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Method for producing 1,3-diglyceride by immobilized lipase

A technology for immobilizing lipase and diglyceride, applied to biochemical equipment and methods, enzymes immobilized on or in inorganic carriers, etc., can solve problems such as difficult to obtain high-purity diglyceride and difficult reaction , to achieve the effect of simple separation and purification, simple recovery and high purification efficiency

Active Publication Date: 2018-07-17
CABIO BIOTECH WUHAN CO LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the existing process, after the reaction, there will be some unreacted oleic acid and the monoglyceride produced in the reaction. At the same time, it is difficult to obtain high-purity diglyceride due to the limitation of lipase selectivity.
From the perspective of immobilized enzymes, traditional resin-immobilized enzymes are only suitable for use in packed beds, and the reaction is difficult to carry out in packed beds due to the immiscibility of fatty acids and glycerol

Method used

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  • Method for producing 1,3-diglyceride by immobilized lipase
  • Method for producing 1,3-diglyceride by immobilized lipase
  • Method for producing 1,3-diglyceride by immobilized lipase

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0087] Example 1 Nano Fe 3 O 4 Synthesis

[0088] 11.75g ​​FeCl 3 ·6H 2 O, 5.97g FeSO 4 ·7H 2 O was dissolved in 250ml of deionized water, heated to 80°C, and 14ml of ammonia was quickly added under rapid stirring, and kept for 2h under the protection of nitrogen. Wash the precipitate with deionized water until the aqueous solution is neutral to obtain Fe 3 O 4 . Its transmission electron microscope is like figure 1 (a) Shown.

[0089] Will get Fe 3 O 4 Add 500ml of 80% ethanol, slowly add 20ml of APTES (3-aminopropyl-triethoxysilane) at 30℃, react for 12h at room temperature, wash with deionized water and ethanol three times to obtain aminated Fe 3 O 4 . Aminated Fe 3 O 4 Add to 100ml 2% glutaraldehyde solution, react at 30℃ for 2h, wash and freeze-dry to get about 5g Fe with aldehyde groups on the surface 3 O 4 . Its transmission electron microscope such as figure 1 (b) Shown.

[0090] by figure 1 (a) and figure 1 (b) It can be seen that after the ferroferromagnetic particles ar...

Embodiment 2

[0091] Example 2 Screening of Lipase

[0092] Weigh 0.115g glycerol (1.25mM), 0.76g oleic acid (2.5mM), 10mg lipase to be screened (the amount of liquid enzyme added is 10μl), place it in a 2ml flat-bottom centrifuge tube, add about 0.23g (oleic acid) 5 times the mass of water produced by the complete reaction) activated 4A molecular sieve and a 2×5mm jujube-shaped magnet.

[0093] The reaction temperature was controlled to 30°C by a constant temperature water bath, and the reaction was stirred magnetically for 12 hours. After the reaction, the reaction was centrifuged, and 50 μl of the upper layer sample was taken and analyzed by liquid chromatography. The lipase with catalytic activity in the solvent-free system and the highest selectivity to the 1,3 hydroxyl groups of glycerol was obtained.

[0094] The screening results are as follows:

[0095] Table 1 Catalytic performance of different lipases

[0096]

[0097] It can be seen from Table 1 that the oleic acid conversion rate of th...

Embodiment 3

[0098] Example 3 Immobilized Lipase

[0099] Take 1g Rhizopus oryzae lipase enzyme powder (protein content is about 10.5%) dissolved in 30ml pH 6.5 phosphate buffer (100mM), add 0.3g Tween 80, stir to dissolve the enzyme powder, and then add 2g activated Ferric oxide was stirred for 4 hours at 20°C, and the immobilized enzyme obtained was magnetically separated and washed with buffer for several times to wash away unbound proteins and Tween 80.

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Abstract

The invention discloses a method for producing 1,3-diglyceride by immobilized lipase. The method comprises the following steps: mixing unsaturated long-chain fatty acid, glycerinum and the immobilizedlipase, performing stirring reaction at 25 to 40 DEG C for 1 to 12 hours, and performing post-treatment to obtain the 1,3-diglyceride, wherein the unsaturated long-chain fatty acid is at least one ofoleic acid, linoleic acid, linolenic acid, arachidonic acid and docosahexenoic acid; a mole ratio of the unsaturated long-chain fatty acid to the glycerinum is (1 to 3):1; based on the volume of a reaction system, the mass volume concentration of the immobilized lipase is 0.4 to 1 percent; the immobilized lipase is at least one of immobilized rhizopus lipase, myrhizopus lipase, lipase RM-IM, palatase 20000L lipase and Xhlip-O lipase. The method disclosed by the invention has the characteristics of high raw material conversion rate, high enzyme catalysis efficiency, a few of produced impurities, low production cost, suitability for large-scale industrial production and the like.

Description

Technical field [0001] The present invention relates to the field of biochemical industry, in particular to a method for producing 1,3-diglyceride by immobilized lipase. Background technique [0002] Diglyceride is a natural component in fats and oils, but it is less than the content of triglyceride, usually less than 5%. About 70% of diglycerides are in the form of 1,3-diglycerides in weight-loss cooking edible fats; and in the intermediate products of ordinary fat metabolism, they are mainly 1,2(2,3)-glycerol Exist in diester form. The metabolism of 1,3-diglycerides is different from triglycerides and 1,2-diglycerides. It is hydrolyzed into free fatty acids and 1(3)-monoglycerides under the action of digestive enzymes mainly pancrelipase. , 1(3)-monoglyceride is released in the form of energy through β-oxidation, so that it is rarely stored in the body to prevent the accumulation of body fat. Therefore, eating fats containing 1,3-diglycerides has the effect of reducing fat a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P7/64C12N11/14
CPCC12N9/20C12N11/14C12P7/6454C12P7/6472C12Y301/01003
Inventor 杨立荣李翔宇吴绵斌赵炯烽汪志明马凡提
Owner CABIO BIOTECH WUHAN CO LTD
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