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CN107858342B - Stable lactase liquid preparation - Google Patents

Stable lactase liquid preparation Download PDF

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CN107858342B
CN107858342B CN201710973692.1A CN201710973692A CN107858342B CN 107858342 B CN107858342 B CN 107858342B CN 201710973692 A CN201710973692 A CN 201710973692A CN 107858342 B CN107858342 B CN 107858342B
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lactase
oligosaccharide
preparation
liquid preparation
liquid
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CN107858342A (en
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李斌
邓紫玙
李晶
杨梅
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Huazhong Agricultural University
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/96—Stabilising an enzyme by forming an adduct or a composition; Forming enzyme conjugates
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
    • A23C9/00—Milk preparations; Milk powder or milk powder preparations
    • A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
    • A23C9/1203—Addition of, or treatment with, enzymes or microorganisms other than lactobacteriaceae
    • A23C9/1206—Lactose hydrolysing enzymes, e.g. lactase, beta-galactosidase
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
    • A23C9/00—Milk preparations; Milk powder or milk powder preparations
    • A23C9/152—Milk preparations; Milk powder or milk powder preparations containing additives
    • A23C9/1522—Inorganic additives, e.g. minerals, trace elements; Chlorination or fluoridation of milk; Organic salts or complexes of metals other than natrium or kalium; Calcium enrichment of milk
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
    • A23C9/00—Milk preparations; Milk powder or milk powder preparations
    • A23C9/152—Milk preparations; Milk powder or milk powder preparations containing additives
    • A23C9/156—Flavoured milk preparations ; Addition of fruits, vegetables, sugars, sugar alcohols or sweeteners
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
    • C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y302/00—Hydrolases acting on glycosyl compounds, i.e. glycosylases (3.2)
    • C12Y302/01—Glycosidases, i.e. enzymes hydrolysing O- and S-glycosyl compounds (3.2.1)
    • C12Y302/01108—Lactase (3.2.1.108)

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  • Food Science & Technology (AREA)
  • Polymers & Plastics (AREA)
  • Biotechnology (AREA)
  • Biomedical Technology (AREA)
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  • Medicinal Chemistry (AREA)
  • Inorganic Chemistry (AREA)
  • Dairy Products (AREA)
  • Enzymes And Modification Thereof (AREA)

Abstract

The invention discloses a stable lactase liquid preparation, and belongs to the field of food additives. The lactase liquid preparation consists of lactase, phosphate buffer solution, oligosaccharide and degraded sodium carboxymethylcellulose, wherein the oligosaccharide comprises one or more of isomaltooligosaccharide, trehalose, konjac oligosaccharide and chitosan oligosaccharide. The invention solves the technical problems of poor stability of lactase under the liquid condition, easy reduction of enzyme activity and the like, and the preparation can be directly added into dairy products and can quickly act with lactose, thereby being particularly suitable for infants.

