CN111363766A - Preparation method of structural lipid for improving DHA bioavailability and product thereof - Google Patents

Preparation method of structural lipid for improving DHA bioavailability and product thereof Download PDF

Info

Publication number
CN111363766A
CN111363766A CN202010198628.2A CN202010198628A CN111363766A CN 111363766 A CN111363766 A CN 111363766A CN 202010198628 A CN202010198628 A CN 202010198628A CN 111363766 A CN111363766 A CN 111363766A
Authority
CN
China
Prior art keywords
dha
content
product
monoglyceride
diglyceride
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Granted
Application number
CN202010198628.2A
Other languages
Chinese (zh)
Other versions
CN111363766B (en
Inventor
邹孝强
张晖
金青哲
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Jiangnan University
Original Assignee
Jiangnan University
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Jiangnan University filed Critical Jiangnan University
Priority to CN202010198628.2A priority Critical patent/CN111363766B/en
Publication of CN111363766A publication Critical patent/CN111363766A/en
Application granted granted Critical
Publication of CN111363766B publication Critical patent/CN111363766B/en
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • 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
    • C12P7/00—Preparation of oxygen-containing organic compounds
    • C12P7/64—Fats; Fatty oils; Ester-type waxes; Higher fatty acids, i.e. having at least seven carbon atoms in an unbroken chain bound to a carboxyl group; Oxidised oils or fats
    • C12P7/6409—Fatty acids
    • C12P7/6427—Polyunsaturated fatty acids [PUFA], i.e. having two or more double bonds in their backbone
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/115—Fatty acids or derivatives thereof; Fats or oils
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/115—Fatty acids or derivatives thereof; Fats or oils
    • A23L33/12—Fatty acids or derivatives thereof
    • 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
    • C12P7/00—Preparation of oxygen-containing organic compounds
    • C12P7/64—Fats; Fatty oils; Ester-type waxes; Higher fatty acids, i.e. having at least seven carbon atoms in an unbroken chain bound to a carboxyl group; Oxidised oils or fats
    • C12P7/6436—Fatty acid esters
    • C12P7/6445—Glycerides
    • C12P7/6472—Glycerides containing polyunsaturated fatty acid [PUFA] residues, i.e. having two or more double bonds in their backbone
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs

Landscapes

  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Organic Chemistry (AREA)
  • Engineering & Computer Science (AREA)
  • Oil, Petroleum & Natural Gas (AREA)
  • Zoology (AREA)
  • Health & Medical Sciences (AREA)
  • Wood Science & Technology (AREA)
  • Nutrition Science (AREA)
  • Mycology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Microbiology (AREA)
  • Polymers & Plastics (AREA)
  • Food Science & Technology (AREA)
  • Biotechnology (AREA)
  • Biochemistry (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)
  • Edible Oils And Fats (AREA)

Abstract

The invention discloses a preparation method of structural lipid for improving DHA bioavailability and a product thereof, wherein the preparation method comprises the steps of carrying out enzyme catalysis alcoholysis reaction to prepare an intermediate enzymolysis product rich in DHA; adding the intermediate enzymolysis product and glycerol into an intermittent reactor according to a molar ratio of 1: 3-10, adding 3-12 wt% of lipase, and reacting for 3-12 hours at the temperature of 40-70 ℃ and at a stirring speed of 300-800 rpm to obtain a monoglyceride product rich in DHA; adding the intermediate enzymolysis product and glycerol into an intermittent reactor according to a molar ratio of 1: 0.5-2, adding 3-15 wt% of lipase, and reacting for 4-10 hours at the temperature of 40-70 ℃ and at a stirring speed of 300-800 rpm to obtain a diglyceride product rich in DHA. According to the invention, a DHA-enriched monoglyceride or diglyceride product is prepared by two-step enzymatic catalysis for the first time, and the DHA content in the product exceeds that reported in the same type, so that the product can play a role of predigestion, is beneficial to the digestion and absorption of DHA, and improves the bioavailability of DHA.

