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A kind of purification preparation method of simonoside I

A siamanside, eluent technology, applied in chemical instruments and methods, organic chemistry, glycoside steroids, etc., can solve the problems of difficult large-scale production, low natural content, difficult product quality control, etc., to reduce process costs, Simple operation and good repeatability

Active Publication Date: 2022-02-22
CHINA PHARM UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the natural content of simonoside I is extremely low, less than 0.5%, and it is difficult to produce on a large scale
However, due to the particularity of bacterial components, the purified simonoside I is often mixed with impurities such as proteins and water-soluble pigments, making it difficult to control the quality of the product

Method used

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  • A kind of purification preparation method of simonoside I
  • A kind of purification preparation method of simonoside I
  • A kind of purification preparation method of simonoside I

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] The HPLC detection method of embodiment 1 symonoside I

[0026] The HPLC analysis of symmonoside I sample involved in the present invention, its method parameter is all as follows:

[0027] Instrument model: Agilent 1260

[0028] Chromatographic column: YMC-Pack ODS-A (250mm×4.6mm, 5μm)

[0029] Mobile phase A: water (containing 1‰ formic acid); mobile phase B: acetonitrile (containing 1‰ formic acid)

[0030] Column temperature: 30°C; Flow rate: 1mL / min; Detection wavelength: 205nm; Injection volume: 5μL

[0031] Elution gradient: 10% to 90%; gradient time 25min.

[0032] Under the above-mentioned HPLC detection conditions, the retention times of mogroside IIIE and simonoside I are respectively 8.7min and 9.5min ( figure 1 ).

Embodiment 2

[0033] The biosynthesis of embodiment 2 simonoside I and the comparison of the method for removing bacterial fragments

[0034] Utilize the whole-cell biocatalyst in CN112063678A to construct a 0.5L biosynthesis system of simonoside I, wherein the concentration of the substrate mogroside IIIE is 15g / L, the concentration of sucrose is 500mg / mL, and the reaction solution is M9 medium with pH=9.0, Reaction time 12 hours. After the reaction, the pH of the reaction solution was adjusted to 5.0-5.5 with dilute hydrochloric acid (0.1M), and divided into two equal parts (A and B). A reaction solution was boiled for 5 minutes, filtered to remove protein and bacterial fragments. Add an equal amount of absolute ethanol to the B reaction solution, mix thoroughly and let it stand for 5 minutes, then filter to remove protein and bacterial fragments. The obtained filtrate showed a slight yellow color, and still contained protein detected by SDS-PAGE, and there was no significant difference...

Embodiment 3

[0035] The adsorption investigation of embodiment 3 macroporous adsorption resin

[0036] Referring to literature reports, the present invention investigates the purification effects of five nonpolar macroporous resins D3520, HPD100, D101, HP20 and AB-8 on symanoside I. First soak the macroporous resin in 95% ethanol solution for 8 hours, wash with distilled water until there is no ethanol smell; then soak in 5% NaOH solution for 4 hours, wash with distilled water until neutral, then soak with 5% hydrochloric acid for 4 hours, and finally use Wash with distilled water until neutral.

[0037] Weigh 5 g of each of the above pretreated five pretreated resins and place them in shake flasks, add 40 mL of boiled reaction solution to each bottle, and vibrate and adsorb for 12 hours at room temperature. Take out the resin, shake and desorb with 95% ethanol solution for 6 hours, measure the content of symanoside I in the reaction solution before and after adsorption, and the analysis ...

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Abstract

The invention discloses a method for purifying and preparing high-purity natural sweetener siamenoside I from a biocatalytic reaction system. Specifically, it involves the removal of bacterial debris and proteins, the removal of pigments, and the preparation of product purification. The present invention comprehensively applies the protein removal method and separation materials such as ion exchange resin and macroporous resin, and establishes a process for preparing high-quality simanoside I from a biosynthesis system, and the product yield exceeds 70%. Only water and ethanol are used in the purification preparation process, the process is simple, the cost is low, and the safety is high, and industrial application is expected.

Description

technical field [0001] The invention belongs to the technical field of natural product biosynthesis and separation and purification, and in particular relates to the biosynthesis, purification and preparation of mogroside natural sweetener simonoside I. Background technique [0002] After entering the 21st century, with the continuous improvement of human living standards, the number of "three high" people has also shown explosive growth, among which diabetes has become a global chronic disease, occupying a large amount of medical resources. In recent years, people have gradually realized the importance of a low-sugar diet to health. Therefore, natural sweeteners such as stevioside, rubusoside, glycyrrhizic acid, rebaudioside A, mogroside, etc. have gradually entered people's field of vision and become synonymous with low-calorie dietary additives, showing extremely high commercial value. [0003] Siamenoside I (siamenoside I) is a cucurbitane-type tetracyclic triterpene sa...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07J17/00
CPCC07J17/005
Inventor 吴旭日杜雅丽苏咏欣刘世强陈漫婷陈依军
Owner CHINA PHARM UNIV