Description

Stable lactase liquid preparation
Technical Field
The invention belongs to the field of food additives, and particularly relates to a lactase liquid preparation for a dairy product additive.
Background
The activity of lactase of adults is gradually reduced with age, and the activity of the lactase is only 5% -10% of the normal infant level after adults, and the lactase intolerance develops. Lactase deficiency occurs to varying degrees in most of the world's population, except for caucasian races, with asians and africans occurring at 75% -100%. The incidence rate of lactase deficiency of Chinese adults is as high as 75% -95.7%. The existence of lactose intolerance limits the intake of milk and dairy products to a great extent, and can generate a series of adverse effects.
Lactase can alter lactose malabsorption, but there are some problems: firstly, lactase is added into milk, and the lactase can hydrolyze lactose in the milk in advance under the optimal condition, so that the sweetness of the milk is increased by 3-4 times, and people do not like the lactase; ② the lactase tablet taken in can be degraded in gastrointestinal tract, at the same time other food can destroy the activity of enzyme, and the temperature and pH value of gastrointestinal tract can also affect the activity of enzyme. The above problems can be solved by the following means: lactase is prepared into tablets and water aqua and is taken orally before milk is drunk or is directly dripped into milk for drinking; or making into lactase lipid carrier to prevent degradation of lactase in gastrointestinal tract; and searching for the lactase with the optimum conditions similar to the pH value and the temperature of the gastrointestinal tract.
Currently, there are a wide variety of lactase products on the market, roughly divided into 4 types: 1, tablet preparation: the adopted lactase is acidic lactase, namely the most suitable pH of the lactase is about 3.5 (the product is not easy to be damaged by gastric acid); 2: solid particle beverage: is a probiotic (bifidobacteria) containing lactase, rather than directly added; 3, capsule preparation: the enteric capsule is wrapped, so that the capsule cannot be damaged by gastric acid; 4 liquid lactase product. The first 3 dosage forms are useful for adults but also not for infants who cannot swallow tablets or granules, and therefore liquid lactase formulations are most suitable for use by infants. Meanwhile, when the lactase is taken as a tablet or a capsule, the lactase does not easily and quickly act with lactose in milk products, so that the problem can be solved by developing a stable lactase liquid preparation, and the lactase liquid preparation is convenient to use and quick in action.
Oligosaccharides are a food ingredient that can be selectively fermented and alter the composition and activity of the beneficial flora of the gastrointestinal tract for the health of the host, and are frequently used in dairy products to improve the nutrition and absorption of milk. Degradation products of hydrocolloids have also been tried for protein protectants, but there has been no report to date of degradation of sodium carboxymethylcellulose as a protein protectant. It is of great significance to obtain cheap and efficient compound to stabilize liquid lactase preparation.
Disclosure of Invention
Aiming at the defects in the prior art, the prebiotics, the degraded sodium carboxymethylcellulose and the lactase are combined, so that the stability of the liquid lactase can be improved, the lactose intolerance can be relieved, the intestinal flora can be adjusted, and the gastrointestinal environment can be improved.
A stable lactase liquid preparation, which consists of the following components:
Figure BDA0001430645050000021
preferably, the lactase liquid preparation consists of the following components:
Figure BDA0001430645050000022
preferably, the oligosaccharide is one or more of isomaltooligosaccharide, trehalose, konjac oligosaccharide and chitosan oligosaccharide.
Preferably, the oligosaccharide is a composition of isomaltooligosaccharide and konjac oligosaccharide, and the weight ratio of the isomaltooligosaccharide to the konjac oligosaccharide is 3: 4.
preferably, the molecular weight of the degraded sodium carboxymethylcellulose is 5-10KDa, and the viscosity is 100-150mPa & s.
Preferably, the phosphate buffer has a pH of 5 to 7 and comprises dipotassium hydrogen phosphate and potassium dihydrogen phosphate in a weight ratio of 1:0.8 to 1.1.
The lactase liquid preparation can be added into dairy products, and the addition amount is 1-3% (volume).
Compared with the prior art, the invention has the advantages that:
1. the invention provides a lactase liquid preparation, which solves the technical problems that the lactase has poor stability under the liquid condition, the enzyme activity is easy to reduce and the like, and the preparation can be directly added into dairy products and can quickly play a role with lactose, so the preparation is particularly suitable for infants.
2. The raw materials used by the invention are all natural products, and the invention has the advantages of safety, reliability, low cost, convenient use and the like.