Description

Preparation method of structural lipid for improving DHA bioavailability and product thereof
Technical Field
The invention belongs to the technical field of grease, and particularly relates to a preparation method of structural lipid for improving DHA bioavailability and a product thereof.
Background
Docosahexaenoic acid (DHA, C22: 6n-3) is an important substance for invertebrate and vertebrate development and plays an important role in improving the neural and retinal development of infants and in reducing the incidence of cardiovascular disease DHA is an end product of n-3 fatty acids, which can be synthesized from its essential dietary precursor, α -linolenic acid (C18: 3n-3), the transformation pathway consisting of a series of fatty acid desaturation and elongation processes.
Breast milk is the best food for infants and contains a certain amount of Triglycerides (TAG) or phospholipids containing DHA (0.32 ± 0.22%, range: 0.06-1.4%). As a substitute for breast milk, infant formulas are currently supplemented with oils rich in DHA. DHA may be derived from deep-sea fish and microalgae. The fish oil contains large amount of eicosapentaenoic acid (EPA, C20:5n-3), and high content of EPA reduces synthesis of eicosanoid containing arachidonic acid, and has effect in inhibiting growth of newborn. Most fish oils are unsuitable as supplements for infant formula, DHA-rich microbial oils, produced by fermentation of microalgae such as Crypthecodinium cohnii, schizochytrium sp, and Ulkenia sp, are free of EPA and possible contaminants such as dioxins, polychlorinated biphenyls, heavy metals, etc., and thus, microbially produced DHA-rich oils are widely used in infant formula as supplements.
It is well known that the digestive system of infants, especially newborns, has not developed well. Gastric lipase levels are similar to those of adults, while pancreatic lipase and bile salt levels are relatively low and the ability to digest and absorb fat is limited, which severely affects the bioavailability of some important physiologically active lipids. Therefore, it is necessary to modify lipids having important physiological functions into a form that is easily absorbed. In view of the important physiological function of DHA, DHA is converted into diglyceride or monoglyceride form by a certain method, and the process can play a role of predigestion and is beneficial to the digestive absorption of DHA. Meanwhile, the diglyceride and monoglyceride rich in DHA can be used as an emulsifier in food and also an important food additive for supplementing DHA to general people.
Currently, many studies report the synthesis of diglycerides and monoglycerides by using conventional edible oils such as soybean oil, rapeseed oil, and the like, using glycerolysis, alcoholysis, and esterification. In the preparation of DHA-rich diglycerides and monoglycerides, researchers have mainly used marine fish oil as a raw material. As mentioned above, marine oils contain EPA, which is detrimental to the growth and development of infants.
Disclosure of Invention
This section is for the purpose of summarizing some aspects of embodiments of the invention and to briefly introduce some preferred embodiments. In this section, as well as in the abstract and the title of the invention of this application, simplifications or omissions may be made to avoid obscuring the purpose of the section, the abstract and the title, and such simplifications or omissions are not intended to limit the scope of the invention.
The present invention has been made keeping in mind the above and/or other problems occurring in the prior art.
Therefore, the present invention aims to overcome the defects in the prior art and provide a method for preparing structural lipid for improving the bioavailability of DHA.
In order to solve the technical problems, the invention provides the following technical scheme: a method for preparing structural lipid for improving DHA bioavailability comprises an enzyme catalysis alcoholysis reaction: adding microbial oil containing DHA and ethanol into an intermittent reactor according to a molar ratio of 1: 2-10, adding 3-12 wt% of lipase, and reacting for 3-12 hours at a temperature of 30-50 ℃ and a stirring speed of 300-800 rpm to prepare an intermediate enzymolysis product rich in DHA, wherein the content of monoglyceride and diglyceride in the intermediate enzymolysis product is more than 30%, and the content of monoglyceride is more than 3%; preparing monoglyceride: after ethanol and fatty acid ethyl ester of the intermediate enzymolysis product are respectively removed by adopting reduced pressure distillation and molecular distillation, adding the intermediate enzymolysis product and glycerol into an intermittent reactor according to the molar ratio of 1: 3-10, adding 3-12 wt% of lipase, and reacting for 3-12 hours at the temperature of 40-70 ℃ and at the stirring speed of 300-800 rpm to obtain a monoglyceride product rich in DHA, wherein the monoglyceride content is more than 60%, the DHA content is more than 65%, the polyunsaturated fatty acid content is more than 80%, the saturated fatty acid content is less than 20%, and the DHA content in the monoglyceride is more than 65%; preparing diglyceride: after ethanol and fatty acid ethyl ester of the intermediate enzymolysis product are respectively removed by adopting reduced pressure distillation and molecular distillation, adding the intermediate enzymolysis product and glycerol into an intermittent reactor according to the molar ratio of 1: 0.5-2, adding 3-15 wt% of lipase, and reacting for 4-10 hours at the temperature of 40-70 ℃ and at the stirring speed of 300-800 rpm to obtain a diglyceride product rich in DHA, wherein the content of diglyceride is more than 50%, the content of DHA is more than 65%, the content of polyunsaturated fatty acid is more than 80%, the content of saturated fatty acid is less than 20%, and the content of DHA in diglyceride is more than 65%.
