CN118813729B - A preparation method and application of diglyceride - Google Patents
A preparation method and application of diglyceride Download PDFInfo
- Publication number
- CN118813729B CN118813729B CN202411312663.7A CN202411312663A CN118813729B CN 118813729 B CN118813729 B CN 118813729B CN 202411312663 A CN202411312663 A CN 202411312663A CN 118813729 B CN118813729 B CN 118813729B
- Authority
- CN
- China
- Prior art keywords
- oil
- lipase
- seed oil
- seed
- hydrolysate
- 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.)
- Active
Links
- 238000002360 preparation method Methods 0.000 title claims abstract description 20
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 claims abstract description 50
- 108090001060 Lipase Proteins 0.000 claims abstract description 39
- 239000004367 Lipase Substances 0.000 claims abstract description 39
- 102000004882 Lipase Human genes 0.000 claims abstract description 39
- 235000019421 lipase Nutrition 0.000 claims abstract description 39
- 239000002904 solvent Substances 0.000 claims abstract description 18
- 238000002156 mixing Methods 0.000 claims abstract description 17
- 239000000413 hydrolysate Substances 0.000 claims abstract description 15
- 238000000034 method Methods 0.000 claims abstract description 15
- 108010031797 Candida antarctica lipase B Proteins 0.000 claims abstract description 11
- 239000004519 grease Substances 0.000 claims abstract description 11
- 238000005886 esterification reaction Methods 0.000 claims abstract description 7
- 239000003921 oil Substances 0.000 claims description 39
- 235000019198 oils Nutrition 0.000 claims description 39
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 17
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 16
- 235000015112 vegetable and seed oil Nutrition 0.000 claims description 15
- 235000012424 soybean oil Nutrition 0.000 claims description 13
- 239000003549 soybean oil Substances 0.000 claims description 13
- IIZPXYDJLKNOIY-JXPKJXOSSA-N 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCC\C=C/C\C=C/C\C=C/C\C=C/CCCCC IIZPXYDJLKNOIY-JXPKJXOSSA-N 0.000 claims description 8
- 239000000787 lecithin Substances 0.000 claims description 8
- 235000010445 lecithin Nutrition 0.000 claims description 8
- 229940067606 lecithin Drugs 0.000 claims description 8
- 239000011259 mixed solution Substances 0.000 claims description 8
- 238000006460 hydrolysis reaction Methods 0.000 claims description 6
- 239000000047 product Substances 0.000 claims description 5
- 239000008158 vegetable oil Substances 0.000 claims description 5
- 108010048733 Lipozyme Proteins 0.000 claims description 4
- 238000006243 chemical reaction Methods 0.000 claims description 4
- 235000005687 corn oil Nutrition 0.000 claims description 4
- 239000002285 corn oil Substances 0.000 claims description 4
- 230000007062 hydrolysis Effects 0.000 claims description 4
- 235000007650 Aralia spinosa Nutrition 0.000 claims description 3
- 235000019483 Peanut oil Nutrition 0.000 claims description 3
- 241000949456 Zanthoxylum Species 0.000 claims description 3
- 235000013399 edible fruits Nutrition 0.000 claims description 3
- 239000008157 edible vegetable oil Substances 0.000 claims description 3
- 239000000312 peanut oil Substances 0.000 claims description 3
- 230000035484 reaction time Effects 0.000 claims description 3
- 241000219226 Acer truncatum Species 0.000 claims description 2
- 235000019489 Almond oil Nutrition 0.000 claims description 2
- 240000001548 Camellia japonica Species 0.000 claims description 2
- 241000195493 Cryptophyta Species 0.000 claims description 2
- 240000000950 Hippophae rhamnoides Species 0.000 claims description 2
- 235000003145 Hippophae rhamnoides Nutrition 0.000 claims description 2
- 240000007817 Olea europaea Species 0.000 claims description 2
- 240000007594 Oryza sativa Species 0.000 claims description 2
- 235000007164 Oryza sativa Nutrition 0.000 claims description 2
- 235000006484 Paeonia officinalis Nutrition 0.000 claims description 2
- 244000170916 Paeonia officinalis Species 0.000 claims description 2
- 235000019484 Rapeseed oil Nutrition 0.000 claims description 2
- 235000019774 Rice Bran oil Nutrition 0.000 claims description 2
- 235000019498 Walnut oil Nutrition 0.000 claims description 2
- 244000248162 Xanthoceras sorbifolium Species 0.000 claims description 2
- 235000009240 Xanthoceras sorbifolium Nutrition 0.000 claims description 2
