CN113796457A - Prebiotics-containing de-lactose pet milk and preparation method thereof - Google Patents
Prebiotics-containing de-lactose pet milk and preparation method thereof Download PDFInfo
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- CN113796457A CN113796457A CN202110966073.6A CN202110966073A CN113796457A CN 113796457 A CN113796457 A CN 113796457A CN 202110966073 A CN202110966073 A CN 202110966073A CN 113796457 A CN113796457 A CN 113796457A
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- milk
- lactose
- pectin
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/20—Animal feeding-stuffs from material of animal origin
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/10—Animal feeding-stuffs obtained by microbiological or biochemical processes
- A23K10/14—Pretreatment of feeding-stuffs with enzymes
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/10—Organic substances
- A23K20/163—Sugars; Polysaccharides
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K50/00—Feeding-stuffs specially adapted for particular animals
- A23K50/40—Feeding-stuffs specially adapted for particular animals for carnivorous animals, e.g. cats or dogs
- A23K50/48—Moist feed
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/0006—Homoglycans, i.e. polysaccharides having a main chain consisting of one single sugar, e.g. colominic acid
- C08B37/0045—Homoglycans, i.e. polysaccharides having a main chain consisting of one single sugar, e.g. colominic acid alpha-D-Galacturonans, e.g. methyl ester of (alpha-1,4)-linked D-galacturonic acid units, i.e. pectin, or hydrolysis product of methyl ester of alpha-1,4-linked D-galacturonic acid units, i.e. pectinic acid; Derivatives thereof
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
Abstract
The invention discloses a prebiotics-containing delactose pet milk and a preparation method thereof, wherein the prebiotics-containing delactose pet milk is milk with the mass content of lactose less than or equal to 0.5 percent and the mass content of unsaturated oligosaccharide of 2-20 g/L. The lactose content in the prebiotics-containing de-lactose pet milk is extremely low, and meets the standard of zero-lactose pet milk (the lactose mass content is lower than 0.5%) required in GB 28040-; the pectin oligosaccharide contains multiple active ingredients, and has effects in improving immunity, promoting health of intestine and stomach, and resisting tumor; the nutrition is rich, no chemical food additive is contained, the palatability is good, and the food can be accepted by animals; the preparation process is advanced and reasonable, the enzyme engineering technology is comprehensively applied, the process conditions are simple, the production period is short, and the product has wide market prospect.
Description
(I) technical field
The invention relates to a prebiotics-containing delactose pet milk and a preparation method thereof.
(II) background of the invention
Pure milk is one of the oldest natural beverages and is called 'white blood'. The preservation rate of the nutrient components in the sterilized fresh milk is usually more than 90%. Milk and its products are one of the important sources of protein, calcium, phosphorus, vitamin A, vitamin D, vitamin B2, etc. in the diet, contain a large amount of bioactive substances, have high nutritive value and are easy to digest and absorb. However, not only humans, but also animals, suffer from lactose intolerance. Lactose intolerance is a condition in which the patient's intestinal tract cannot secrete lactose-decomposing enzymes, but cannot digest and absorb lactose for use by the body, which is likely to cause diarrhea, abdominal distension, etc.
Pectin oligosaccharide is a low molecular carbohydrate, has a plurality of bioactive functions, and as a novel functional oligosaccharide, the excellent prebiotic property attracts more and more attention and researches, particularly the regulation effect on intestinal flora. At present, researches show that pectin oligosaccharide can reduce the number of bacteroides and clostridia and participate in growth regulation and adhesion processes of bifidobacteria in intestinal tracts, thereby reducing secretion of exotoxin and invasive enzymes, and particularly having good prevention effect on pseudomembranous colitis. The pectin oligosaccharide is also a natural bacteriostatic preservative, and can effectively inhibit or even kill putrefying bacteria in food and beverage, thereby playing a role in prolonging the shelf life of the food and beverage.
With the rapid development of social economy, the breeding heat of pets is raised nationwide, and with the continuous increase of the breeding quantity and types of family pets, the nutrition, safety and health of pet food are more and more valued by consumers, which also brings great opportunity for the development of the industries of pet food and health care products.
Disclosure of the invention
The invention aims to provide the pet milk containing prebiotics and the preparation method thereof, and the pectin oligosaccharide containing unsaturated double bonds is added as the prebiotics of the pet milk, so that the antioxidant property of the milk product can be improved, free radicals can be eliminated, and the pet milk product with a health care function is formed; the desugarized pet milk containing prebiotics can reduce the lactose content in milk, can adjust the intestinal flora of pets, and solves the problems of diarrhea, flatulence, vomiting and constipation, lactose intolerance of pets and the like caused by lactose intolerance in the prior art.