As a preferred embodiment of the method for preparing structural lipid for improving DHA bioavailability according to the present invention, the method comprises: the enzyme catalyzes alcoholysis reaction, wherein the used Lipase comprises Lipozyme RM IM, Lipozyme TL IM or Lipase AY-30 SD.
As a preferred embodiment of the method for preparing structural lipid for improving DHA bioavailability according to the present invention, the method comprises: the preparation of monoglyceride, wherein the lipase used is Novozym 435.
As a preferred embodiment of the method for preparing structural lipid for improving DHA bioavailability according to the present invention, the method comprises: the preparation of diglyceride, wherein the lipase used is Novozym 435.
As a preferred embodiment of the method for preparing structural lipid for improving DHA bioavailability according to the present invention, the method comprises: the DHA-containing microbial oil is one or more of microbial oils produced by fermenting microalgae such as Crypthecodinium cohnii, Schizochytrium sp.
As a preferred embodiment of the method for preparing structural lipid for improving DHA bioavailability according to the present invention, the method comprises: the glycerol is food grade glycerol.
As a preferred embodiment of the method for preparing structural lipid for improving DHA bioavailability according to the present invention, the method comprises: the enzymatic alcoholysis reaction is carried out, wherein the DHA content of the intermediate enzymolysis product is more than 65%, the polyunsaturated fatty acid content is more than 80%, the saturated fatty acid content is less than 20%, the content of monoglyceride and diglyceride is more than 30%, and the content of monoglyceride is more than 3%.
As a preferred embodiment of the method for preparing structural lipid for improving DHA bioavailability according to the present invention, the method comprises: the DHA-rich monoglyceride product is obtained, wherein the content of monoglyceride in the product is more than 60%, the content of DHA is more than 65%, the content of polyunsaturated fatty acid is more than 80%, the content of saturated fatty acid is less than 20%, and the content of DHA in monoglyceride is more than 65%.
As a preferred embodiment of the method for preparing structural lipid for improving DHA bioavailability according to the present invention, the method comprises: the DHA-rich diglyceride product is obtained, wherein the content of diglyceride in the product is more than 50%, the content of DHA in the product is more than 65%, the content of polyunsaturated fatty acid is more than 80%, the content of saturated fatty acid is less than 20%, and the content of DHA in diglyceride is more than 65%.
It is another object of the present invention to overcome the deficiencies of the prior art and to provide a product obtained by the method for preparing structural lipids for improving the bioavailability of DHA.
As a preferred embodiment of the product obtained by the preparation method of the structural lipid for improving the bioavailability of DHA of the present invention, the method comprises: the product comprises a DHA-rich monoglyceride product and a DHA-rich diglyceride product, wherein the content of monoglyceride in the DHA-rich monoglyceride product is more than 60%, the content of diglyceride in the DHA-rich diglyceride product is more than 50%, and the content of DHA in both the monoglyceride and the diglyceride is more than 65%.
The invention has the beneficial effects that:
(1) according to the invention, conventional fatty acid in microbial oil containing DHA is replaced by utilizing the selectivity of lipase to a substrate through alcoholysis reaction, and DHA is enriched in an intermediate enzymolysis product; and then the enzymatic glycerolysis reaction is utilized to respectively obtain the monoglyceride or diglyceride product rich in DHA. In the second step of reaction, the reaction system is changed into a microemulsion system after the glycerol is added and stirred, so that the reaction rate is higher in the second step of reaction, and the production efficiency is greatly improved.
(2) The method creatively adopts the two-step enzyme method to catalyze and prepare the DHA-enriched monoglyceride or diglyceride product, the content of the monoglyceride in the prepared DHA-enriched monoglyceride product is more than 60%, the content of the diglyceride in the DHA-enriched diglyceride product is more than 50%, the content of the DHA in both the monoglyceride and the diglyceride is more than 65%, and the DHA content in the product exceeds that reported by the same kind, so that the predigestion effect can be achieved, the digestion and absorption of the DHA are facilitated, and the bioavailability of the DHA is improved.
Detailed Description
In order to make the aforementioned objects, features and advantages of the present invention more comprehensible, specific embodiments thereof are described in detail below with reference to examples of the specification.
In the following description, numerous specific details are set forth in order to provide a thorough understanding of the present invention, but the present invention may be practiced in other ways than those specifically described and will be readily apparent to those of ordinary skill in the art without departing from the spirit of the present invention, and therefore the present invention is not limited to the specific embodiments disclosed below.
