Method for synthesizing diacylglycerol in reverse micelle enzyme system

A diglyceride and enzyme system technology, applied in chemical instruments and methods, preparation of organic compounds, biochemical equipment and methods, etc., can solve problems such as unusable for a long time, uneven droplet size, incomplete enzyme activity, etc. , to achieve the effects of mild generation conditions, easy operation, and simple synthesis method

Pending Publication Date: 2020-03-20
NANCHANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The preparation method of the existing reverse micellar enzyme system is relatively simple. The surfactant solution and the enzyme solution are mixed and fully shaken until clear. However, the reverse micellar system prepared by this method has uneven droplet size and relatively low stability. Bad question
On the one hand, poor stability makes the system unusable for a long time; on the other hand, the relationship between the droplet size in reverse micelles and the size of lipase is one of the important factors that determine the activity of lipase in reverse micelles. Therefore, due to the inhomogeneity of the droplet size, the enzyme activity is not fully improved and the reproducibility of the results is poor.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Dissolve a certain amount of AOT in isooctane, and mix thoroughly with a vortex oscillator until it becomes clear to obtain an AOT isooctane solution, and then add a certain amount of Candida Antarctica B lipase buffered aqueous solution (50 mol / mL PBS, pH 7.5) to obtain a reverse micellar enzyme solution, in which [enzyme] (mg / mL): [water] (mmol / mL): [AOT] (mmol / mL) =3:375:25, then homogenized by focused ultrasound, the frequency of focused ultrasound was 25 kHz, the power was 300 W, the ultrasonic temperature was 50 °C, the total ultrasonic time was 42 s, and the ultrasonic mode was "ultrasonic 5 s- stop for 2 s". Then 0.7 g / mL peanut oil and glycerol were mixed into the reverse micelles enzyme system at a molar ratio of 1:2, and reacted for 8 h under the condition that the enzyme load was 2% of the total substrate and the reaction temperature was 50 °C. Supplemented by divergent ultrasound, the ultrasonic power is 35 kHz, the power is 300 W, and the ultrasonic mod...

Embodiment 2

[0020] Dissolve a certain amount of AOT in isooctane, and mix thoroughly with a vortex oscillator until it becomes clear to obtain an AOT isooctane solution, and then add a certain amount of Candida Antarctica B lipase buffered aqueous solution (50 mol / mL PBS, pH 7), where [enzyme] (mg / mL): [water] (mmol / mL): [AOT] (mmol / mL) = 4:400:20, Then, homogenization was carried out by focused ultrasound. The frequency of focused ultrasound was 20 kHz, the power was 300 W, the ultrasound temperature was 60 °C, the total ultrasound time was 56 s, and the ultrasound mode was "ultrasound 5 s-stop 2 s". Then, 0.6 g / mL peanut oil and glycerol were mixed into the reverse micelles enzyme system at a molar ratio of 1:1.5, and reacted for 7 h under the condition that the enzyme load was 1.8% of the total substrate and the reaction temperature was 60 °C. The process is supplemented by divergent ultrasound. The ultrasonic power is 30 kHz, the power is 350 W, and the ultrasonic mode is "ultrasoun...

Embodiment 3

[0022] Dissolve a certain amount of AOT in isooctane, and mix thoroughly with a vortex oscillator until it becomes clear to obtain an AOT isooctane solution, and then add a certain amount of Candida Antarctica B lipase buffered aqueous solution (50 mol / mL PBS, pH 6.5), where [enzyme] (mg / mL): [water] (mmol / mL): [AOT] (mmol / mL) = 5:600:30, Then, homogenization was performed by focused ultrasound. The frequency of focused ultrasound was 35 kHz, the power was 350 W, the ultrasound temperature was 40 °C, the total time of ultrasound was 35 s, and the ultrasound mode was "5 s of ultrasound - 2 s off". Then, 0.5 g / mL peanut oil and glycerol were mixed into the reverse micelles enzyme system at a molar ratio of 1:1, and the enzyme load was 1.5% of the total substrate, and the reaction temperature was 40 °C for 6 h. Supplemented by divergent ultrasound, the ultrasonic power is 35 kHz, the power is 300 W, and the ultrasonic mode is "ultrasound 10 s-stop 20 s". After the reaction, add...

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PUM

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Abstract

The invention discloses a method for synthesizing diacylglycerol in a reverse micelle enzyme system using activation characteristics of a lipase interface. The specific synthesis method is as follows:first, constructing the reverse micelle enzyme system, specifically, adding an immobilized water-soluble lipase buffer aqueous solution (50 mmol/mL sodium dihydrogen phosphate-disodium hydrogen phosphate, pH 6.5 to 8) in an isooctane solution of sodium bis(2-ethylhexyl) sulfosuccinate (AOT), and conducting spot energy ultrasound to obtain a clarified reverse micelle enzyme system, wherein the ratio of an enzyme (mg/mL) to water (mmol/mL) to the AOT (mmol/mL) is equal to (3-8):(200-600):(20-30); secondly, mixing 0.5 to 0.8 g/(mL reaction system) plant oil and glycerin with a molar ratio of 1:(1-4), adding a formed mixture into the reverse micelle enzyme system, conducting a reaction for 6-8 h under the conditions that the enzyme load is 1.5-3% of the amount of the total substrate and the reaction temperature is 40 to 60 DEG C, after reacting, adding water with the amounting being 2 times the volume of the reaction liquid, conducting shaking for layering, conducting centrifugation at 4200 rpm for 5min, and taking an upper oil phase which has a diacylglycerol content of 50.45% to 63.23% measured by HPLC; and finally performing twice molecular distillation to purify a product which has a purity of 80.82% to 95.87% measured by HPLC. The method has the advantage of producing diacylglycerol with high efficiency and high yield.

Description

technical field [0001] The invention belongs to the field of grain and oil processing, and in particular relates to a method for synthesizing diglyceride in a reverse micellar enzyme system. Background technique [0002] Structural lipid is a kind of oil with special nutritional effect or physiological function obtained through structural recombination or modification of natural oil. Diacylglycerol is one of them. Concerned, the study found that after 40 healthy men participated in the intake of triglyceride oil or diglyceride oil, the levels of chylomicrons and serum triglycerides in the triglyceride group were significantly higher than those in the diglyceride group. [0003] The preparation of diglycerides is mainly realized under the action of catalysts. According to different catalysts, it can be roughly divided into chemical methods and enzymatic methods. According to different reaction mechanisms, it can also be divided into direct esterification methods, glycerol hyd...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P7/64C12N9/20C12N9/96C07C67/54C07C69/003
CPCC12P7/6445C12N9/20C12N9/96C12Y301/01003C07C67/54C07C69/003
Inventor 邓泽元张兵陈孙妮李静李红艳
Owner NANCHANG UNIV
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