according to the weight-volume ratio of 1:9 for homogenizing, centrifuging for 10min at 4200rpm in a refrigerated centrifuge, taking supernatant to obtain 10% of liver homogenate, and preparing the liver homogenate under the low temperature condition. A small amount of 10% liver homogenate was taken and diluted with cold physiological saline to 1% liver homogenate.

Loading samples in a 96-well plate according to the instruction of a Coomassie brilliant blue kit, SOD and MDA kit, respectively, measuring respective OD values by an enzyme labeling instrument, and respectively calculating the concentration of the hepatic protein in 1 percent of liver homogenate, the activity of the SOD in the liver and the concentration of the MDA in 10 percent of the liver homogenate.

2. Influence on acute liver injury of mice caused by alcohol

Mice were acclimatized for one week during which they had free access to water. After one week, the animals were randomly divided into 14 groups of 10 animals by weight, which were: a normal control group, an alcohol model group, a stevia leaf extract low dose group (50mg/kg/d) prepared in examples 1-3, a middle dose group (100mg/kg/d), a high dose group (200mg/kg/d), a stevia leaf extract group (100mg/kg/d) prepared in comparative example 1, a stevia leaf mixed nutrient syrup group (100mg/kg/d) prepared in example 4, a stevia leaf mixed nutrient syrup group (100mg/kg/d) prepared in example 5, a stevia leaf mixed nutrient syrup group (100mg/kg/d) prepared in example 6, a stevia leaf mixed nutrient syrup group (100mg/kg/d) prepared in example 7, a stevia leaf mixed nutrient syrup group (100mg/kg/d) prepared in example 8, the stevia rebaudiana mixed nutrient syrup group prepared in comparative example 2 (100mg/kg/d), the stevia rebaudiana mixed nutrient syrup group prepared in comparative example 3 (100mg/kg/d), the stevia rebaudiana mixed nutrient syrup group prepared in comparative example 4 (100mg/kg/d), the stevia rebaudiana mixed nutrient syrup group prepared in comparative example 5 (100mg/kg/d), and the bifendate group (a positive control group, 150mg/kg/d), all the medicines were dissolved by normal saline and the dissolution was assisted by ultrasound. Except for the blank control group and the model group, the same amount of normal saline is used for intragastric administration according to the body weight, and other administration groups are used for administrating corresponding medicines according to the body weight. Gavage is performed 1 time per day for 7 days. After the administration of the drug for 12h in the evening on the 6 th day, and after the normal administration for 1h on the next day, the rest groups except the blank control group are subjected to intragastric administration of 50% ethanol according to the dose of 12ml/kg to cause acute alcoholic liver injury. After 12h, the eye was bled, the serum was centrifuged at 4200rpm for 10min, and the supernatant was stored in a 4 ℃ refrigerator. After blood sampling, mice are killed by cervical spondylolysis immediately, livers and spleens are dissected and taken out, after weighing, blood stains on the surfaces of the livers are washed off by normal saline, after the normal saline on the surfaces is wiped off, a part of the livers are soaked and fixed by 10% formalin, and the other part of the livers are wrapped by tinfoil and then are immediately placed into a refrigerator at the temperature of minus 80 ℃ for storage.

Third, experimental results

1. For CCl₄Influence of acute liver injury in mice

The results of this experiment show that the test is carried out by CCl₄After injection, ALT and AST of the model mice are obviously increased and higher than those of the normal control group, which indicates that the model building is successful (p)<0.05), high dose, medium dose groups, and the groups given in examples 4-6 all reduced the increased ALT and AST activity in serum with significant differences (p) compared to the model group<0.05), examples 7-8 had very significant differences (p)<0.01); meanwhile, the high and medium dose groups of stevia rebaudiana leaf extract and the groups of examples 4-6 can effectively reduce the MDA level and increase the SOD level of liver tissues, and have significant difference (p)<0.05), examples 7-8 had very significant differences (p)<0.01), groups of comparative examples 1 to 5 had a decreasing trend and were not statistically significant (p)>0.05). Specific results are shown in table 1.

TABLE 1 influence of stevia rebaudiana leaf extract on serum ALT and AST and liver SOD and MDA of mice with CCl4 acute liver injury

Note that: #, dosing vs model group, p < 0.05; ﹡, dosing group vs model group, p <0.05, administration group vs model group, p < 0.01.

2. Influence on acute liver injury of mice caused by alcohol

The experimental result shows that after the stomach is perfused by 50% alcohol, ALT and AST of the model group mice are obviously increased and higher than those of a normal control group, which indicates that the model building is successful (p is less than 0.05), and the high-dose and medium-dose groups and the administration groups of examples 4-6 are compared with the model group, so that the ALT and AST activities increased in serum can be reduced, and the test results show that the test results have significant differences (p is less than 0.05), and the test results show that the test results in examples 7-8 have very significant differences (p is less than 0.01); meanwhile, the stevia rebaudiana leaf extract high and medium dose groups and the groups of examples 4 to 6 can effectively reduce the MDA level and increase the SOD level of the liver tissue, have significant difference (p <0.05), have extremely significant difference (p <0.01) in examples 7 to 8, have reduction trend in the groups of comparative examples 1 to 5, and have no statistical significance (p > 0.05). The specific results are shown in Table 2.

