CN104672082B - A kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid - Google Patents
A kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid Download PDFInfo
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- CN104672082B CN104672082B CN201510070107.8A CN201510070107A CN104672082B CN 104672082 B CN104672082 B CN 104672082B CN 201510070107 A CN201510070107 A CN 201510070107A CN 104672082 B CN104672082 B CN 104672082B
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- danshensu
- macroporous
- fermentation liquid
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C51/00—Preparation of carboxylic acids or their salts, halides or anhydrides
- C07C51/42—Separation; Purification; Stabilisation; Use of additives
- C07C51/47—Separation; Purification; Stabilisation; Use of additives by solid-liquid treatment; by chemisorption
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Abstract
The invention discloses and a kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid, comprise the following steps: the large intestine Erichsen of deposit number CGMCC No.7961 is wished the fermentation liquid that bacterium SyBE 002444 fermentation obtains by (1), centrifugal, obtain supernatant, regulation pH=1.5～3.0, as sample solution, standby；(2) sample solution is joined in macroporous adsorptive resins；(3) adding the ethanol water that volumetric concentration is 70%～90% in macroporous adsorptive resins, the flow velocity with 0.5～2BV/h passes through macroporous adsorptive resins eluting, collects eluent；(4) eluent is concentrated, be dried, obtain danshensu.The method of the present invention, have simple to operate, yield is high, advantages of environment protection.The present invention can be used for the isolated and purified of Production by Microorganism Fermentation danshensu.
Description
Technical field
The present invention relates to the isolated and purified of tunning, use danshensu in macroporous adsorbing resin for purification fermentation liquid particularly to one
Method.
Background technology
Macroporous adsorbent resin is a kind of organic polymer copolymer with macroporous structure, is the organic polymer of a class synthetic
Adsorbent.Absorption with macroporous adsorbent resin technology is the earliest for waste water process, medical industry, chemical industry, analytical chemistry, clinic
Calibrating and the field such as treatment, be widely used in the extracting and developing of Chinese herbal medicine effective ingredients, purification work in recent years, such as flavone,
Saponin, the polishing purification of the Chinese medicine ingredients such as alkaloid.
Danshensu (Salvianic acid A, Danshensu), belongs to acidic aromatic compounds, its chemical entitled β-(3,4-
Dihydroxy phenyl) lactic acid, also referred to as danshensu first, molecular formula is C9H10O5, molecular weight is 198.17.Research shows, danshensu
Tool is significantly improved the pharmacologically actives such as hemorheology, lipid lowering, anti-lipid peroxidation, antitumor and visceral organ injury protection.
This laboratory constructs the engineering colon bacillus of synthesis danshensu, it is achieved that the fermentable of danshensu produces, and applies for
Chinese patent, patent name is the biological production of a kind of danshensu, Publication No. CN 103667371 A.At present from greatly
In enterobacteria fermentation liquid, the method for isolated and purified danshensu is not yet reported in media.
Summary of the invention
It is an object of the invention to provide and a kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid.
Technical scheme is summarized as follows:
A kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid, comprise the following steps:
(1) the large intestine Erichsen producing danshensu is wished the fermentation liquid that bacterium fermentation obtains, centrifugal, obtain supernatant, regulate pH=1.5～3.0,
As sample solution, standby；
(2) it is 1～3BV/h sample solution to be joined in macroporous adsorptive resins with loading flow velocity；Sample solution and macroporous adsorbent resin
Ratio is 3～5mL:1g；
(3) adding the ethanol water that volumetric concentration is 70%～90% in macroporous adsorptive resins, the flow velocity with 0.5～2BV/h leads to
Cross macroporous adsorptive resins eluting, collect eluent；The ratio of described ethanol water and macroporous adsorbent resin is 1～2mL:1g；
(4) eluent is concentrated, be dried, obtain danshensu.
PH=2.0 in step (1).
The model of macroporous adsorbent resin is preferably X-5 or ADS-17.