Furthermore, reference herein to "one embodiment" or "an embodiment" means that a particular feature, structure, or characteristic described in connection with the embodiment is included in at least one implementation of the invention. The appearances of the phrase "in one embodiment" in various places in the specification are not necessarily all referring to the same embodiment, nor are separate or alternative embodiments mutually exclusive of other embodiments.
Example 1
This example provides a method for preparing structural lipids that increase DHA bioavailability:
(1) adding DHA-enriched algae oil from Schizochytrium sp into a sealable batch reactor, adding ethanol according to the molar ratio of 1:8 (algae oil: ethanol), adding 4 wt% Lipozyme RM IM, sealing the reactor, and reacting at 45 ℃ and the stirring speed of 500rpm for 10 hours to obtain an intermediate enzymolysis product enriched in DHA.
(2) Firstly, removing ethanol from the intermediate product by rotary evaporation, wherein the conditions of the rotary evaporation are as follows: the temperature is 50 ℃, the rotating speed is 60rpm, the time is 20min, and the vacuum degree is 0.095 MPa.
(3) Further removing the generated fatty acid ethyl ester from the ethanol-removed intermediate product by molecular distillation, wherein the conditions of the molecular distillation are as follows: the evaporation temperature is 185 ℃; heat exchanger temperature, 60 ℃; rotational speed, 120 rpm; absolute pressure, 2 Pa. The glyceride and fatty acid compositions of the intermediate products obtained are shown in table 1.
TABLE 1 chemical composition of intermediate enzymatic products
Fatty acids Algae oil Intermediate product Glyceride composition Content (wt.)
C14:0 2.8 0.7 Triglycerides 63.3
C16:0 33.7 9.7 Diglyceride 31.9
C18:0 2.5 0.5 Monoglyceride 4.8
C18:1 1.2 0.2
C20:4 0.78 0.6
C20:5 1.12 0.4
C22:5 10.6 17.9
C22:6 47.1 68.9
Saturated fatty acid 39 10.9
Polyunsaturated fatty acids 59.6 87.8
(4) Taking the DHA-enriched intermediate enzymolysis product as an initiator, adding glycerol into a batch reactor according to the molar ratio of 1:1 (intermediate enzymolysis product: glycerol), and reacting for 5 hours under the conditions that the addition amount of Novozym 435 is 8 wt%, the temperature is 50 ℃ and the stirring speed is 600rpm, so as to obtain a DHA-enriched diglyceride product. The glyceride composition of the product obtained is shown in table 2.
TABLE 2 composition of diglyceride product
Glyceride composition Content (wt.)
Triglycerides 26.0
Diglyceride 55.8
Monoglyceride 18.2
The fatty acid composition of the resulting glycerides is shown in table 3.
TABLE 3 glyceride fatty acid composition of the resulting product
Figure BDA0002418540610000051
Figure BDA0002418540610000061
(5) Taking the obtained DHA-enriched intermediate product as an initial material, adding glycerol into a batch reactor according to a molar ratio (intermediate enzymolysis product: glycerol) of 1:3, and reacting for 10 hours under the conditions that the addition amount of Novozym 435 is 12 wt%, the temperature is 50 ℃ and the stirring speed is 600rpm, so as to obtain a DHA-enriched monoglyceride product. The glyceride composition of the product obtained is shown in table 4.
TABLE 4 composition of monoglyceride product
Glyceride composition Content (wt.)
Triglycerides 8.1
Diglyceride 23.3
Monoglyceride 68.6
The fatty acid composition of the resulting glycerides is shown in table 5.
TABLE 5 glyceride fatty acids of the resulting products
Fatty acids Triglycerides Diglyceride Monoglyceride
C14:0 1.5 0.5 0.4
C16:0 11.3 9.1 8.8
C18:0 0.6 0.3 0.5
C18:1 0.4 0.2 0.1
C20:4 0.5 0.6 0.5
C20:5 0.3 0.5 0.4
C22:5 16.5 18.3 17.1
C22:6 67.1 69.2 71.3
Saturated fatty acid 13.4 9.9 9.7
Polyunsaturated fatty acids 84.4 88.6 89.3
Example 2
This example provides a method for preparing structural lipids that increase DHA bioavailability:
(1) adding algae oil rich in DHA from Crypthecodinium cohnii into a sealable batch reactor, adding ethanol according to the molar ratio of 1:6 (algae oil: ethanol), adding 8 wt% Lipozyme TL IM, sealing the reactor, and reacting at the temperature of 40 ℃ and the stirring speed of 500rpm for 8 hours to obtain an intermediate product rich in DHA.
(2) Firstly, removing ethanol from the intermediate product by adopting a rotary evaporation mode, wherein the conditions of the rotary evaporation are as follows: the temperature is 50 ℃, the rotating speed is 60rpm, the time is 20min, and the vacuum degree is 0.095 MPa.
(3) Further removing the generated fatty acid ethyl ester from the ethanol-removed intermediate product by molecular distillation, wherein the conditions of the molecular distillation are as follows: the evaporation temperature is 185 ℃; heat exchanger temperature, 60 ℃; rotational speed, 120 rpm; absolute pressure, 2 Pa. The glyceride and fatty acid compositions of the intermediate products obtained are shown in table 6.
TABLE 6 chemical composition of intermediate enzymatic products
Fatty acids Algae oil Intermediate product Glyceride composition Content (wt.)
C14:0 15.5 5.7 Triglycerides 64.3
C16:0 17.1 8.7 Diglyceride 30.6
C18:0 12.1 4.5 Monoglyceride 5.1
C18:1 0.97 0.1
C22:5 0.2 0.5
C22:6 52.3 79.9
Saturated fatty acid 44.7 18.9
Polyunsaturated fatty acids 52.5 80.4
(4) Taking the obtained DHA-enriched intermediate product as an initial material, adding 1:1.5 of glycerol in a molar ratio (intermediate enzymolysis product: glycerol) into a batch reactor, and reacting for 8 hours under the conditions that the addition amount of Novozym 435 is 10 wt%, the temperature is 60 ℃, and the stirring speed is 800rpm, so as to obtain a DHA-enriched diglyceride product. The glyceride composition of the product obtained is shown in Table 7.