Detailed Description
The present invention will be described in detail below with reference to specific examples.
Example 1
A stable lactase liquid preparation, which consists of the following components:
Figure BDA0001430645050000031
the preparation method comprises the following steps:
(1) 0.8g of dipotassium hydrogenphosphate and 0.88g of potassium dihydrogenphosphate were dissolved in 100mL of ultrapure water, and the pH was adjusted to about 6.5.
(2) 30g of isomaltooligosaccharide, 40g of konjac oligosaccharide and 0.5g of degraded sodium carboxymethylcellulose (molecular weight of 5-10KDa, viscosity of 100-150mPa · s, degree of substitution of 0.7, obtained by degrading sodium carboxymethylcellulose with cellulase) were dissolved in the phosphate buffer solution of step (1) and stirred for about 3 hours.
(3) 0.8g lactase (1000-3000U/g) was added to the solution of step (2) and stirred for 1 h.
The liquid preparation can be applied to breast milk or formula milk, and the addition amount is as follows: 1-3mL/100mL milk.
Example 2
A stable lactase liquid preparation, which consists of the following components:
Figure BDA0001430645050000032
the preparation method comprises the following steps:
(1) 0.8g of dipotassium hydrogenphosphate and 0.88g of potassium dihydrogenphosphate were dissolved in 100mL of ultrapure water, and the pH was adjusted to about 6.5.
(2) 30g of isomaltooligosaccharide, 20g of chitosan oligosaccharide and 0.3g of degraded sodium carboxymethylcellulose (molecular weight of 10-15KDa, viscosity of 150-200mPa & s, degree of substitution of 0.9) were dissolved in the buffer solution of step (1) and stirred for about 3 hours.
(3) 0.8g lactase (1000-3000U/g) was added to the solution of step (2) and stirred for 1 h.
Example 3
A stable lactase liquid preparation, which consists of the following components:
Figure BDA0001430645050000041
the preparation method comprises the following steps:
(1) 0.8g of dipotassium hydrogenphosphate and 0.7g of potassium dihydrogenphosphate were dissolved in 100mL of ultrapure water, and the pH was adjusted to about 6.5.
(2) 25g of isomaltose hypgather, 40g of trehalose and 0.3g of degraded sodium carboxymethylcellulose (molecular weight of 15-20KDa, viscosity of 200-250 mPas, degree of substitution of 1.2) were dissolved in the buffer solution of step (1) and stirred for about 3 hours.
(3) 0.8g lactase (1000-3000U/g) was added to the solution of step (2) and stirred for 1 h.
Example 4
A stable lactase liquid preparation, which consists of the following components:
Figure BDA0001430645050000042
the preparation method comprises the following steps:
(1) 0.8g of dipotassium hydrogenphosphate and 0.8g of monopotassium dihydrogenphosphate were dissolved in 100mL of ultrapure water, and the pH was adjusted to about 6.5.
(2) 20g of isomaltooligosaccharide, 30g of konjac oligosaccharide, 20g of trehalose and 0.1g of sodium carboxymethylcellulose (molecular weight of 20-30KDa, viscosity of 250-300mPa & s and substitution degree of 1.2) were dissolved in the buffer solution in the step (1) and stirred for about 3 hours.
(3) 0.8g lactase (1000-3000U/g) was added to the solution of step (2) and stirred for 1 h.
Example 5
A stable lactase liquid preparation, which consists of the following components:
Figure BDA0001430645050000051
the preparation method is the same as that of example 1.
Example 6
A stable lactase liquid preparation, which consists of the following components:
Figure BDA0001430645050000052
the preparation method is the same as that of example 1.
Example 7
A stable lactase liquid preparation, which consists of the following components:
Figure BDA0001430645050000053
the preparation method is the same as that of example 1.
Test examples
The stability studies were performed on the lactase liquid formulations prepared in examples 1-7: respectively storing the lactase liquid at 60 ℃ for 1 h; storing at 4 deg.C, 25 deg.C and 40 deg.C for 20 days, measuring the retention rate of enzyme activity and examining thermal stability.
Comparative example 1: the other ingredients and preparation method are the same as example 1 without degraded sodium carboxymethyl cellulose.
Comparative example 2: the degraded sodium carboxymethylcellulose was replaced with ordinary sodium carboxymethylcellulose (CMC), and other ingredients and preparation methods were the same as in example 1.
Blank: lactase liquid preparation without oligosaccharide and without sodium carboxymethyl cellulose degradation.
The results are given in the following table:
TABLE 1 enzyme Activity Retention (%)
Figure BDA0001430645050000061
As can be seen from Table 1, the lactase liquid preparation added with oligosaccharide has high stability, especially the lactase liquid preparation added with isomaltooligosaccharide and konjac oligosaccharide has the enzyme activity retention rate of more than 80% under the high temperature condition (60 ℃), so the stability is the best. The addition of degraded sodium carboxymethylcellulose to the lactase liquid formulation further improves stability, while the addition of sodium carboxymethylcellulose has no protective effect.