TABLE 2 influence of stevia rebaudiana leaf extract on serum ALT and AST and liver SOD and MDA of mice with acute liver injury caused by alcohol

Fourth, conclusion of experiment

Stevia leaf extract prepared in examples 1 to 3, stevia leaf sugar prepared in examples 4 to 8 of the present inventionThe pulp can obviously reduce CCl₄Or ALT and AST activity in serum of a model mouse with alcohol-induced liver injury and liver tissue MDA content, and liver SOD activity are enhanced, which shows that the active extract has obvious liver protection effect on chemical liver injury and alcoholic liver injury.

Comparative example 1 the preparation process differs from the inventive example 3 in that the stevia rebaudiana leaf extract prepared has a low yield and a poor activity of protecting liver, and has a tendency of recovering various indexes of liver, but is not obvious.

Comparative example 2 stevia leaf syrup prepared using the stevia leaf extract prepared in comparative example 1 also has poor activity of protecting liver.

Comparative example 3 compared to example 8, the stevia leaf extract prepared in example 3 was replaced with a general stevia leaf extract, and there was almost no activity of protecting liver.

Compared with the example 8, the comparative examples 4 and 5 are not added with the traditional Chinese medicine extract and the whey protein respectively, the activity of protecting the liver is poorer, although various indexes of the liver are recovered, the activity is not obvious, and the activity is obviously reduced greatly compared with the example 6. Therefore, the traditional Chinese medicine extract and the whey protein have synergistic effect.

Test example 2Blood sugar reduction experiment of stevia rebaudian leaf extract

1 materials and methods

1.1 materials

SPF-grade KM mice, male, (22 ± 2) g, purchased from slagoff test animals limited, hunan, production license number: SCXK (Xiang) 2018-.

1.2 Experimental methods

1.2.1 method for establishing hyperglycemic mouse model

SPF-grade KM male mice, about 22g, were fed with free diet for 3 days. Fasting for 16h, measuring fasting blood glucose and body weight, determining required streptozotocin amount according to body weight, preparing solution, and performing intraperitoneal injection of 50mg/kg^-1Injections were given for 5 days 1 time per day. After the last injection, fasting for 16h, measuring blood sugar and bloodThe sugar value is higher than 11.1 mmoL.L^-1The molding is regarded as successful; meanwhile, the physiological changes of the weight, drinking water, diet and the like of the mice are observed and recorded.

1.2.2 animal test design

Based on the established hyperglycemic mouse model method, 42 SPF-level KM mice are adaptively raised for 7 days at 22-25 ℃, weighed, 6 mice are randomly selected as normal groups, the rest mice are modeled, and 36 mice successfully modeled are randomly divided into a model group and a stevia rebaudiana extract high-dose group (the dose is 700mg kg. kg)^-1·d^-1) And stevia leaf extract middle dose group (dose is 350mg kg)^-1·d^-1) Stevia leaf extract low dose group (dose of 175mg kg)^-1·d^-1) The traditional Chinese medicine composition comprises a high-dose group of common stevia leaf extract and 6 metformin hydrochloride groups. The normal group and the model group are fed with water with equal volume, and the feeding volume is 10 mL/kg^-1·d^-1The mice were continuously fed with 28d, and the blood glucose levels of the mice in each group were measured at 7, 14, 21, and 28d, respectively.

1.2.3 data processing was performed using SPSS 22.0 software and results are expressed as "mean. + -. standard deviation (x. + -. sd)". The difference between groups is analyzed by adopting ONE-WAY analysis of variance (ONE-WAY ANOVA) and Duncan multiple comparison, wherein p is less than 0.05 to indicate that the difference is obvious, and p is less than 0.01 to indicate that the difference is extremely obvious. Correlation analysis was performed by Pearson's method.

2. Effects on blood glucose in hyperglycemic mice

The results are shown in FIG. 1, note: significant differences (p <0.01) compared to normal group; compared with the model group, # indicates significant difference (p <0.05) and # indicates very significant difference (p < 0.01).

As can be seen from FIG. 1, the blood glucose levels of the mice in each group before molding were all at normal levels, and the fasting blood glucose levels of the mice in each group after molding were significantly increased (P <0.01), and the blood glucose levels were all higher than 11.1 mmoL.L^-1Thus, the success of modeling the hyperglycemia model is demonstrated. After 28 days of feeding, compared with the model group, the blood sugar level of the stevia leaf extract in the high and medium dose groups and the metformin hydrochloride group is extremely reduced (P is less than 0.01), and the blood sugar level of the low dose group is significantly reduced (P is less than 0.05), thereby indicating that the sweet taste is generatedThe chrysanthemum syrup has obvious blood sugar reducing effect. The blood sugar of mice in the common stevia syrup group is slightly reduced, and the difference with the model group is not obvious, which indicates that the common stevia syrup does not have obvious blood sugar reducing effect.