TABLE 7 composition of diglyceride product
Glyceride composition Content (wt.)
Triglycerides 18.1
Diglyceride 60.5
Monoglyceride 21.4
The fatty acid composition of the resulting glycerides is shown in table 8.
TABLE 8 fatty acid composition of the resulting product glycerides
Fatty acids Triglycerides Diglyceride Monoglyceride
C14:0 6.4 4.4 3.7
C16:0 9.6 7.8 7.3
C18:0 5.2 4.1 3.5
C18:1 0.1 0.1 0.1
C22:5 0.4 0.6 0.7
C22:6 77.5 81.5 82.8
Saturated fatty acid 21.2 16.3 14.5
Polyunsaturated fatty acids 77.9 82.1 83.5
(5) Taking the obtained DHA-enriched intermediate product as an initial material, adding glycerol into a batch reactor according to the molar ratio of 1:6 (intermediate enzymolysis product: glycerol), and reacting for 5 hours under the conditions that the addition amount of Novozym 435 is 5 wt%, the temperature is 70 ℃, and the stirring speed is 800rpm, so as to obtain a DHA-enriched monoglyceride product. The glyceride composition of the product obtained is shown in Table 9.
TABLE 9 composition of monoglyceride products
Glyceride composition Content (wt.)
Triglycerides 6.4
Diglyceride 18.3
Monoglyceride 75.3
The fatty acid composition of the resulting glycerides is shown in table 10.
TABLE 10 fatty acid composition of the resulting product glycerides
Fatty acids Triglycerides Diglyceride Monoglyceride
C14:0 6.7 3.6 3.2
C16:0 9.9 7.7 7.1
C18:0 5.5 4.4 3.3
C18:1 0.1 0.1 0.1
C22:5 0.3 0.5 0.8
C22:6 76.5 81.7 84.3
Saturated fatty acid 22.1 15.7 13.6
Polyunsaturated fatty acids 76.8 82.2 85.1
Example 3
This example provides a method for preparing structural lipids that increase DHA bioavailability:
(1) adding DHA-enriched algae oil from Ulkenia sp into a sealable batch reactor, adding ethanol according to the molar ratio of 1:3 (algae oil: ethanol), adding 12 wt% of Lipase AY-30SD, sealing the reactor, and reacting at 30 ℃ and the stirring speed of 600rpm for 4 hours to obtain an intermediate enzymolysis product enriched in DHA.
(2) The intermediate enzymolysis product is firstly subjected to ethanol removal by adopting a rotary evaporation mode, and the conditions of the rotary evaporation are as follows: the temperature is 50 ℃, the rotating speed is 60r/min, the time is 20min, and the vacuum degree is 0.095 MPa.
(3) Further removing the generated fatty acid ethyl ester from the ethanol-removed intermediate product by molecular distillation, wherein the conditions of the molecular distillation are as follows: the evaporation temperature is 185 ℃; heat exchanger temperature, 60 ℃; rotational speed, 120 rpm; absolute pressure, 2 Pa. The glyceride and fatty acid compositions of the intermediate products obtained are shown in Table 11.
TABLE 11 chemical composition of intermediate enzymatic products
Figure BDA0002418540610000091
(4) Taking the obtained DHA-enriched intermediate product as an initial material, adding glycerol into a batch reactor according to a molar ratio (intermediate enzymolysis product: glycerol) of 1:0.5, and reacting for 5 hours under the conditions that the addition amount of Novozym 435 is 15 wt%, the temperature is 40 ℃, and the stirring speed is 400rpm, so as to obtain a DHA-enriched diglyceride product. The glyceride composition of the resulting product is shown in fig. 12.
TABLE 12 composition of diglyceride product
Figure BDA0002418540610000092
Figure BDA0002418540610000101
The fatty acid composition of the resulting glycerides is shown in table 13.
TABLE 13 fatty acid composition of the resulting product glycerides
Fatty acids Triglycerides Diglyceride Monoglyceride
C14:0 1.4 0.7 0.6
C16:0 9.7 7.4 7.1
C18:0 0.8 0.4 0.4
C20:4 0.5 0.7 0.8
C20:5 0.6 1.1 0.9
C22:5 18.8 20.1 20.8
C22:6 67.3 68.8 69.1
Saturated fatty acid 11.9 8.5 8.1
Polyunsaturated fatty acids 87.2 90.7 91.6
(5) Taking the obtained DHA-enriched intermediate product as an initial material, adding glycerol into a batch reactor according to a molar ratio (intermediate enzymolysis product: glycerol) of 1:9, and reacting for 8 hours under the conditions that the addition amount of Novozym 435 is 8 wt%, the temperature is 60 ℃, and the stirring speed is 400rpm, so as to obtain a DHA-enriched monoglyceride product. The glyceride composition of the product obtained is shown in Table 14.
TABLE 14 compositions of monoglyceride products
Glyceride composition Content (wt.)
Triglycerides 7.3
Diglyceride 20.6
Monoglyceride 72.1
The fatty acid composition of the resulting glycerides is shown in table 15.
TABLE 15 fatty acid composition of the resulting product glycerides
Figure BDA0002418540610000102
Figure BDA0002418540610000111
Example 4
This example provides a control example of a method for preparing structural lipids that increase DHA bioavailability:
(1) adding DHA-rich algal oil from Ulkenia sp, adding ethanol according to the molar ratio of 1:1 (algal oil: ethanol), adding 4 wt% Lipozyme TL IM, sealing the reactor, and reacting at the temperature of 30 ℃ and the stirring speed of 600rpm for 6 hours to obtain an intermediate product rich in DHA, wherein the DHA content is 57.5%, the polyunsaturated fatty acid content is 69.8%, the monoglyceride content is 1.1%, and the diglyceride content is 16.4%.
(2) Firstly, removing ethanol from the intermediate product by adopting a rotary evaporation mode, wherein the conditions of the rotary evaporation are as follows: the temperature is 50 ℃, the rotating speed is 60rpm, the time is 20min, and the vacuum degree is 0.095 MPa.