- 239000008168 almond oil Substances 0.000 claims description 2
- 239000010775 animal oil Substances 0.000 claims description 2
- 239000010495 camellia oil Substances 0.000 claims description 2
- 239000003240 coconut oil Substances 0.000 claims description 2
- 235000019864 coconut oil Nutrition 0.000 claims description 2
- 235000018597 common camellia Nutrition 0.000 claims description 2
- 239000002385 cottonseed oil Substances 0.000 claims description 2
- 235000012343 cottonseed oil Nutrition 0.000 claims description 2
- 239000008169 grapeseed oil Substances 0.000 claims description 2
- 239000010460 hemp oil Substances 0.000 claims description 2
- 239000000944 linseed oil Substances 0.000 claims description 2
- 235000021388 linseed oil Nutrition 0.000 claims description 2
- 239000003346 palm kernel oil Substances 0.000 claims description 2
- 235000019865 palm kernel oil Nutrition 0.000 claims description 2
- 239000001335 perilla frutescens leaf extract Substances 0.000 claims description 2
- 239000008171 pumpkin seed oil Substances 0.000 claims description 2
- 235000009566 rice Nutrition 0.000 claims description 2
- 239000008165 rice bran oil Substances 0.000 claims description 2
- 239000003813 safflower oil Substances 0.000 claims description 2
- 239000008159 sesame oil Substances 0.000 claims description 2
- 235000011803 sesame oil Nutrition 0.000 claims description 2
- 235000020238 sunflower seed Nutrition 0.000 claims description 2
- UFTFJSFQGQCHQW-UHFFFAOYSA-N triformin Chemical compound O=COCC(OC=O)COC=O UFTFJSFQGQCHQW-UHFFFAOYSA-N 0.000 claims description 2
- 239000008170 walnut oil Substances 0.000 claims description 2
- 239000010497 wheat germ oil Substances 0.000 claims description 2
- 235000002566 Capsicum Nutrition 0.000 claims 1
- 240000008574 Capsicum frutescens Species 0.000 claims 1
- 239000001390 capsicum minimum Substances 0.000 claims 1
- 150000004668 long chain fatty acids Chemical class 0.000 claims 1
- 235000010746 mayonnaise Nutrition 0.000 claims 1
- 239000008268 mayonnaise Substances 0.000 claims 1
- 150000004667 medium chain fatty acids Chemical class 0.000 claims 1
- 230000003301 hydrolyzing effect Effects 0.000 abstract description 11
- 108010009736 Protein Hydrolysates Proteins 0.000 abstract description 9
- 238000013329 compounding Methods 0.000 abstract description 4
- 230000009286 beneficial effect Effects 0.000 abstract description 3
- 235000013305 food Nutrition 0.000 abstract description 2
- 210000000748 cardiovascular system Anatomy 0.000 abstract 1
- 238000000105 evaporative light scattering detection Methods 0.000 description 14
- 235000011187 glycerol Nutrition 0.000 description 14
- 239000002994 raw material Substances 0.000 description 10
- FPWNQPQTICPCOM-UHFFFAOYSA-N acetonitrile;propan-2-ol Chemical group CC#N.CC(C)O FPWNQPQTICPCOM-UHFFFAOYSA-N 0.000 description 7
- 238000004128 high performance liquid chromatography Methods 0.000 description 7
- 238000010829 isocratic elution Methods 0.000 description 7
- 238000001474 liquid chromatography-evaporative light scattering detection Methods 0.000 description 7
- 239000003643 water by type Substances 0.000 description 7
- 235000019197 fats Nutrition 0.000 description 5
- 108090000790 Enzymes Proteins 0.000 description 3
- 102000004190 Enzymes Human genes 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 230000002255 enzymatic effect Effects 0.000 description 3
- 238000004519 manufacturing process Methods 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 230000002051 biphasic effect Effects 0.000 description 2
- 230000004060 metabolic process Effects 0.000 description 2
- 239000004006 olive oil Substances 0.000 description 2
- 235000008390 olive oil Nutrition 0.000 description 2
- 239000000758 substrate Substances 0.000 description 2
- 150000003626 triacylglycerols Chemical class 0.000 description 2
- VZSRBBMJRBPUNF-UHFFFAOYSA-N 2-(2,3-dihydro-1H-inden-2-ylamino)-N-[3-oxo-3-(2,4,6,7-tetrahydrotriazolo[4,5-c]pyridin-5-yl)propyl]pyrimidine-5-carboxamide Chemical compound C1C(CC2=CC=CC=C12)NC1=NC=C(C=N1)C(=O)NCCC(N1CC2=C(CC1)NN=N2)=O VZSRBBMJRBPUNF-UHFFFAOYSA-N 0.000 description 1
- 102100021851 Calbindin Human genes 0.000 description 1
- 240000004160 Capsicum annuum Species 0.000 description 1
- 101000898082 Homo sapiens Calbindin Proteins 0.000 description 1
- 235000019482 Palm oil Nutrition 0.000 description 1
- 244000025272 Persea americana Species 0.000 description 1