Claims (6)

1. A stable lactase liquid preparation, which is characterized by comprising the following components:
Figure FDA0002560885530000011
the oligosaccharide is one or more of isomaltooligosaccharide, trehalose, konjac oligosaccharide and chitosan oligosaccharide.
2. The liquid lactase formulation according to claim 1, consisting of:
Figure FDA0002560885530000012
3. the lactase liquid formulation of claim 1, wherein: the oligosaccharide is a composition of isomaltooligosaccharide and konjac oligosaccharide, and the weight ratio of the isomaltooligosaccharide to the konjac oligosaccharide is 3: 4.
4. the lactase liquid formulation of claim 1, wherein: the molecular weight of the degraded sodium carboxymethylcellulose is 5-10KDa, and the viscosity is 100-150mPa & s.
5. The lactase liquid formulation of claim 1, wherein: the phosphate buffer has a pH of 5-7 and comprises dipotassium hydrogen phosphate and potassium dihydrogen phosphate in a weight ratio of 1: 0.8-1.1.
6. Use of the lactase liquid formulation of any one of claims 1 to 5 as a dairy additive in an amount of 1 to 3% by volume of the dairy product.
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Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH02110198A (en) * 1988-10-18 1990-04-23 Kao Corp Method for stabilization of enzyme
CN100447242C (en) * 2001-04-04 2008-12-31 Dsmip资产有限公司 Purified lactase
CN102172254A (en) * 2011-04-02 2011-09-07 内蒙古蒙牛乳业（集团）股份有限公司 Liquid dairy product and preparation method thereof
CN102232417A (en) * 2011-06-30 2011-11-09 北京三元食品股份有限公司 Galactosyl oligosaccharide-enriching milk and preparation method thereof
CN102559647A (en) * 2011-12-31 2012-07-11 湖南利尔康生物有限公司 Method for improving application effects of liquid enzyme
CN103082000A (en) * 2013-02-06 2013-05-08 内蒙古蒙牛乳业（集团）股份有限公司 Low-lactose liquid milk product containing fish oil and production method thereof

Patent Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH02110198A (en) * 1988-10-18 1990-04-23 Kao Corp Method for stabilization of enzyme
CN100447242C (en) * 2001-04-04 2008-12-31 Dsmip资产有限公司 Purified lactase
CN102172254A (en) * 2011-04-02 2011-09-07 内蒙古蒙牛乳业（集团）股份有限公司 Liquid dairy product and preparation method thereof
CN102232417A (en) * 2011-06-30 2011-11-09 北京三元食品股份有限公司 Galactosyl oligosaccharide-enriching milk and preparation method thereof
CN102559647A (en) * 2011-12-31 2012-07-11 湖南利尔康生物有限公司 Method for improving application effects of liquid enzyme
CN103082000A (en) * 2013-02-06 2013-05-08 内蒙古蒙牛乳业（集团）股份有限公司 Low-lactose liquid milk product containing fish oil and production method thereof

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
HECTOR L. RAMI＇REZ等.Invertase Stabilization by Chemical Modification of Sugar Chains with Carboxymethylcellulose.《Journal of BIOACTIVE AND COMPATIBLE POLYMERS》.2002,第17卷 *
Xyloglucan-based diblock co-oligomer: Synthesis, self-assembly and steric stabilization of proteins;Cony Gauche等;《Carbohydrate Polymers》;20130806;第98卷;第1271-1280页 *
多糖/蛋白质体系相转变及其在食品中的应用;许威等;《食品科技》;20170131;第42卷(第01期);第76-81页 *

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