(3) Further removing the generated fatty acid ethyl ester from the ethanol-removed intermediate product by molecular distillation, wherein the conditions of the molecular distillation are as follows: the evaporation temperature is 185 ℃; heat exchanger temperature, 60 ℃; rotational speed, 120 rpm; absolute pressure, 2 Pa.
(4) Taking the obtained DHA-enriched intermediate product as an initial material, adding 1:0.5 of glycerol in a molar ratio (intermediate enzymolysis product: glycerol) into a batch reactor, and reacting for 6 hours under the conditions that the addition amount of Novozym 435 is 20 wt%, the temperature is 70 ℃, and the stirring speed is 800rpm, so as to obtain a DHA-enriched diglyceride product, wherein the diglyceride content is 33.4%, the DHA content in the diglyceride is 56.3%, and the polyunsaturated fatty acid content is 68.7%.
(5) Taking the obtained DHA-enriched intermediate product as an initiator, adding glycerol into a batch reactor according to the molar ratio of 1:10 (intermediate enzymolysis product: glycerol), and reacting for 4 hours under the conditions that the addition amount of Novozym 435 is 8 wt%, the temperature is 50 ℃, and the stirring speed is 800rpm, so as to obtain a DHA-enriched monoglyceride product, wherein the monoglyceride content is 52.4%, the DHA content in the monoglyceride is 58.6%, and the polyunsaturated fatty acid content is 71.3%.
Example 5
This example provides a control example of a method for preparing structural lipids that improve DHA bioavailability:
(1) adding DHA-rich algae oil from Crypthecodinium cohnii into a sealable batch reactor, adding ethanol according to the molar ratio of 1:8 (algae oil: ethanol), adding 10 wt% Lipozyme TL IM, sealing the reactor, and reacting at the temperature of 40 ℃ and the stirring speed of 500rpm for 10 hours to obtain an intermediate DHA-rich enzymolysis product, wherein the DHA content in the intermediate enzymolysis product is 82.3%, the polyunsaturated fatty acid content is 81.2%, the diglyceride content is 32.1%, and the monoglyceride content is 6.3%.
(2) Firstly, removing ethanol from the intermediate product by adopting a rotary evaporation mode, wherein the conditions of the rotary evaporation are as follows: the temperature is 50 ℃, the rotating speed is 60rpm, the time is 20min, and the vacuum degree is 0.095 MPa.
(3) Further removing the generated fatty acid ethyl ester from the ethanol-removed intermediate product by molecular distillation, wherein the conditions of the molecular distillation are as follows: the evaporation temperature is 185 ℃; heat exchanger temperature, 60 ℃; rotational speed, 120 rpm; absolute pressure, 2 Pa.
(4) The obtained DHA-enriched intermediate product is used as an initiator, 1:0.3 of glycerol is added into a batch reactor according to the molar ratio (intermediate enzymolysis product: glycerol), and the mixture reacts for 8 hours under the conditions that the addition amount of Novozym 435 is 8 wt%, the temperature is 50 ℃, and the stirring speed is 800rpm, so that the content of diglyceride in the product is 46.7 percent.
(5) The obtained DHA-enriched intermediate product is used as an initiator, glycerol is added into a batch reactor according to the mol ratio of 1:2.5 (intermediate enzymolysis product: glycerol), and the mixture reacts for 5 hours under the conditions that the addition amount of Novozym 435 is 5 wt%, the temperature is 70 ℃, and the stirring speed is 800rpm, so that the content of monoglyceride in the obtained product is 54.3 percent.
In the above examples 1-3, a certain amount of glyceride products are generated by the alcoholysis reaction in the first step, and in the second step, because of the existence of monoglyceride and diglyceride, the synergistic emulsification can be achieved in the reaction process, and after the glycerol is added and stirred, the reaction system is changed into a microemulsion system, so that the contact area between the glycerol and the oil is increased, and the specific surface area of the lipase in play is increased.
Therefore, the method replaces the conventional fatty acid in the microbial oil containing DHA by utilizing the selectivity of lipase to the substrate through alcoholysis reaction, and enriches the DHA in the intermediate enzymolysis product; the method realizes that the content of monoglyceride in the DHA-rich monoglyceride product is more than 60%, the content of diglyceride in the DHA-rich diglyceride product is more than 50%, the content of DHA in the obtained product is more than 65%, the content of polyunsaturated fatty acids is more than 80%, and the content of saturated fatty acids is less than 20% through the synergistic effect of all processes and preferably all process conditions.
The method adopts the two-step enzyme method for catalytic preparation to obtain DHA-enriched monoglyceride or diglyceride products for the first time, the DHA and polyunsaturated fatty acid content in the products exceeds similar reports, the process is simple, the operation is easy, and the method is suitable for industrial production. Meanwhile, the product can play a role in predigestion, is beneficial to the digestion and absorption of DHA, and improves the bioavailability of DHA.
It should be noted that the above-mentioned embodiments are only for illustrating the technical solutions of the present invention and not for limiting, and although the present invention has been described in detail with reference to the preferred embodiments, it should be understood by those skilled in the art that modifications or equivalent substitutions may be made on the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention, which should be covered by the claims of the present invention.