- 235000008673 Persea americana Nutrition 0.000 description 1
- 101001021643 Pseudozyma antarctica Lipase B Proteins 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 239000002671 adjuvant Substances 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000003125 aqueous solvent Substances 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 230000003197 catalytic effect Effects 0.000 description 1
- 238000006555 catalytic reaction Methods 0.000 description 1
- 239000002131 composite material Substances 0.000 description 1
- 239000002537 cosmetic Substances 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 230000000214 effect on organisms Effects 0.000 description 1
- 239000003995 emulsifying agent Substances 0.000 description 1
- 230000032050 esterification Effects 0.000 description 1
- 150000004665 fatty acids Chemical group 0.000 description 1
- 235000021588 free fatty acids Nutrition 0.000 description 1
- 238000000703 high-speed centrifugation Methods 0.000 description 1
- 238000000265 homogenisation Methods 0.000 description 1
- FCCDDURTIIUXBY-UHFFFAOYSA-N lipoamide Chemical compound NC(=O)CCCCC1CCSS1 FCCDDURTIIUXBY-UHFFFAOYSA-N 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 239000002540 palm oil Substances 0.000 description 1
- 230000004584 weight gain Effects 0.000 description 1
- 235000019786 weight gain Nutrition 0.000 description 1
- 239000013585 weight reducing agent Substances 0.000 description 1
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
- C12P7/6436—Fatty acid esters
- C12P7/6445—Glycerides
- C12P7/6454—Glycerides by esterification
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Zoology (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Wood Science & Technology (AREA)
- Biotechnology (AREA)
- Microbiology (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Oil, Petroleum & Natural Gas (AREA)
- Health & Medical Sciences (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)
Abstract
The invention relates to the technical field of foods, in particular to a preparation method and application of diglycerides. The method comprises the steps of firstly, hydrolyzing grease partially by adding a solvent under the action of a first lipase to obtain a hydrolysate, then, mixing the hydrolysate with glycerol, and carrying out esterification reaction by adopting a second lipase, wherein the first lipase is prepared by compounding lipase RM and candida antarctica lipase B. The invention can specifically synthesize the 1, 3-diglyceride which is beneficial to the cardiovascular system of the organism, has high yield and good application prospect.
Description
Technical Field
The invention relates to the technical field of preparation processes, in particular to a preparation method and application of diglycerides.
Background
Diglycerides are a type of fatty molecules having only two fatty acid chains, and are useful as emulsifiers, fatty plasticity improvers, or as substrates for foods, pharmaceuticals, cosmetics, etc.
In recent years, it has been found that diglycerides are more easily converted into energy and utilized in the body, and triglycerides are more easily accumulated in the body, because of different absorption and metabolism patterns of diglycerides and triglycerides. The edible diglyceride-containing fat has an effect of suppressing weight gain, and therefore, the diglyceride fat can be eaten as a healthy fat.
The diglyceride can be produced by various processes, a corresponding production method is disclosed in a plurality of patents in the related field, such as China patent application CN112513235A, CN116042736A, CN105400837A and the like, wherein CN105400837A discloses a preparation method of enzyme-catalyzed diglyceride, and the preparation method comprises the following steps of (1) partially hydrolyzing grease, namely, partially hydrolyzing edible oil by adopting an enzyme-catalyzed hydrolysis method, dehydrating to obtain a hydrolysate, wherein the content of the diglyceride in the hydrolysate is 30.0-35.0 wt% and the content of free fatty acid is 26.0-30.0 wt%, and (2) re-esterifying, namely, adding glycerol with the mass of 2.5-4 times of the glycerol into the hydrolysate obtained in the step (1), and esterifying by adopting an enzyme-catalyzed esterification method to synthesize the diglyceride, thereby removing the superfluous diglyceride and obtaining the edible diglyceride.