Loading flow velocity in step (2) is preferably 2BV/h.
Preferably the ratio of step (2) sample solution and macroporous adsorbent resin is 5mL:1g.
In step (3), the volumetric concentration of ethanol water is preferably 80%.
Preferably in step (3) ethanol water be the ratio of macroporous adsorbent resin be 1mL:1g.
In step (3), flow velocity is preferably 1BV/h.
The method of the present invention, have simple to operate, yield is high, advantages of environment protection.The present invention can be used for microbe fermentation method
Produce the isolated and purified of danshensu, there is good prospects for commercial application.
Detailed description of the invention
" the large intestine Erichsen in " the large intestine Erichsen producing danshensu wishes the fermentation liquid that bacterium fermentation obtains " used by various embodiments of the present invention
Uncommon bacterium " it is as a example by the large intestine Erichsen of deposit number CGMCC No.7961 wishes bacterium SyBE-002444.
Large intestine Erichsen involved in the present invention wishes bacterium SyBE-002444, it is proposed that Classification And Nomenclature be that large intestine Erichsen wishes bacterium
Escherichia coli, existing in the center preservation of China Committee for Culture Collection of Microorganisms's common micro-organisms, preservation center is registered
Entering volume numbered CGMCC No.7961, the preservation time is on July 22nd, 2013, and address is BeiChen West Road, Chaoyang District, BeiJing City 1
Number No. 3 Institute of Microorganism, Academia Sinica of institute, postcode 100101.
The large intestine Erichsen of deposit number CGMCC No.7961 is wished bacterium SyBE-002444 fermentation process and is seen Chinese patent CN 103667371
A " biological production of a kind of danshensu ", obtains fermentation liquid by the method.
It is noted that the present invention is as a example by above-mentioned large intestine Erichsen wishes bacterium SyBE-002444, is the technology in order to make this area
Personnel better understood when the present invention, but does not impose any restrictions the present invention, it will be appreciated by those skilled in the art that other
The large intestine Erichsen that can produce danshensu wish recombinant bacterium and fall within protection scope of the present invention.
Below in conjunction with specific embodiment, the present invention is further illustrated.
Embodiment 1
A kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid, comprise the following steps:
(1) the large intestine Erichsen producing danshensu is wished the fermentation liquid that bacterium fermentation obtains, centrifugal, obtain supernatant, regulate pH=2.0, make
For sample solution, standby；
(2) each 50g of the macroporous adsorbent resin that model is X-5, ADS-17 of pretreatment is accurately weighed, respectively wet method dress post (25*300
Band core) sample solution that 250mL step (1) obtains is joined in macroporous adsorptive resins with loading flow velocity for 2BV/h respectively；
(3) adding 50mL volumetric concentration respectively in macroporous adsorptive resins is the ethanol water of 80%, leads to the flow velocity of 1BV/h
Cross macroporous adsorptive resins eluting, collect 10～40mL this section of eluents；
(4) eluent is concentrated, be dried, obtain danshensu.
Calculate adsorbance, elution amount and eluting rate.
The adsorbance of X-5 resin is 6.646mg/g, and parsing amount is 5.712mg/g, and resolution factor is 85.9%.
The adsorbance of ADS-17 resin is 3.373mg/g, and parsing amount is 3.357mg/g, and resolution factor is 99.5%.
Embodiment 2
A kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid, comprise the following steps:
(1) the large intestine Erichsen producing danshensu is wished the fermentation liquid that bacterium fermentation obtains, centrifugal, obtain supernatant, take 3 parts and regulate respectively
PH=1.5, pH=2.0, pH=3.0, as sample solution, standby；
(2) model accurately weighing 3 parts of pretreatment is each 50g of X-5 macroporous adsorbent resin, respectively wet method dress post (25*300 band core)
Respectively the sample solution of 250mLpH=1.5, pH=2.0, pH=3.0 is joined X-5 macroporous absorption with loading flow velocity for 2BV/h
In resin column；
(3) adding 50mL volumetric concentration respectively in macroporous adsorptive resins is the ethanol water of 80%, leads to the flow velocity of 1BV/h
Cross macroporous adsorptive resins eluting, collect 10～40mL this section of eluents；
(4) eluent is concentrated, be dried, obtain danshensu.
