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Imprinted lipase and application thereof

A lipase and imprinting technology, applied in the directions of enzymes, hydrolases, enzymes, etc., can solve the problems of difficult separation, low optical purity of L-lactic acid, and high impurity content, and achieve high optical purity, simple operation, and high enzyme activity. Effect

Active Publication Date: 2021-09-28
WANHUA CHEM GRP
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The technical problem mainly solved by the present invention is that the optical purity of L-lactic acid in the existing lactic acid fermentation products is low (generally only 98%), the content of impurities is large, and the problem of difficult separation

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0063] 1. Preparation of molecularly imprinted cross-linked enzyme polymers

[0064] Weigh 100mg of Candida antarctica lipase B and dissolve it in 10ml of phosphate buffer (pH7.0, 10mM), add 500mg of high optical purity L-lactic acid as a template for imprinting, mix at 4°C and 400rpm for 20min, then add Freeze overnight in liquid nitrogen or in an ultra-low temperature freezer, then transfer to a vacuum freeze dryer to freeze dry. Add 50 mL of isopropanol reagent to the lyophilized enzyme powder to obtain an isopropanol enzyme solution, in which the isopropanol reagent is used as a precipitant to precipitate enzyme protein molecules, and at the same time as a detergent to remove the imprinted template. Then add 0.05 g / 100 ml (calculated by the volume of isopropanol) of SMCC to the isopropanol enzyme solution as a cross-linking agent for cross-linking. After mixing at 400rpm for 20min, refrigerated centrifugation at 4°C and 6000rpm for 20min. Discard the supernatant, add 50 ...

Embodiment 2

[0074] 1. Preparation of molecularly imprinted cross-linked enzyme polymers

[0075] Weigh 200mg of Candida antarctica lipase B and dissolve it in 10ml of phosphate buffer (pH7.0, 10mM), add 300mg of high optical purity L-lactic acid as a template for imprinting, mix at 4°C and 300rpm for 25min, then add Freeze overnight in liquid nitrogen or in an ultra-low temperature freezer, then transfer to a vacuum freeze dryer to freeze dry. Add 80 mL of isopropanol reagent to the lyophilized enzyme powder to obtain an isopropanol enzyme solution, in which the isopropanol reagent is used as a precipitant to precipitate enzyme protein molecules, and at the same time as a detergent to remove the imprinted template. Then add 0.07 g / 100 ml (calculated by the volume of isopropanol) of SMCC to the isopropanolase solution as a cross-linking agent for cross-linking. After mixing at 300rpm for 25min, refrigerated centrifugation at 4°C and 6000rpm for 20min. The supernatant was discarded, and 8...

Embodiment 3

[0085] 1. Preparation of molecularly imprinted cross-linked enzyme polymers

[0086] Weigh 400mg of porcine pancreatic lipase and dissolve it in 10ml of phosphate buffer (pH7.0, 10mM), add 900mg of high optical purity L-lactic acid as a template for imprinting, mix at 4°C and 500rpm for 35min, then add liquid nitrogen or Put it into an ultra-low temperature refrigerator to freeze overnight, and then transfer it to a vacuum freeze dryer to freeze dry. Add 90 mL of isopropanol reagent to the lyophilized enzyme powder to obtain an isopropanol enzyme solution, in which the isopropanol reagent is used as a precipitant to precipitate enzyme protein molecules, and at the same time as a detergent to remove the imprinted template. Then add 0.09 g / 100 ml (calculated by the volume of isopropanol) of SMCC to the isopropanol enzyme solution as a cross-linking agent for cross-linking. After mixing at 500rpm for 35min, refrigerated centrifugation at 4°C and 6000rpm for 20min. Discard the s...

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PUM

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Abstract

The invention discloses imprinted lipase and application thereof. A method for preparing the imprinted lipase comprises the step of performing biological imprinting on lipase by taking L-lactic acid with high optical purity as an imprinting template. The imprinted lipase is used for preparing the L-lactic acid through fermentation, the obtained L-lactic acid is high in optical purity, bacterial strains do not need to be modified, operation is easy and convenient, cost is low, and the method can be suitable for industrial production of the L-lactic acid with the high optical purity.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to imprinted lipase and application thereof, in particular to the application of imprinted lipase in preparing L-lactic acid with high optical purity by a fermentation method. Background technique [0002] Lactic acid, also known as α-hydroxypropionic acid, is one of the three major organic acids in nature. As an important raw material, it is widely used in the fields of food, medicine, chemical industry and materials. In recent years, with the intensification of the greenhouse effect and environmental pollution, polylactic acid (PLA) materials prepared by polymerization of lactic acid monomers have attracted widespread attention. Polylactic acid can be degraded into small molecules under specific conditions to produce carbon dioxide and water, which can alleviate the environmental pressure caused by white pollution and improve the ecological environment. Moreover, polylactic acid is saf...

Claims

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Application Information

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IPC IPC(8): C12N9/20C12P7/56
CPCC12N9/20C12Y301/01003C12P7/56Y02P20/10
Inventor 单雨瑶王竞辉吴计划杨付伟张稳陈长生王坤黎源孔令晓
Owner WANHUA CHEM GRP
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