The Chinese patent application CN116042736A discloses an enzymatic production process of diglyceride, which adopts enzymatic glycerolysis to prepare diglyceride oil, namely, grease and glycerin react under the catalysis of lipase to prepare diglyceride, and the reaction substrate is subjected to homogenization treatment before the enzymatic glycerolysis, so that the catalytic efficiency of the lipase in a reaction system can be improved, the purposes of reducing the reaction time, reducing the enzyme addition amount and improving the content of the diglyceride in the product are achieved, and the purpose of reducing the production cost is also achieved.
In the prior art, a certain research and study is carried out on the preparation of diglyceride to obtain a certain effect, but a plurality of processes are difficult to be applied in a real large scale, the product yield is required to be further improved, in addition, the research shows that the 1,3-DG has good regulation effects on organisms, such as blood fat reduction, weight reduction and the like, but most of the obtained products are mixtures of 1,2-, 1,3-DG and the ratio of the mixture to the mixture is usually 7:3-6:4, so the research on a preparation method with higher 1,3-DG content is a problem to be solved in the art.
Disclosure of Invention
Aiming at the defects existing in the prior art, the invention provides a preparation method of diglyceride, which solves the problems of low 1,3-DG content and low yield.
The invention aims at realizing the following technical scheme:
A process for the preparation of diglycerides comprising the steps of:
(1) Partially hydrolyzing the grease by adding a solvent under the action of first lipase to obtain a hydrolysate;
(2) Mixing the hydrolysate with glycerol, and performing esterification reaction by using a second lipase;
wherein the first lipase is compounded by lipase Lipozyme RM and candida antarctica lipase B in a mass ratio of 1:1-3.
Preferably, the solvent is a mixed solution of lecithin, ethanol and water in a mass ratio of 0.01-0.05:0.1-0.2:1.
Preferably, the temperature of the hydrolysis is 40-65 ℃ and the time of the hydrolysis is 3-8h.
Preferably, the amount of the first lipase is 1-10% of the mass of the fat.
Preferably, the solvent is used in an amount of 10-50% by mass of the grease.
Preferably, the second lipase is lipase 435.
Preferably, the mass ratio of the hydrolysate to the glycerol is 1:0.5-2.
Preferably, the temperature of the esterification reaction is 40-60 ℃ and the reaction time is 5-10h.
Preferably, the grease is selected from vegetable oils;
Preferably, the vegetable oil is selected from one or more of linseed oil, soybean oil, rapeseed oil, peanut oil, corn oil, sunflower seed oil, camellia seed oil, coconut oil, palm kernel oil, olive fruit residue oil, walnut oil, rice bran oil, rice oil, cotton seed oil, perilla seed oil, safflower seed oil, grape seed oil, tea seed oil, peony seed oil, sesame oil, wheat germ oil, acer truncatum seed oil, shinyleaf yellowhorn oil, sea buckthorn seed oil, DHA algae oil, medium chain triglyceride, medium-long chain fatty acid edible oil, hemp seed oil, pricklyash peel oil, pumpkin seed oil, chilli oil, almond oil and merry seed oil;
or, the grease is selected from animal oil.
Preferably, the vegetable oil is selected from corn oil, soybean oil, palm oil, peanut oil, olive oil or avocado.
The invention also provides application of the preparation method in preparation of 1,3 diglycerides.
Compared with the prior art, the invention has the beneficial effects that:
the invention is repeatedly researched and screened to obtain the optimal preparation process, which adopts the composite lipase to carry out hydrolysis reaction under a specific biphasic solvent, then glycerol is added to carry out esterification reaction under the action of second lipase, and finally oil phase is separated by high-speed centrifugation. The method has higher yield of diglyceride, and more importantly, can improve the ratio of 1, 3-diglyceride beneficial to metabolism of organisms.
Detailed Description
The invention is further described in connection with the following detailed description.