The technical scheme adopted by the invention is as follows:
the invention provides prebiotics-containing delactose pet milk, which is milk with the mass content of lactose less than or equal to 0.5 percent and the mass content of unsaturated oligosaccharide between 2 and 20 g/L.
The unsaturated oligosaccharide is unsaturated pectin oligosaccharide, has molecular weight of 3-5kD, and unsaturation degree of 7-10.
The unsaturated pectin oligosaccharide is prepared by the following method: adding high ester citrus pectin into citric acid-sodium dihydrogen phosphate buffer solution with pH of 4, adding pectin lyase, reacting at 35-55 deg.C for 4-8h (preferably 40 deg.C for 4h), boiling for 8-15min to inactivate enzyme, centrifuging (preferably 12000rpm for 5min), and retaining supernatant; adding yeast into the supernatant, reacting at 38 deg.C for 24 hr, centrifuging (preferably at 8000rpm for 8min), retaining the supernatant, respectively intercepting with 10KDa, 5KDa, and 3KDa membranes to obtain 3-5KDa retentate, lyophilizing at-80 deg.C and 100Pa for 48 hr to obtain unsaturated pectin oligosaccharide powder; the volume consumption of the citric acid-sodium dihydrogen phosphate buffer solution is 100mL/g based on the mass of the high-ester citrus pectin; the mass ratio of the volume consumption of the pectin lyase to the high-ester citrus pectin is 1mL/g, and the mass ratio of the high-ester citrus pectin to the yeast is 1: 0.5-2 (preferably 1: 1); the yeast is high-sugar-resistant Angel high-activity dry yeast.
The invention also provides a preparation method of the prebiotics-containing delactosed milk, which comprises the following steps: adding lactase into fresh milk, performing enzymolysis at 30-40 deg.C for 2-5h, inactivating enzyme at 65 deg.C for 10-30min, cooling to room temperature (25-30 deg.C), adding unsaturated pectin oligosaccharide, and stirring to obtain the final product.
The lactase specifically hydrolyzes lactose to glucose and galactose to reduce lactose content in milk; the enzyme activity of the lactase is more than or equal to 1 ten thousand U/g; the lactase is added in an amount of 5-40g/L, preferably 12g/L, based on the volume of the milk.
The unsaturated oligosaccharide is added in the form of unsaturated oligosaccharide powder or unsaturated oligosaccharide aqueous solution, and the mass addition amount of the unsaturated oligosaccharide is 2-20g/L (preferably 20 g/L) calculated by the volume of milk.
The pectin oligosaccharide is a novel functional saccharide source, and has good biological activity, such as inhibiting growth of pathogenic microorganisms in urinary tract, promoting apoptosis of human colon adenocarcinoma cells, resisting obesity, toxicity, infection, bacteria and oxidation. The unsaturated pectin oligosaccharide has excellent oxidation resistance due to the double bonds, has good free radical scavenging activity, and also has the functions of bacteriostasis, fresh keeping and the like.
Compared with the prior art, the invention has the following beneficial effects:
1. the lactose content in the prebiotics-containing de-lactose pet milk is extremely low, and meets the standard of zero-lactose pet milk (the lactose mass content is lower than 0.5%) required in GB 28040-;
2. the pectin oligosaccharide contains multiple active ingredients, and has effects in improving immunity, promoting health of intestine and stomach, and resisting tumor;
3. the pectin oligosaccharide milk is rich in nutrient components, does not contain any chemical food additive, is good in palatability and can be accepted by animals, the quality guarantee period of the product is greatly prolonged due to the addition of the pectin oligosaccharide, and the product is safer in the feeding process;
4. the preparation process is advanced and reasonable, the enzyme engineering technology is comprehensively applied, the process conditions are simple, the production period is short, and the product has wide market prospect.
(IV) description of the drawings
FIG. 1 is a diagram showing a model of the action of pectin lyase.
(V) detailed description of the preferred embodiments
The invention will be further described with reference to specific examples, but the scope of the invention is not limited thereto:
the milk used by the invention is fresh milk and is purchased from a mountain cattle house in a western lake region in Hangzhou city of Zhejiang province. The pectin is high ester citrus pectin (HMP) and is purchased from Henan Elite Biotech limited; the enzyme activity of the pectin lyase is 600U/mL, the enzyme activity of the lactase is more than or equal to 1 ten thousand U/g, and the pectin lyase and the lactase are purchased from Ningxia Xixia Seisan industry group GmbH. The yeast is high-sugar-resistant Angel high-activity dry yeast which is purchased from Angel Yeast GmbH. The room temperature is 25-30 ℃.