Claims (10)

1. A method for preparing structural lipid for improving DHA bioavailability is characterized by comprising the following steps: comprises the steps of (a) preparing a mixture of a plurality of raw materials,
enzyme catalysis alcoholysis reaction: adding microbial oil containing DHA and ethanol into an intermittent reactor according to a molar ratio of 1: 2-10, adding 3-12 wt% of lipase, and reacting for 3-12 hours at a temperature of 30-50 ℃ and a stirring speed of 300-800 rpm to prepare an intermediate enzymolysis product rich in DHA, wherein the content of monoglyceride and diglyceride in the intermediate enzymolysis product is more than 30%, and the content of monoglyceride is more than 3%;
preparing monoglyceride: after ethanol and fatty acid ethyl ester of the intermediate enzymolysis product are respectively removed by adopting reduced pressure distillation and molecular distillation, adding the intermediate enzymolysis product and glycerol into an intermittent reactor according to the molar ratio of 1: 3-10, adding 3-12 wt% of lipase, and reacting for 3-12 hours at the temperature of 40-70 ℃ and at the stirring speed of 300-800 rpm to obtain a monoglyceride product rich in DHA, wherein the monoglyceride content is more than 60%, the DHA content is more than 65%, the polyunsaturated fatty acid content is more than 80%, the saturated fatty acid content is less than 20%, and the DHA content in the monoglyceride is more than 65%;
preparing diglyceride: after ethanol and fatty acid ethyl ester of the intermediate enzymolysis product are respectively removed by adopting reduced pressure distillation and molecular distillation, adding the intermediate enzymolysis product and glycerol into an intermittent reactor according to the molar ratio of 1: 0.5-2, adding 3-15 wt% of lipase, and reacting for 4-10 hours at the temperature of 40-70 ℃ and at the stirring speed of 300-800 rpm to obtain a diglyceride product rich in DHA, wherein the content of diglyceride is more than 50%, the content of DHA is more than 65%, the content of polyunsaturated fatty acid is more than 80%, the content of saturated fatty acid is less than 20%, and the content of DHA in diglyceride is more than 65%.
2. The method of making a structural lipid for improving DHA bioavailability according to claim 1, wherein: the enzyme catalyzes alcoholysis reaction, wherein the used Lipase comprises Lipozyme RMIM, Lipozyme TLIM or Lipase AY-30 SD.
3. The method of making a structural lipid for improving DHA bioavailability according to claim 1, wherein: the preparation of monoglyceride, wherein the lipase used is Novozym 435.
4. The method of making a structural lipid for improving DHA bioavailability according to claim 1, wherein: the preparation of diglyceride, wherein the lipase used is Novozym 435.
5. The method of making a structural lipid for improving DHA bioavailability according to claim 1, wherein: the DHA-containing microbial oil is one or more of microbial oils produced by fermenting microalgae such as Crypthecodinium cohnii, Schizochytrium sp.
6. The method of making a structural lipid for improving DHA bioavailability according to claim 1, wherein: the glycerol is food grade glycerol.
7. The method of making a structural lipid for improving DHA bioavailability according to claim 1, wherein: the enzymatic alcoholysis reaction is carried out, wherein the DHA content of the intermediate enzymolysis product is more than 65%, the polyunsaturated fatty acid content is more than 80%, the saturated fatty acid content is less than 20%, the content of monoglyceride and diglyceride is more than 30%, and the content of monoglyceride is more than 3%.
8. The method of making a structural lipid for improving DHA bioavailability according to claim 1, wherein: the DHA-rich monoglyceride product is obtained, wherein the content of monoglyceride in the product is more than 60%, the content of DHA is more than 65%, the content of polyunsaturated fatty acid is more than 80%, the content of saturated fatty acid is less than 20%, and the content of DHA in monoglyceride is more than 65%.
9. The method of making a structural lipid for improving DHA bioavailability according to claim 1, wherein: the DHA-rich diglyceride product is obtained, wherein the content of diglyceride in the product is more than 50%, the content of DHA in the product is more than 65%, the content of polyunsaturated fatty acid is more than 80%, the content of saturated fatty acid is less than 20%, and the content of DHA in diglyceride is more than 65%.
10. The product of the method for preparing structured lipid for improving bioavailability of DHA according to any one of claims 1 to 9, wherein: the product comprises a DHA-rich monoglyceride product and a DHA-rich diglyceride product, wherein the content of monoglyceride in the DHA-rich monoglyceride product is more than 60%, the content of diglyceride in the DHA-rich diglyceride product is more than 50%, and the content of DHA in both the monoglyceride and the diglyceride is more than 65%.
CN202010198628.2A 2020-03-20 2020-03-20 Preparation method of structural lipid for improving DHA bioavailability and product thereof Active CN111363766B (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN202010198628.2A CN111363766B (en) 2020-03-20 2020-03-20 Preparation method of structural lipid for improving DHA bioavailability and product thereof