The following raw materials are all conventional in the art, wherein candida antarctica lipase B is available from hangzhou department biotechnology limited and is available in the model number Lipozyme CALB. The supplier of lipase RM is Norwesterner.
Example 1
The preparation method of the diglyceride comprises the following steps:
(1) Adding 5% of first lipase (which is prepared by compounding lipase RM and candida antarctica lipase B in a mass ratio of 1:2) into soybean oil, mixing, then adding 25% of solvent (a mixed solution of lecithin, ethanol and water in a mass ratio of 0.02:0.1:1) into the soybean oil, hydrolyzing for 5 hours at 55 ℃, and standing for layering to obtain an oil phase as a hydrolysate;
(2) Adding 1 times of glycerol into the hydrolysate, mixing, adding a second lipase Lipozyme435, esterifying at 50deg.C for 8 hr, centrifuging at 5500rad for 30min, separating oil from water, collecting oil phase, and measuring content by high performance liquid chromatography-evaporative light scattering detector (HPLC-ELSD). The high performance liquid chromatography conditions are that the chromatographic column is a Waters C18 chromatographic column (250 mm multiplied by 4.6 mm,5 μm), the mobile phase is acetonitrile-isopropanol with the volume ratio of 95:5, the isocratic elution is carried out, the flow rate is 1.0 mL/min, and the column temperature is 35 ℃. The conditions of the evaporative light scattering detector are that the temperature of the ELSD drift tube is 90 ℃, and the flow rate of N 2 is 2.2L/min.
Example 2
(1) Adding 2% of first lipase (compounded by lipase RM and candida antarctica lipase B in a mass ratio of 1:2) into corn oil, mixing, then adding 15% of solvent (mixed solution of lecithin, ethanol and water in a mass ratio of 0.05:0.2:1), partially hydrolyzing for 4 hours at 65 ℃, standing and layering to obtain an oil phase as a hydrolysate;
(2) Adding 0.5 times of glycerol into the hydrolysate, mixing, adding a second lipase Lipozyme435, esterifying at 40deg.C for 5 hr, centrifuging at 5500rad for 30min after the reaction, separating oil from water, collecting oil phase, and measuring content by high performance liquid chromatography-evaporative light scattering detector (HPLC-ELSD). The high performance liquid chromatography conditions are that the chromatographic column is a Waters C18 chromatographic column (250 mm multiplied by 4.6 mm,5 μm), the mobile phase is acetonitrile-isopropanol with the volume ratio of 95:5, the isocratic elution is carried out, the flow rate is 1.0 mL/min, and the column temperature is 35 ℃. The conditions of the evaporative light scattering detector are that the temperature of the ELSD drift tube is 90 ℃, and the flow rate of N 2 is 2.2L/min.
Example 3
(1) Adding 7% of first lipase (which is prepared by compounding lipase RM and candida antarctica lipase B in a mass ratio of 1:3) into olive oil, mixing, then adding 45% of solvent (a mixed solution of lecithin, ethanol and water in a mass ratio of 0.01:0.2:1), partially hydrolyzing for 8 hours at 40 ℃, standing and layering to obtain an oil phase as a hydrolysate;
(2) Adding 2 times of glycerol into the hydrolysate, mixing, adding a second lipase Lipozyme435, esterifying at 60deg.C for 10 hr, centrifuging at 5500rad for 30min, separating oil from water, collecting oil phase, and measuring content by high performance liquid chromatography-evaporative light scattering detector (HPLC-ELSD). The high performance liquid chromatography conditions are that the chromatographic column is a Waters C18 chromatographic column (250 mm multiplied by 4.6 mm,5 μm), the mobile phase is acetonitrile-isopropanol with the volume ratio of 95:5, the isocratic elution is carried out, the flow rate is 1.0 mL/min, and the column temperature is 35 ℃. The conditions of the evaporative light scattering detector are that the temperature of the ELSD drift tube is 90 ℃, and the flow rate of N 2 is 2.2L/min.
