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Method for enriching DHA and EPA in fish oil based on liquid immobilized enzyme

An immobilized enzyme enrichment fish, immobilized lipase technology, applied in immobilized enzymes, biochemical equipment and methods, enzymes and other directions, can solve the difficulty of lipase separation, difficult to reuse immobilization cost, lipase volatile In order to reduce the loss and purification cost, reduce the application cost, and weaken the inactivation

Active Publication Date: 2019-02-01
SOUTH CHINA UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to provide a novel liquid-based immobilization method for the current biological enzymatic method to enrich EPA and DHA, the existing problems of lipase separation difficulty, easy inactivation of lipase, difficulty in reutilization and high cost of immobilization, etc. A method for efficiently reusing enzymatic enrichment of EPA and DHA in fish oil

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Take 6.5g of crude enzyme solution (75U / mL, pH 6) containing lipase AYS and 20mM phosphate buffer system, add 1.9g of sodium sulfate, mix well, add 1.6g of polyethylene glycol 400, separate liquid Upper layer gets 3g liquid immobilized lipase; Liquid immobilized lipase is placed in reactor, adds 30% sodium sulfate solution (containing the phosphate buffered saline of 1Mm, pH is 6) of 6.8g, 2g fish oil, under stirring at 200rpm The reaction was controlled at 37°C for 1h. After the reaction, centrifuge at 3000rpm for 3min. The experimental components are divided into three liquid phases: upper, middle and lower. The middle liquid phase is liquid immobilized enzyme. After taking it out, add 5mL of the salt-enriched lower phase solution of the above system, 2g of fish oil , reacted under stirring at 200rpm, controlled to repeat the reaction at 37°C for 1h, and so on, repeated 3 times in total, and measured the enzyme activity by the p-nitrophenol method. After the three rea...

Embodiment 2

[0033]Take 6.5g of crude enzyme solution (75U / mL, pH 6) containing lipase AYS and 20mM phosphate buffer system, add 1.9g of sodium sulfate, mix well, add 1.6g of polyethylene glycol 400, separate liquid 3 g of liquid immobilized lipase was obtained from the upper layer; the liquid immobilized lipase was placed in a reactor, 7 g of 35% sodium sulfate solution and 2 g of fish oil were added, and the mixture was stirred at 200 rpm and reacted at 37° C. for 1 h. After the reaction, centrifuge at 3000rpm for 3min. The experimental components are divided into three liquid phases: upper, middle and lower. The middle liquid phase is liquid immobilized enzyme. After taking it out, add 5mL of the salt-enriched lower phase solution of the above system, 2g of fish oil , reacted under stirring at 200rpm, controlled to repeat the reaction at 37°C for 1h, and so on, repeated 7 times in total, and measured the enzyme activity by the p-nitrophenol method. After seven reactions, the enzyme activ...

Embodiment 3

[0035] Take 6.5g of crude enzyme solution (80U / mL) containing lipase MAS1, add 1.5g of sodium sulfate, mix well and add 2g of [BMIM]BF 4 , liquid separation and take the upper layer to get 3.5g liquid immobilized lipase; put the liquid immobilized lipase in the reactor, add 6.2g of 36% sodium sulfate solution, 2g fish oil, react under stirring at 200rpm, and control it at 37°C Reaction 2h. After the reaction, centrifuge at 3000rpm for 3min. The experimental components are divided into three liquid phases: upper, middle and lower. The middle liquid phase is liquid immobilized enzyme. After taking it out, add 5mL of the salt-enriched lower phase solution of the above system, 2g of fish oil , react under stirring at 200rpm, repeat the reaction at 37°C for 2h, and measure the enzyme activity by the p-nitrophenol method. After the reaction, the enzyme activity retention rate is 90% of the initial reaction enzyme activity, and the EPA and DHA on the glyceride The enrichment rates w...

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Abstract

The invention discloses a method for enriching DHA and EPA in fish oil based on an immobilized enzyme. The method comprises the following steps: preparing a lipase solution, adding a soluble salt anda hydrophilic solvent to the lipase solution to form a two-phase system, and carrying out liquid separation to obtain an upper liquid immobilized lipase phase and a salt-rich lower phase; blending theobtained liquid immobilized lipase with a soluble salt solution and fish oil to form a three-liquid phase system, carrying out an enzyme catalyzed reaction under a stirring condition, and standing orcentrifuging the obtained solution after the reaction is finished in order to obtain three layers which are an upper phase being a fish oil product, a middle phase being the liquid immobilized lipaseand a lower phase being a reaction medium rich in glycerol hydrolysate; and adding the upper phase fish oil product to an alkali solution, then adding a hydrophobic organic solvent, carrying out standing extraction, taking the obtained upper phase, and distilling the upper layer to obtain a glyceride product rich in EPA and DHA. The method realizes the rapid separation of the product and the catalyst, and the lipase can be repeatedly used, and can improve the catalytic efficiency, so the enzyme loss and the purifying cost are greatly reduced.

Description

technical field [0001] The invention belongs to the field of bioengineering and food technology, and relates to enzyme separation and application technology, in particular to a method for hydrolyzing DHA and EPA in fish oil with lipase. Background technique [0002] w-3 polyunsaturated fatty acids (EPA, DHA) have excellent blood lipid-lowering, anti-inflammatory and cholesterol-lowering and other physiological activities, so they have received extensive attention in recent years and become widely sought after anti-cardiovascular and cerebrovascular diseases. Functional health products. According to statistics, its global market sales have soared from US$150 million in 2004 to US$1.6 billion in 2016. However, the content of EPA and DHA in natural fish oil is low, which cannot meet the relevant needs. It is necessary to refine and enrich the EPA and DHA in natural fish oil through enzymatic hydrolysis, alcoholysis, molecular distillation and other methods to remove relatively...

Claims

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Application Information

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IPC IPC(8): C12N9/20C12N11/00C12P7/64
CPCC12N9/20C12N11/00C12P7/6427C12Y301/01003
Inventor 李志刚杨博王永华陈华勇陆德林陈华
Owner SOUTH CHINA UNIV OF TECH
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