It is computed adsorbance and is respectively 6.325mg/g, 6.646mg/g, 6.418mg/g.
Embodiment 3
A kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid, comprise the following steps:
(1) the large intestine Erichsen producing danshensu is wished the fermentation liquid that bacterium fermentation obtains, centrifugal, obtain supernatant, regulate pH=2.0, make
For sample solution, standby；
(2) model accurately weighing 3 parts of pretreatment is each 50g of X-5 macroporous adsorbent resin, respectively wet method dress post (25*300 band core)
Take sample solution 150mL, 200mL, 250mL respectively, join X-5 macroporous adsorptive resins with loading flow velocity for 2BV/h
In；
(3) adding 50mL volumetric concentration respectively in macroporous adsorptive resins is the ethanol water of 80%, leads to the flow velocity of 1BV/h
Cross macroporous adsorptive resins eluting, collect 10～40mL this section of eluents；
(4) eluent is concentrated, be dried, obtain danshensu.
It is computed adsorbance and is respectively 4.47mg/g, 5.674,6.646mg/g.
Embodiment 4
A kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid, comprise the following steps:
(1) the large intestine Erichsen producing danshensu is wished the fermentation liquid that bacterium fermentation obtains, centrifugal, obtain supernatant, regulate pH=2.0, make
For sample solution, standby；
(2) model accurately weighing 3 parts of pretreatment is each 50g of X-5 macroporous adsorbent resin, respectively wet method dress post (25*300 band core)
Taking 3 parts of 250mL sample solutions respectively, be respectively 1BV/h with loading flow velocity, 2BV/h, 3BV/h join X-5 macropore and inhale
In attached resin column；
(3) adding 50mL volumetric concentration respectively in macroporous adsorptive resins is the ethanol water of 80%, leads to the flow velocity of 1BV/h
Cross macroporous adsorptive resins eluting, collect 10～40mL this section of eluents；
(4) eluent is concentrated, be dried, obtain danshensu.
It is computed adsorbance and is respectively 6.651mg/g, 6.646mg/g, 5.268mg/g.
Embodiment 5
A kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid, comprise the following steps:
(1) the large intestine Erichsen producing danshensu is wished the fermentation liquid that bacterium fermentation obtains, centrifugal, obtain supernatant, regulate pH=2.0, make
For sample solution, standby；
(2) model accurately weighing 3 parts of pretreatment is each 50g of X-5 macroporous adsorbent resin, respectively wet method dress post (25*300 band core)
Take 3 parts of 250mL sample solutions respectively, join in X-5 macroporous adsorptive resins for 2BV/h with loading flow velocity；
(3) in macroporous adsorptive resins, add 50mL volumetric concentration respectively and be respectively 70%, the ethanol water of 80%, 90%,
Pass through macroporous adsorptive resins eluting with the flow velocity of 1BV/h, collect 10～40mL this section of eluents；
(4) eluent is concentrated, be dried, obtain danshensu.
Calculate desorption efficiency and be respectively 83.5%, 85.9%, 84%.
Embodiment 6
A kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid, comprise the following steps:
(1) the large intestine Erichsen producing danshensu is wished the fermentation liquid that bacterium fermentation obtains, centrifugal, obtain supernatant, regulate pH=2.0, make
For sample solution, standby；
(2) model accurately weighing 3 parts of pretreatment is each 50g of X-5 macroporous adsorbent resin, respectively wet method dress post (25*300 band core)
Take 3 parts of 250mL sample solutions respectively, join in X-5 macroporous adsorptive resins for 2BV/h with loading flow velocity；
(3) adding 50mL volumetric concentration respectively in macroporous adsorptive resins is the ethanol water of 80%, with 0.5BV/h, 1BV/h,
The flow velocity of 2BV/h passes through macroporous adsorptive resins eluting, collects 10～40mL this section of eluents；
(4) eluent is concentrated, be dried, obtain danshensu.