Compared with the prior art, the method adopts water soaking and supercritical fluid extraction to prepare the active extract of the stevia rebaudiana leaf on the basis of the research on the original chemical components and the traditional efficacy of the stevia rebaudiana leaf, and compared with the prior art, the extracted stevia rebaudiana leaf extract can obviously reduce CCl₄Or ALT and AST activity in serum of a model mouse with alcohol-induced liver injury and liver tissue MDA content, and liver SOD activity are enhanced, which shows that the active extract has obvious liver protection effect on chemical liver injury and alcoholic liver injury. Therefore, the stevia rebaudiana leaf extract is expected to be developed into a new generation of safe and effective medicine, health-care product or food for preventing and treating various types of liver protection.

The stevia rebaudiana leaf syrup provided by the invention reduces the sugar content of the syrup, the stevioside component can obviously improve the sweetness of the syrup without increasing the blood sugar content of eaters, can be eaten by patients with diabetes, essential hypertension, hyperglycemia and hyperlipidemia, further has the effects of reducing blood pressure, blood sugar and blood fat and the like, and has wide application prospect.

The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents, improvements and the like that fall within the spirit and principle of the present invention are intended to be included therein.

Claims

1. A preparation method of stevia rebaudiana leaf mixed nutrient syrup is characterized in that the syrup at least comprises stevia rebaudiana leaf extract, nutrient extract, sweetener, brewed vinegar, fruit juice and a flavoring agent, and the preparation method comprises the following steps:

and (3) standing for 24 hours, filling and sterilizing to obtain the product.

2. The method of claim 1, wherein the stevia rebaudiana leaf extract is prepared by:

s1, cleaning stevia rebaudiana leaves, cutting into pieces, adding the stevia rebaudiana leaves into pure water for soaking, performing ultrasonic-assisted extraction for 30min, filtering, reserving filter residues, and freeze-drying the filtrate to obtain water extract freeze-dried powder;

the extraction conditions were:

3. The method for preparing the stevia rebaudiana leaf mixed nutrient syrup as claimed in claim 2, wherein the ultrasonic power is 1500-2000W, the freeze drying condition is that the mixture is frozen at-15 ℃ for 30min and then cooled to-35 ℃ for 10h, and the low-temperature drying is drying at-5 ℃ for 1 h.

4. The preparation method of the stevia rebaudiana leaf mixed nutrient syrup according to claim 1, wherein the mass ratio of the stevia rebaudiana leaf extract to the nutrient to the sweetener to the brewed vinegar to the fruit juice to the flavoring agent is 100 (5-10): 2: (3-7): (10-20): (2-5).

5. The method for preparing mixed nutrient syrup of stevia rebaudiana leaves as claimed in claim 1, wherein the nutrient is selected from one or more of Chinese medicinal extracts, vitamins, animal proteins, plant proteins and amino acids.

6. The method for preparing stevia rebaudiana leaf mixed nutrient syrup as claimed in claim 5, wherein the traditional Chinese medicine extract comprises the following raw materials in parts by weight: 100-200 parts of liquorice, 50-120 parts of sugarcane and 10-20 parts of dendrobium officinale.

7. The method for preparing stevia rebaudiana leaf mixed nutrient syrup as claimed in claim 5, wherein the Chinese herbal extract is prepared by the following method:

s2, extracting: adding the Chinese medicinal powder into pure water, adding cellulase, adjusting pH and temperature, performing enzymolysis for 0.5-1 hr, inactivating enzyme, adding sodium carbonate to adjust pH to neutral, performing ultrasonic-assisted extraction for 1-3 hr, centrifuging, collecting supernatant, concentrating, and freeze drying to obtain Chinese medicinal extract.

8. The method for preparing mixed nutrient syrup of stevia rebaudiana leaves as claimed in claim 7, wherein the pH value is adjusted to 5, the temperature is adjusted to 45-60 ℃, the enzyme deactivation method is heating at 100 ℃ for 10min, the ultrasonic power is 1200-: (5-10); the addition amount of the cellulase is 1000-1200U/kg, the concentration method adopts ceramic membrane ultrafiltration, the aperture of the ceramic membrane is 0.05-0.10 mu m, and the operation conditions of the ceramic membrane ultrafiltration are as follows: the transmembrane pressure difference is 0.17-0.30MPa, the filtration temperature is 52-57 ℃, and the membrane surface flow rate is 7-10 m/s.

9. Use of a stevia rebaudiana leaf mixed nutrient syrup prepared by the method of any one of claims 1 to 8 in the preparation of a medicament, health food or food for preventing and treating liver injury.

10. The use of claim 9, wherein the liver injury is chemical liver injury or alcoholic liver injury.