CN105274158B

CN105274158B - method for extracting polyglutamic acid from fermentation liquor by using aqueous two-phase technology

Info

Publication number: CN105274158B
Application number: CN201510817118.8A
Authority: CN
Inventors: 马霞; 赵帅; 李路遥
Original assignee: Shanghai Institute of Technology
Current assignee: Shanghai Institute of Technology
Priority date: 2015-11-23
Filing date: 2015-11-23
Publication date: 2020-01-31
Anticipated expiration: 2035-11-23
Also published as: CN105274158A

Abstract

The invention discloses methods for extracting polyglutamic acid from fermentation liquor by adopting a double aqueous phase technology, which comprises the steps of firstly preparing a mixed solution, establishing a double aqueous phase extraction system, enabling the mass fraction of PEG1000 to be 10-30%, (NH)₄)₂SO₄5-25% of mass fraction, 10% of fermentation liquor mass fraction, 3.75-8.46 of fermentation liquor pH value, then controlling the environmental temperature to be 20-40 ℃, naturally carrying out phase separation on the mixed liquor, taking the upper phase, adopting an ultrafiltration membrane with the molecular weight cutoff of 30KDa to permeate PEG1000, finally freezing the trapped polyglutamic acid concentrated solution at the low temperature of-40 ℃, and carrying out vacuum freeze drying at the temperature of-50 ℃ for 24 hours to obtain a pure product. The invention solves the problems of viscous polyglutamic acid fermentation liquor and overlarge pretreatment cost, and the subsequent aqueous two-phase system has mild reaction, thereby avoiding the damage of the traditional precipitation method to the molecular weight of the polyglutamic acid.

Description

method for extracting polyglutamic acid from fermentation liquor by using aqueous two-phase technology

Technical Field

The invention relates to methods for extracting polyglutamic acid from fermentation liquor by adopting an aqueous two-phase technology, belonging to the technical field of biological separation engineering.

Background

The gamma-PGA has excellent water solubility, super-strong adsorbability and biodegradability, the degradation product is pollution-free glutamic acid, the gamma-PGA is excellent environment-friendly high polymer materials, can be used as a water retention agent, a heavy metal flocculant, a slow release agent, a drug carrier and the like, has great commercial value and social value in industries such as cosmetics, environmental protection, food, medicine, agriculture and the like, but the polyglutamic acid fermentation liquor is viscous, the pretreatment cost is overlarge, and the traditional precipitation method can damage the molecular weight of the polyglutamic acid.

Disclosure of Invention

The invention aims at to solve the problems that the polyglutamic acid fermentation liquor is viscous, the pretreatment cost is overlarge, the traditional precipitation method causes damage to the molecular weight of the polyglutamic acid and the like, provides aqueous two-phase systems with simple process, low cost and mild reaction, and simultaneously avoids the damage to the molecular weight of the polyglutamic acid.

The second purpose of the invention is to provide methods for extracting polyglutamic acid from fermentation liquor by adopting a two-aqueous-phase technology.

The technical scheme of the invention is specifically introduced as follows.

method for extracting polyglutamic acid from fermentation broth by aqueous two-phase technology, comprises preparing mixed solution, establishing aqueous two-phase extraction system to make PEG1000 mass fraction 10-30%, (NH)₄)₂SO₄The mass fraction is 5 to 25 percentThe mass fraction of the fermentation liquor is 10 percent, the pH value of the fermentation liquor is 3.75-8.46, then the environmental temperature is controlled to be 20-40 ℃, the mixed solution is naturally split, the upper phase is taken, an ultrafiltration membrane with the molecular weight cutoff of 30KDa is adopted to permeate PEG1000, finally, the intercepted polyglutamic acid concentrated solution is frozen at the low temperature of minus 40 ℃, and then is frozen and dried in vacuum at the temperature of minus 50 ℃ for 24 hours, and the pure product is obtained.

method for extracting polyglutamic acid from fermentation liquor by adopting aqueous two-phase technology, which comprises the following steps:

(1) preparation of Mixed solution

Preparing mixed solution, establishing a double water phase extraction system to ensure that the PEG1000 mass fraction is 10-30%, (NH)₄)₂SO₄5 to 25 percent of mass fraction, 10 percent of mass fraction of fermentation liquor and 3.75 to 8.46 percent of pH value of the fermentation liquor;

(2) natural phase splitting

Controlling the ambient temperature to be 20-40 ℃, and naturally splitting the phases of the mixed solution prepared in the step (1);

(3) and (3) taking the upper phase of the mixed solution in the step (2), adopting an ultrafiltration membrane with the molecular weight cutoff of 30KDa to permeate PEG1000, freezing the intercepted polyglutamic acid concentrated solution at the low temperature of minus 40 ℃, and carrying out vacuum freeze drying for 24 hours at the temperature of minus 50 ℃ to obtain a pure product.

The invention has the following beneficial effects:

the method for extracting polyglutamic acid from fermentation liquor by adopting the aqueous two-phase technology has the advantages of simple process and low cost, solves the problem of viscosity of the polyglutamic acid fermentation liquor, and avoids the damage of the traditional precipitation method to the molecular weight of the polyglutamic acid.