Example 1 preparation of unsaturated pectin oligosaccharides
Weighing 2g of high-ester citrus pectin, adding 200mL of citric acid-sodium dihydrogen phosphate buffer solution with pH of 4, adding 2mL of pectin lyase, reacting at 40 ℃ for 4h, boiling for 15min, and inactivating enzyme; centrifuge at 12000rpm for 5min, and retain the supernatant. 2g of yeast was added to the whole supernatant, reacted at 38 ℃ for 24 hours, centrifuged at 8000rpm for 8min, and the supernatant was retained. The supernatant fluid passes through a 10KDa filter membrane, filtrate passing through the 10KDa filter membrane passes through a 5KDa filter membrane, filtrate passing through the 5KDa filter membrane passes through a 3KDa filter membrane, retentate of the 3KDa filter membrane (namely, unsaturated pectin oligosaccharide solution of 3-5 KDa) is taken, and freeze-drying is carried out at 80 ℃ and 100Pa, so as to obtain 1.247g of unsaturated pectin oligosaccharide powder.
The unsaturation degree detection method comprises the following steps: the unsaturation degree of the pectin oligosaccharide is determined by an ultraviolet spectrophotometry method, because the pectin oligosaccharide containing unsaturated double bonds has ultraviolet absorption under 235nm wavelength, the absorbance value is in direct proportion to the content of the unsaturated double bonds. The unsaturated pectin oligosaccharide is prepared into a sample with the mass concentration of 1% by using distilled water, the unsaturated degree of the sample is calculated by using the absorbance value of the distilled water as a reference under the wavelength of 235nm, and the result is 7-10.
Formula for the unsaturation (DU):
C2H4containing a single double bond, UPOS represents unsaturation as a reference for unsaturated pectin oligosaccharidesPectin oligosaccharides.
Example 2:
taking 100mL of milk, adding 0.5g of lactase, carrying out enzymolysis for 2h at 35 ℃, inactivating the enzyme for 15min at 65 ℃, and cooling to room temperature to obtain the de-lactose pet milk, wherein the mass content of lactose is 1.271% through liquid phase detection, the in-vitro DPPH free radical clearance rate reaches 41.53%, and the unsealing, feeding and fresh-keeping time of the pet milk at room temperature is 6 h.
The lactose content detection method comprises the following steps: according to the national food safety standard GB 5009.8-2016, the lactose content is measured by high performance liquid chromatography. The liquid chromatographic column is an amino chromatographic column, and the mobile phase is acetonitrile: 70 parts of water: 30 (volume ratio), flow rate of 1.0ml/min, column temperature of 40 ℃, sample injection amount of 20 mul, detector of differential refraction detector, temperature of 40 ℃.
Method for determining DPPH radical scavenging Activity: accurately transferring 2mL of the delactosed pet milk As a sample, adding 2mL of 0.1mg/mL DPPH ethanol solution, uniformly mixing, standing at room temperature in the dark for 30min, measuring the light absorption value (As) at 517nm, using Vc to replace the sample As a standard reference substance, and using ethanol to replace DPPH solution As a blank reference. DPPH radical scavenging activity was calculated according to the following formula:
where Ac represents the absorbance at 517nm of the sample set replaced with 2mL of Vc, and Ab represents the absorbance at 517nm of the DPPH ethanol solution set replaced with 2mL of ethanol.
Ethanol solution of DPPH: weighing 10mg of DPPH (1, 1-diphenyl-2-trinitrophenylhydrazine), adding an ethanol solution to a constant volume of 100mL, preparing a 0.1mg/mL DPPH ethanol solution, and storing in a dark place.
Example 3:
taking 100mL of milk, adding 0.5g of lactase, carrying out enzymolysis for 2h at 35 ℃, inactivating the enzyme for 15min at 65 ℃, cooling to room temperature, then adding 0.2g of unsaturated pectin oligosaccharide prepared by the method in the embodiment 1, and uniformly stirring to obtain the desugarized pet milk containing prebiotics, wherein the detection by the method in the embodiment 1 shows that the mass content of lactose is 1.271%, the in-vitro DPPH free radical clearance rate reaches 70.08%, and the unsealing, feeding and fresh-keeping time of the pet milk at room temperature is prolonged by 4h and reaches 10 h.
Example 4:
taking 100mL of milk, adding 2g of lactase, carrying out enzymolysis for 3h at 30 ℃, inactivating enzyme at 65 ℃ for 15min, then adding 0.4g of unsaturated pectin oligosaccharide prepared by the method in the embodiment 1, and uniformly stirring to obtain the desugarized pet milk containing prebiotics, wherein the detection by the method in the embodiment 1 shows that the mass content of lactose is 0.578%, the in-vitro DPPH free radical clearance rate reaches 81.27%, and the unsealing, feeding and fresh-keeping time of the pet milk at room temperature is prolonged by 6h to 12 h.