Example 1-1
The method comprises the following steps:
(1) Adding 5% of a first lipase RM into soybean oil, mixing, adding a solvent (mixed solution of lecithin, ethanol and water in a mass ratio of 0.02:0.1:1) accounting for 25% of the mass of the soybean oil, hydrolyzing for 5 hours at 55 ℃, standing and layering to obtain an oil phase as a hydrolysate;
(2) Adding 1 times of glycerol into the hydrolysate, mixing, adding a second lipase Lipozyme435, esterifying at 50deg.C for 8 hr, centrifuging at 5500rad for 30min, separating oil from water, collecting oil phase, and measuring content by high performance liquid chromatography-evaporative light scattering detector (HPLC-ELSD). The high performance liquid chromatography conditions are that the chromatographic column is a Waters C18 chromatographic column (250 mm multiplied by 4.6 mm,5 μm), the mobile phase is acetonitrile-isopropanol with the volume ratio of 95:5, the isocratic elution is carried out, the flow rate is 1.0 mL/min, and the column temperature is 35 ℃. The conditions of the evaporative light scattering detector are that the temperature of the ELSD drift tube is 90 ℃, and the flow rate of N 2 is 2.2L/min.
Examples 1 to 2
The method comprises the following steps:
(1) Adding 5% of first lipase candida antarctica lipase B into soybean oil, mixing, adding 25% of solvent (mixed solution of lecithin, ethanol and water in a mass ratio of 0.02:0.1:1) into the soybean oil, hydrolyzing for 5 hours at 55 ℃, standing and layering to obtain an oil phase as a hydrolysate;
(2) Adding 1 times of glycerol into the hydrolysate, mixing, adding a second lipase Lipozyme435, esterifying at 50deg.C for 8 hr, centrifuging at 5500rad for 30min, separating oil from water, collecting oil phase, and measuring content by high performance liquid chromatography-evaporative light scattering detector (HPLC-ELSD). The high performance liquid chromatography conditions are that the chromatographic column is a Waters C18 chromatographic column (250 mm multiplied by 4.6 mm,5 μm), the mobile phase is acetonitrile-isopropanol with the volume ratio of 95:5, the isocratic elution is carried out, the flow rate is 1.0 mL/min, and the column temperature is 35 ℃. The conditions of the evaporative light scattering detector are that the temperature of the ELSD drift tube is 90 ℃, and the flow rate of N 2 is 2.2L/min.
Examples 1 to 3
The solvent is different from example 1, and is specifically as follows:
(1) Adding 5% of first lipase (which is prepared by compounding lipase RM and candida antarctica lipase B in a mass ratio of 1:2) into soybean oil, mixing, adding 25% of solvent (mixed solution of ethanol and water in a mass ratio of 0.1:1) into the soybean oil, hydrolyzing for 5 hours at 55 ℃, standing and layering to obtain an oil phase as a hydrolysate;
(2) Adding 1 times of glycerol into the hydrolysate, mixing, adding a second lipase Lipozyme435, esterifying at 50deg.C for 8 hr, centrifuging at 5500rad for 30min, separating oil from water, collecting oil phase, and measuring content by high performance liquid chromatography-evaporative light scattering detector (HPLC-ELSD). The high performance liquid chromatography conditions are that the chromatographic column is a Waters C18 chromatographic column (250 mm multiplied by 4.6 mm,5 μm), the mobile phase is acetonitrile-isopropanol with the volume ratio of 95:5, the isocratic elution is carried out, the flow rate is 1.0 mL/min, and the column temperature is 35 ℃. The conditions of the evaporative light scattering detector are that the temperature of the ELSD drift tube is 90 ℃, and the flow rate of N 2 is 2.2L/min.
Examples 1 to 4
The solvent is water. The method comprises the following steps:
(1) Adding 5% of first lipase (which is compounded by lipase RM and candida antarctica lipase B in a mass ratio of 1:2) into soybean oil, mixing, adding 25% of water in the mass of the soybean oil, hydrolyzing for 5 hours at 55 ℃, standing and layering to obtain an oil phase as a hydrolysate;
(2) Adding 1 times of glycerol into the hydrolysate, mixing, adding a second lipase Lipozyme435, esterifying at 50deg.C for 8 hr, centrifuging at 5500rad for 30min, separating oil from water, collecting oil phase, and measuring content by high performance liquid chromatography-evaporative light scattering detector (HPLC-ELSD). The high performance liquid chromatography conditions are that the chromatographic column is a Waters C18 chromatographic column (250 mm multiplied by 4.6 mm,5 μm), the mobile phase is acetonitrile-isopropanol with the volume ratio of 95:5, the isocratic elution is carried out, the flow rate is 1.0 mL/min, and the column temperature is 35 ℃. The conditions of the evaporative light scattering detector are that the temperature of the ELSD drift tube is 90 ℃, and the flow rate of N 2 is 2.2L/min.