Detailed Description

The invention is further illustrated by, but is not limited to, by the following specific examples.

fermentation liquor for extracting polyglutamic acid from the fermentation liquor by adopting a two-aqueous-phase technology is prepared by laboratories.

miniature tangential flow ultrafiltration devices for extracting polyglutamic acid from fermentation broth by aqueous two-phase technology are manufactured by Millipore corporation of America.

vacuum freeze-drying machines for extracting polyglutamic acid from fermentation liquor by using a two-aqueous-phase technology are produced by Beijing Bo Yi kang laboratory instruments Co.

Example 1

(1) preparation of Mixed solution

Preparing mixed solution, and establishing a two-aqueous phase extraction system to ensure that the PEG1000 mass fraction is 10 percent and (NH)₄)₂SO₄The mass fraction is 5%, the mass fraction of the fermentation liquor is 10%, and the pH value of the fermentation liquor is 3.75;

(2) natural phase splitting

Controlling the ambient temperature to be 20 ℃, and naturally splitting the phases of the mixed solution prepared in the step (1);

The obtained polyglutamic acid aqueous two-phase system has the distribution coefficient of 0.93 and the extraction rate of 51.25 percent.

Example 2

(1) preparation of Mixed solution

Preparing mixed solution, establishing a two-aqueous phase extraction system to ensure that the PEG1000 mass fraction is 30 percent and (NH)₄)₂SO₄The mass fraction is 25%, the mass fraction of the fermentation liquor is 10%, and the pH value of the fermentation liquor is 8.46;

(2) natural phase splitting

Controlling the ambient temperature to be 40 ℃, and naturally splitting the phases of the mixed solution prepared in the step (1);

The obtained polyglutamic acid aqueous two-phase system has a distribution coefficient of 0.81 and an extraction rate of 45.06%.

Example 3

(1) preparation of Mixed solution

Preparing mixed solution, and establishing a two-aqueous phase extraction system to make PEG1000 mass fraction 20%, (NH)₄)₂SO₄15 percent of mass fraction, 10 percent of mass fraction of fermentation liquor and 5.31 percent of pH value of the fermentation liquor;

(2) natural phase splitting

Controlling the ambient temperature to be 30 ℃, and naturally splitting the phases of the mixed solution prepared in the step (1);

The obtained polyglutamic acid aqueous two-phase system has the distribution coefficient of 1.21 and the extraction rate of 67.59 percent.

Example 4

(1) preparation of Mixed solution

Preparing mixed solution, and establishing a two-aqueous phase extraction system to make PEG1000 mass fraction 15%, (NH)₄)₂SO₄The mass fraction is 10%, the mass fraction of the fermentation liquor is 10%, and the pH value of the fermentation liquor is 4.66;

(2) natural phase splitting

Controlling the ambient temperature to be 25 ℃, and naturally splitting the phases of the mixed solution prepared in the step (1);

The obtained polyglutamic acid aqueous two-phase system has the distribution coefficient of 2.31 and the extraction rate of 78.02 percent.

Example 5

(1) preparation of Mixed solution

Preparing mixed solution, establishing a two-aqueous phase extraction system to ensure that the mass fraction of PEG1000 is 25 percent and (NH)₄)₂SO₄The mass fraction is 20%, the mass fraction of the fermentation liquor is 10%, and the pH value of the fermentation liquor is 7.55;

(2) natural phase splitting

Controlling the ambient temperature to be 35 ℃, and naturally splitting phases of the mixed solution prepared in the step (1);

The obtained polyglutamic acid aqueous two-phase system has the distribution coefficient of 1.52 and the extraction rate of 73.25 percent.

In conclusion, the methods for extracting polyglutamic acid from fermentation liquor by adopting the aqueous two-phase technology have the advantages of simple process and low cost, the distribution coefficient is 0.81-2.31, and the extraction rate is 45.06% -78.02%.

The above examples are only for illustrating the present invention, and other embodiments of the present invention are also possible, but those skilled in the art will be able to make equivalents and modifications thereof based on the technical teaching of the present invention, and fall within the scope of the present invention.

Claims

1, method for extracting polyglutamic acid from fermentation liquor by adopting aqueous two-phase technology, which is characterized by comprising the following steps:

(1) preparing mixed solution from the mixed solution, establishing a two-aqueous phase extraction system to ensure that the mass fraction of PEG1000 is 25 percent and (NH)₄)₂SO₄The mass fraction is 20%, the mass fraction of the fermentation liquor is 10%, and the pH value of the fermentation liquor is 4.66;

(2) naturally splitting the phases, controlling the ambient temperature to be 25 ℃, and naturally splitting the phases of the mixed solution prepared in the step (1);

2. The method for extracting polyglutamic acid from fermentation broth by aqueous two-phase technology as claimed in claim 1, wherein: when the polyglutamic acid is extracted, the partition coefficient is 2.31, and the extraction rate is 78.02%.