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN202010198628.2A CN111363766B (en) 2020-03-20 2020-03-20 Preparation method of structural lipid for improving DHA bioavailability and product thereof

Publications (2)

Publication Number Publication Date
CN111363766A true CN111363766A (en) 2020-07-03
CN111363766B CN111363766B (en) 2021-08-10

Family

ID=71204661

Family Applications (1)

Application Number Title Priority Date Filing Date
CN202010198628.2A Active CN111363766B (en) 2020-03-20 2020-03-20 Preparation method of structural lipid for improving DHA bioavailability and product thereof

Country Status (1)

Country Link
CN (1) CN111363766B (en)

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN113584093A (en) * 2021-07-30 2021-11-02 江南大学 Preparation method of structured lipid with high DHA content and product thereof
CN115011642A (en) * 2022-05-24 2022-09-06 浙江工商大学 Method for preparing structured lipid rich in EPA and DHA by using tuna

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103352059A (en) * 2013-06-25 2013-10-16 江南大学 Preparation method of diglyceride
CN103880672A (en) * 2014-03-20 2014-06-25 江苏中邦制药有限公司 High-purity DHA algae oil ethyl ester and preparation method for transferring high-purity DHA algae oil ethyl ester into glyceride
CN105821088A (en) * 2016-05-26 2016-08-03 暨南大学 Method for preparing glyceride rich in EPA and DHA through enzyme catalysis
CN107549332A (en) * 2017-09-11 2018-01-09 江南大学 Strengthen method of baby's long-chain polyunsaturated fatty acid bioavailability and products thereof

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103352059A (en) * 2013-06-25 2013-10-16 江南大学 Preparation method of diglyceride
CN103880672A (en) * 2014-03-20 2014-06-25 江苏中邦制药有限公司 High-purity DHA algae oil ethyl ester and preparation method for transferring high-purity DHA algae oil ethyl ester into glyceride
CN105821088A (en) * 2016-05-26 2016-08-03 暨南大学 Method for preparing glyceride rich in EPA and DHA through enzyme catalysis
CN107549332A (en) * 2017-09-11 2018-01-09 江南大学 Strengthen method of baby's long-chain polyunsaturated fatty acid bioavailability and products thereof

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
YUKIHISA TANAKA等: "Concentration of Docosahexaenoic Acid in Glyceride by Hydrolysis of Fish Oil with Candida cylindracea Lipase", 《JAOCS》 *

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN113584093A (en) * 2021-07-30 2021-11-02 江南大学 Preparation method of structured lipid with high DHA content and product thereof
CN115011642A (en) * 2022-05-24 2022-09-06 浙江工商大学 Method for preparing structured lipid rich in EPA and DHA by using tuna

Also Published As

Publication number Publication date
CN111363766B (en) 2021-08-10

Similar Documents

Publication Publication Date Title
US7767427B2 (en) Production method of oil or fat containing polyunsaturated fatty acid-containing triglyceride
CN111172211A (en) Method for preparing long-chain polyunsaturated fatty acid glyceride rich in fish oil n-3 by enzyme method and product thereof
EP1966387B1 (en) Process for the production of diacylglycerol
Gupta et al. Lipase mediated upgradation of dietary fats and oils
JP4761771B2 (en) Method for producing microbial fats and oils with reduced unsaponifiable matter content and said fats and oils
JP5828612B2 (en) Method for concentrating fatty acid alkyl esters by enzymatic reaction using glycerol
US7998712B2 (en) Process for production of transesterified oils/fats or triglycerides
WO2016176987A1 (en) Preparation method for structured lipid rich in 1,3-dioleate-2-palmitic acid triglyceride
CN111172210A (en) Method for preparing arachidonic acid glyceride rich by two-step enzyme method and product thereof
CN111378701A (en) Preparation method of middle-long carbon chain triglyceride containing polyunsaturated fatty acid and product thereof
CN111363766B (en) Preparation method of structural lipid for improving DHA bioavailability and product thereof
CN109666709B (en) Method for preparing diglyceride by using high-acid-value grease as raw material
JP4212590B2 (en) High purity diglyceride oil composition containing conjugated linoleic acid and method for producing the same
WO2003094625A1 (en) A facile two-step enzyme process for increasing the content of polyunsaturated fatty acids in fish oil
CN113832200B (en) Preparation method of breast milk structured fat
US11396667B2 (en) Enzymatic method for preparation of lecithin polyunsaturated fatty acids (PUFAs)
Shimada et al. Production of functional lipids containing polyunsaturated fatty acids with lipase
JP4310387B2 (en) Omega-3 highly unsaturated fatty acid-containing partial glyceride composition and method for producing the same
CN117089402A (en) Method for reducing glycidol content in enzymatic diglyceride
Shimada 20Enzymatic Modification

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination
GR01 Patent grant
GR01 Patent grant