Calculate resolution factor and be respectively 86.1%, 85.3%, 75.2%.
Embodiment 7
A kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid, comprise the following steps:
(1) the large intestine Erichsen producing danshensu is wished the fermentation liquid that bacterium fermentation obtains, centrifugal, obtain supernatant, regulate pH=2.0, make
For sample solution, standby；
(2) model accurately weighing 3 parts of pretreatment is each 50g of X-5 macroporous adsorbent resin, respectively wet method dress post (25*300 band core)
Take 3 parts of 250mL sample solutions respectively, join in X-5 macroporous adsorptive resins for 2BV/h with loading flow velocity；
(3) adding 50mL respectively in macroporous adsorptive resins, 75mL, 100mL volumetric concentration is the ethanol water of 80%, all
Pass through macroporous adsorptive resins eluting with the flow velocity of 1BV/h, collect 10～40mL this section of eluents；
(4) eluent is concentrated, be dried, obtain danshensu.
Calculate resolution factor and be respectively 85.9%, 86.5%, 86.9%.
Claims (7)
1. one kind uses the method for danshensu in macroporous adsorbing resin for purification fermentation liquid, it is characterised in that comprise the following steps:
(1) the large intestine Erichsen producing danshensu is wished the fermentation liquid that bacterium fermentation obtains, centrifugal, obtain supernatant, regulate pH=1.5～3.0,
As sample solution, standby；
(2) it is 1～3BV/h sample solution to be joined in macroporous adsorptive resins with loading flow velocity；Sample solution and macroporous adsorbent resin
Ratio is 3～5mL:1g；
(3) adding the ethanol water that volumetric concentration is 70%～90% in macroporous adsorptive resins, the flow velocity with 0.5～2BV/h leads to
Cross macroporous adsorptive resins eluting, collect eluent；The ratio of described ethanol water and macroporous adsorbent resin is 1～2mL:1g；
(4) being concentrated by eluent, be dried, obtain danshensu, the model of described macroporous adsorbent resin is X-5 or ADS-17.
The most according to claim 1 a kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid, it is characterised in that institute
State step (1) pH=2.0.
The most according to claim 1 a kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid, it is characterised in that institute
Stating the loading flow velocity in step (2) is 2BV/h.
The most according to claim 1 a kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid, it is characterised in that institute
The ratio stating step (2) sample solution and macroporous adsorbent resin is 5mL:1g.
The most according to claim 1 a kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid, it is characterised in that institute
Stating the volumetric concentration of ethanol water in step (3) is 80%.
The most according to claim 1 a kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid, it is characterised in that institute
Stating the ratio of ethanol water and macroporous adsorbent resin described in step (3) is 1mL:1g.
The most according to claim 1 a kind of use the method for danshensu in macroporous adsorbing resin for purification fermentation liquid, it is characterised in that institute
Stating flow velocity in step (3) is 1BV/h.
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| US5667902A (en) * | 1996-04-30 | 1997-09-16 | Mobil Oil Corporation | High moisture barrier polypropylene-based film |
| AU2003241211A1 (en) * | 2003-06-19 | 2005-01-04 | Biokab, S.A. De C.V. | Method for the recovery of ferulic acid |
| CN1264506C (en) * | 2004-09-10 | 2006-07-19 | 张平 | Pharmaceutical combination containing red sage root element and preparation method thereof |
| CN102464581B (en) * | 2010-11-12 | 2015-02-25 | 天士力制药集团股份有限公司 | Method for preparing high-purity sodium danshensu |
| CN103667371B (en) * | 2013-11-11 | 2016-03-16 | 天津大学 | A kind of biological production of Salvianic acidA |
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