Example 5:
taking 100mL of milk, adding 2g of lactase, carrying out enzymolysis for 4h at 40 ℃, inactivating the enzyme for 15min at 65 ℃, adding 0.6g of unsaturated pectin oligosaccharide prepared by the method in the embodiment 1, and uniformly stirring to obtain the de-lactose pet milk containing prebiotics, wherein the detection of the method in the embodiment 1 does not detect the lactose content, the product meets the standard of the zero-lactose pet milk required by the prepackaged label in GB28040 plus 2011, the in-vitro DPPH free radical clearance rate reaches 85.13%, and the unsealing, feeding and fresh-keeping time of the pet milk at room temperature is prolonged by 7h to 13 h.
Example 6:
taking 100mL of milk, adding 3g of lactase, carrying out enzymolysis for 3h at 35 ℃, inactivating the enzyme for 15min at 65 ℃, then adding 1g of unsaturated pectin oligosaccharide prepared by the method in the embodiment 1, and uniformly stirring to obtain the de-lactose pet milk containing prebiotics, wherein the detection by the method in the embodiment 1 shows that the lactose mass content is 0.422%, the product meets the standard of the zero-lactose pet milk required by a pre-packaged label in GB28040-2011, the in-vitro DPPH free radical clearance rate reaches 90.12%, and the unsealing, feeding and fresh-keeping time of the pet milk at room temperature is prolonged by 8h and reaches 14 h.
Example 7:
taking 100mL of milk, adding 4g of lactase, carrying out enzymolysis for 5h at 35 ℃, inactivating enzyme at 65 ℃ for 15min, then adding 0.4g of unsaturated pectin oligosaccharide prepared by the method in the embodiment 1, uniformly stirring, and uniformly stirring to obtain the de-lactose pet milk containing prebiotics, wherein the detection of the method in the embodiment 1 shows that the lactose content is not detected, the product meets the standard of the zero-lactose pet milk required by a prepackaged label in GB28040-2011, the in-vitro DPPH free radical clearance rate reaches 81.27%, and the unsealing, feeding and fresh-keeping time of the pet milk at room temperature is prolonged by 6h and reaches 12 h.
Claims (7)
1. The pet milk containing prebiotics and having the lactose removed therefrom is characterized in that the pet milk containing prebiotics and having the lactose content is less than or equal to 0.5 percent and the unsaturated oligosaccharide content is 2-20 g/L.
2. The prebiotic-containing delactosed pet milk of claim 1 wherein the unsaturated oligosaccharide is an unsaturated pectin oligosaccharide having a molecular weight of 3-5kD and an unsaturation of 7-10.
3. The prebiotic-containing, delactosed pet milk of claim 1, characterized in that the unsaturated pectin oligosaccharides are prepared as follows: adding high ester citrus pectin into citric acid-sodium dihydrogen phosphate buffer solution with pH of 4, adding pectin lyase, reacting at 35-55 deg.C for 4-8 hr, boiling for 8-15min to inactivate enzyme, centrifuging, and retaining supernatant; adding yeast into the supernatant, reacting at 38 deg.C for 24 hr, centrifuging, retaining supernatant, respectively intercepting with 10KDa, 5KDa and 3KDa membranes to obtain 3-5KDa retentate, and lyophilizing at-80 deg.C and 100Pa for 48 hr to obtain unsaturated pectin oligosaccharide powder.
4. The prebiotic-containing, delactosed pet milk of claim 3 wherein the citric acid-monosodium phosphate buffer solution is present in an amount of 100mL/g by volume based on the mass of high-ester citrus pectin; the mass ratio of the volume consumption of the pectin lyase to the high-ester citrus pectin is 1mL/g, and the mass ratio of the high-ester citrus pectin to the yeast is 1: 0.5 to 2; the yeast is high-sugar-resistant Angel high-activity dry yeast.
5. A method for preparing the prebiotic-containing delactosed milk of claim 1, characterized in that the method comprises: adding lactase into fresh milk, performing enzymolysis at 30-40 deg.C for 2-5h, inactivating enzyme at 65 deg.C for 10-30min, cooling to room temperature, adding unsaturated oligosaccharide, and stirring to obtain the final product.
6. The method according to claim 5, wherein the enzymatic activity of said lactase is greater than or equal to 1 million U/g; the addition amount of lactase is 5-40g/L based on the volume of milk.
7. The method according to claim 5, wherein the unsaturated oligosaccharide is added in an amount of 4-20g/L by volume of milk.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN114766591A (en) * | 2022-04-06 | 2022-07-22 | 天津朗诺宠物食品有限公司 | Freeze-dried lactose-free milk block and preparation method thereof |
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2021
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