As a result, the content of diglycerides prepared by various processes is shown in Table 1.
TABLE 1
Analysis of results:
the preferred preparation process is adopted in the embodiments 1, 2 and 3 of the invention, the content of the finally obtained diglyceride is between 85 and 90 percent, and the content of the 1, 3-diglyceride can reach 59 to 65 percent. Has higher yield. Examples 1-1 and 1-2 use single lipase Lipozyme RM or candida antarctica lipase B for biphasic solvent glycerol hydrolysis reaction, the content of prepared diglyceride is lower than that of specific complex enzyme, and the two enzymes have obvious promotion effect on specific generation of diglyceride and 1, 3-diglyceride in the system. Examples 1-3 and 1-4 used different solvent systems, examples 1-3 did not add lecithin adjuvants, examples 1-4 used only single phase aqueous solvent systems, and the yields of diglycerides were not ideal.
Conclusion the preparation process of the present invention can prepare diglyceride product of excellent quality.
The foregoing detailed description is directed to one of the possible embodiments of the present invention, which is not intended to limit the scope of the invention, but is to be accorded the full scope of all such equivalents and modifications so as not to depart from the scope of the invention.
Claims (5)
1. A process for the preparation of diglycerides, comprising the steps of:
(1) Adding solvent to react the grease under the action of first lipase to obtain hydrolysis products;
(2) Mixing the hydrolysate with glycerol, and performing esterification reaction by using a second lipase;
Wherein the first lipase is compounded by lipase Lipozyme RM and candida antarctica lipase B in a mass ratio of 1:1-3;
the solvent is a mixed solution of lecithin, ethanol and water in a mass ratio of 0.01-0.05:0.1-0.2:1;
the temperature of the reaction in the step (1) is 40-65 ℃ and the reaction time is 3-8h;
the dosage of the first lipase is 1-10% of the mass of the grease;
the dosage of the solvent is 10-50% of the mass of the grease;
the second lipase is lipase 435.
2. The method according to claim 1, wherein the mass ratio of the hydrolysate to glycerol is 1:0.5-2.
3. The process according to claim 1, wherein the esterification reaction is carried out at a temperature of 40 to 60 ℃ for a period of 5 to 10 hours.
4. The method of claim 1, wherein the oil is selected from the group consisting of vegetable oils;
The vegetable oil is selected from one or more of linseed oil, soybean oil, rapeseed oil, peanut oil, corn oil, sunflower seed oil, camellia seed oil, coconut oil, palm kernel oil, olive fruit residue oil, walnut oil, rice bran oil, rice oil, cotton seed oil, perilla seed oil, safflower seed oil, grape seed oil, tea seed oil, peony seed oil, sesame oil, wheat germ oil, acer truncatum seed oil, shinyleaf yellowhorn oil, sea buckthorn seed oil, DHA algae oil, medium chain triglyceride, medium and long chain fatty acid edible oil, hemp oil, pricklyash peel seed oil, pricklyash peel oil, pumpkin seed oil, capsicum oil, almond oil and mayonnaise fruit oil;
or, the grease is selected from animal oil.
