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Method for quickly and efficiently extracting active antimicrobial substance HSAF (Heat Stable Antifungal Factor) in fermentation liquor

A technology of antibacterial substances and fermented liquid, applied in the field of separation engineering, can solve the problems of HSAF industrial production difficulties, easy emulsification of the system, increase of production process, etc., and achieve the effects of shortening extraction time, easy stratification, and improving production efficiency

Active Publication Date: 2017-07-14
JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the presence of various metabolites in the fermentation broth, such as proteins and sugars, the system is prone to emulsification, resulting in difficult phase separation and serious product loss
Generally, multiple repeated extraction strategies are used. Although more target products can be recovered, it is time-consuming and labor-intensive, and the production process is increased, which greatly reduces the production efficiency and brings great difficulties to the laboratory research and industrial production of HSAF.

Method used

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  • Method for quickly and efficiently extracting active antimicrobial substance HSAF (Heat Stable Antifungal Factor) in fermentation liquor
  • Method for quickly and efficiently extracting active antimicrobial substance HSAF (Heat Stable Antifungal Factor) in fermentation liquor
  • Method for quickly and efficiently extracting active antimicrobial substance HSAF (Heat Stable Antifungal Factor) in fermentation liquor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Embodiment 1: the influence of different organic solvents on the extraction of HSAF in the fermented liquid

[0034] Draw 3mL of the fermentation broth into a 15mL centrifuge tube, add concentrated hydrochloric acid dropwise until the pH is 3.0; add 3mL of organic solvent (Table 1) to the mixture; place it on a vortex mixer at 2000rpm for 1min to make the organic solvent and the fermentation broth fully Contact; centrifuge at 8000rpm for 5min to separate the fermentation broth and the organic solvent into two phases, the supernatant is the HSAF organic solvent layer, draw 1mL to dry and add 500μL of methanol to dissolve and then detect its content by HPLC.

[0035] Detection conditions are: reversed-phase column: InterSustainSwift C18 5μm, 250×4.6mm, mobile phase: solution A (0.025% TFA in water) and solution B (0.025% TFA in acetonitrile), flow rate: 1mL / min; injection volume: 20μL , UV absorption value: 318nm; Injection program: 0-10min, solution B from 5% to 25%; 25m...

Embodiment 2

[0039] Embodiment 2: adding CaCl 2 Breaking the emulsified extraction of HSAF in the fermentation broth

[0040] Draw 3mL of fermentation broth into a 15mL centrifuge tube, add concentrated hydrochloric acid dropwise to pH 3.5; add 0.5g CaCl 2 Finally, shake it gently to make it fully dissolved; add 3mL ethyl acetate solvent to the mixture; place it on a vortex mixing oscillator at 2000rpm for 1min to make the extractant fully contact with the fermentation broth; centrifuge at 8000rpm for 5min to make the organic phase and The water phase is separated, and the supernatant is drawn to be the HSAF extract, and its content is detected by HPLC, and other conditions are the same as in Example 1.

[0041] Depend on figure 2 As can be seen from Table 2, CaCl 2 It can effectively break the emulsification phenomenon produced in the extraction process, and can increase the content of HSAF in the extract (336.8mg / L). At the same time, the HSAF content obtained after the full extract...

Embodiment 3

[0042] Embodiment 3: adding NaCl to break the HSAF in the emulsified extraction fermented liquid

[0043] Draw 3mL of fermentation broth into a 15mL centrifuge tube, add concentrated hydrochloric acid dropwise until the pH is 4.0; put in 0.6g NaCl, shake gently to dissolve it fully; add 3mL ethyl acetate solvent to the mixture; place in a vortex mixer React at 2000rpm for 1min to fully contact the extractant with the fermentation broth; centrifuge at 8000rpm for 5min to separate the fermentation broth and the organic solvent into two phases, the supernatant is the HSAF extract, draw 1mL to dry and add 500μL of methanol to dissolve and follow the steps in Example 1 The middle scheme was used to detect its content.

[0044] Depend on figure 2 It can be seen from Table 2 that NaCl can effectively break the emulsification phenomenon produced during the extraction process, and can increase the content of HSAF in the extract (343.6mg / L). At the same time, the HSAF content obtaine...

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Abstract

The invention discloses a method for quickly and efficiently extracting an active antimicrobial substance HSAF (Heat Stable Antifungal Factor) in fermentation liquor. The method comprises the four steps of (1) acidizing the fermentation liquor; (2) breaking emulsification through inert matters; (3) vibrating for extracting ethyl acetate;(4) centrifuging, and absorbing a supernate for extracting the HSAF. According to the method provided by the invention, the added inert matters can effectively break emulsification, the extraction reaction time is shortened to be 1min by adopting the vibrating treatment, and the extraction speed is greatly improved. The method provided by the invention can maximally enable the single extraction efficiency of the HSAF in the fermentation liquor to reach to 88.2 percent which is improved by 1.83 times compared with the control, is convenient to operate and simple in process, and is a summary of an extraction process for the HSAF, which is a product with a great application prospect for the first time.

Description

technical field [0001] The invention belongs to the technical field of separation engineering, and in particular relates to a method for rapidly and efficiently extracting HSAF, an active antibacterial substance in a fermentation broth. Background technique [0002] Biological pesticides refer to pesticides made from living organisms, biological metabolites, or biologically specific genes. At present, agricultural antibiotics (Jinggangmycin, abamectin, etc.), microbial pesticides (Bacillus thuringiensis, Pseudomonas monas, etc.), botanical pesticides (matrine, pyrethrins, etc.), etc. It has the advantages of low toxicity, low residue, and not easy to produce drug resistance, which provides a very important research direction for the sustainable development of agriculture and has broad prospects. [0003] [0004] Lysobacterenzymogenes is a new type of biocontrol bacteria, which can secrete a variety of extracellular enzymes and various secondary metabolites. It has a goo...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07D487/08
CPCC07D487/08
Inventor 刘凤权汤宝史晓敏赵延存赵杨扬徐会永孙伟波
Owner JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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