5. Use of a preparation method according to any one of claims 1-4 for the preparation of 1, 3-diglycerides.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202411312663.7A CN118813729B (en) | 2024-09-20 | 2024-09-20 | A preparation method and application of diglyceride |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202411312663.7A CN118813729B (en) | 2024-09-20 | 2024-09-20 | A preparation method and application of diglyceride |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN118813729A CN118813729A (en) | 2024-10-22 |
| CN118813729B true CN118813729B (en) | 2025-01-03 |
Family
ID=93070107
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202411312663.7A Active CN118813729B (en) | 2024-09-20 | 2024-09-20 | A preparation method and application of diglyceride |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN118813729B (en) |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105400837A (en) * | 2015-12-23 | 2016-03-16 | 华中科技大学 | Method for preparing diglyceride through enzyme catalysis |
| CN116396951A (en) * | 2023-04-14 | 2023-07-07 | 浙江工业大学 | Immobilized composite lipase and preparation method and application thereof |
| CN118652945A (en) * | 2024-08-19 | 2024-09-17 | 广东善百年特医食品有限公司 | A process for producing diglyceride by composite enzyme method |
Family Cites Families (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US7790429B2 (en) * | 2007-11-28 | 2010-09-07 | Transbiodiesel Ltd. | Robust multi-enzyme preparation for the synthesis of fatty acid alkyl esters |
| CN104711298B (en) * | 2015-03-09 | 2017-12-01 | 杭州铎海科技有限公司 | A kind of method that triglycerides enzymolysis prepares 1,3 diglycerides |
| WO2024079301A1 (en) * | 2022-10-14 | 2024-04-18 | Novozymes A/S | Process for selective hydrolysis of diglycerides in an oil/fat with aid of candida antarctica lipase b |
-
2024
- 2024-09-20 CN CN202411312663.7A patent/CN118813729B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105400837A (en) * | 2015-12-23 | 2016-03-16 | 华中科技大学 | Method for preparing diglyceride through enzyme catalysis |
| CN116396951A (en) * | 2023-04-14 | 2023-07-07 | 浙江工业大学 | Immobilized composite lipase and preparation method and application thereof |
| CN118652945A (en) * | 2024-08-19 | 2024-09-17 | 广东善百年特医食品有限公司 | A process for producing diglyceride by composite enzyme method |
Also Published As
| Publication number | Publication date |
|---|---|
| CN118813729A (en) | 2024-10-22 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Sivaramakrishnan et al. | Utilization of microalgae feedstock for concomitant production of bioethanol and biodiesel | |
| Park et al. | Economical DHA (Docosahexaenoic acid) production from Aurantiochytrium sp. KRS101 using orange peel extract and low cost nitrogen sources | |
| Ashokkumar et al. | Production of liquid biofuels (biodiesel and bioethanol) from brown marine macroalgae Padina tetrastromatica | |
| Cho et al. | Effects of enzymatic hydrolysis on lipid extraction from Chlorella vulgaris | |
| Xu et al. | Production of biodiesel from carbon sources of macroalgae, Laminaria japonica | |
| Wang et al. | From microalgae oil to produce novel structured triacylglycerols enriched with unsaturated fatty acids | |
| Carvalho et al. | Biosynthesis, characterization and enzymatic transesterification of single cell oil of Mucor circinelloides–A sustainable pathway for biofuel production | |
| Xu et al. | Bioconversion of volatile fatty acids from macroalgae fermentation into microbial lipids by oleaginous yeast | |
| He et al. | Cost-effective biodiesel production from wet microalgal biomass by a novel two-step enzymatic process | |
| Thangam et al. | Bio-refinery approaches based concomitant microalgal biofuel production and wastewater treatment | |
| US20180340197A1 (en) | Use of an esterase to enhance ethyl ester content in fermentation media | |
| CN103667379B (en) | Method for preparing breast milk fat substitute through lipase-catalyzed acidolysis of algae oil | |
| CN101225415A (en) | Process for Enzymatic Preparation of Diacylglycerol in Organic Medium System | |
| Xiao et al. | Comprehensive study of cultivation conditions and methods on lipid accumulation of a marine protist, Thraustochytrium striatum | |
| Zhang et al. | Utilization of enzymatic cell disruption hydrolysate of Chlorella pyrenoidosa as potential carbon source in algae mixotrophic cultivation | |
| CN114058649A (en) | Solvent-free enzymatic preparation process of diglyceride oil | |
| Gomaa et al. | Enhancement of microalgal biomass, lipid production and biodiesel characteristics by mixotrophic cultivation using enzymatically hydrolyzed chitin waste | |
| CN113774094A (en) | A kind of enzymatic synthesis method of medium and long chain triglyceride | |
| Alavijeh et al. | Enzymatic production of biodiesel from microalgal oil using ethyl acetate as an acyl acceptor | |
| CN104046662A (en) | Transesterification preparation method for 1,3-dioleic acid-2-triglyceride palmitate | |
| CN105400837A (en) | Method for preparing diglyceride through enzyme catalysis | |
| CN107805647A (en) | A kind of method of backbone ester in enzymatic clarification | |
| US10870869B2 (en) | Enzymatic method for preparing glyceryl butyrate | |
| CN106480114B (en) | Method for preparing biodiesel | |
| CN118813729B (en) | A preparation method and application of diglyceride |
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 |