CN103954726A - Method for extracting muramic acid from corynebacterium glutamicum dregs and detection method - Google Patents
Method for extracting muramic acid from corynebacterium glutamicum dregs and detection method Download PDFInfo
- Publication number
- CN103954726A CN103954726A CN201410188150.XA CN201410188150A CN103954726A CN 103954726 A CN103954726 A CN 103954726A CN 201410188150 A CN201410188150 A CN 201410188150A CN 103954726 A CN103954726 A CN 103954726A
- Authority
- CN
- China
- Prior art keywords
- acid
- thin
- muramic acid
- filter residue
- muramic
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Landscapes
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Compounds Of Unknown Constitution (AREA)
Abstract
The invention discloses a thin-layer chromatographic scanning detection method of muramic acid. The method has the advantages of being simple, rapid, accurate, reliable, stable and the like and comprises the following steps: (1) taking a to-be-detected sample liquor, sampling on a high-efficiency thin-layer silica gel plate, simultaneously sampling standard liquor on the same high-efficiency thin-layer silica gel plate, adding a developing solvent in a developing cylinder for pre-balancing, subsequently putting the high-efficiency thin-layer silica gel plate into the developing cylinder for secondary development, taking out and airing; (2) dyeing through a baking process; (3) performing a thin-layer chromatographic scanning process through a thin-layer scanner to obtain an Rf (Retardation Factor Value) and a peak area integral value of the muramic acid, simultaneously calculating a regression equation of the relationship of the quality of the standard liquor and the peak area integral value, and then calculating the mass concentration of the muramic acid in the sample liquor according to the peak area integral value of the sample liquor. The invention also provides a method for extracting the muramic acid from corynebacterium glutamicum dregs, and purified muramic acid can be obtained through various treatments such as washing, degreasing, acidolysis and extraction on the corynebacterium dregs.
Description
Technical field
The present invention relates to a kind of method of extracting muramic acid from corynebacterium glutamicum bacterium slag, and the thin-layer chromatography detection method of muramic acid.
Background technology
Muramic acid is the constituent of peptide glycan in bacteria cell wall.
Corynebacterium glutamicum (Corynebacterium glutamicum) is aerobic bacteria, bar-shaped, sometimes microbend, the blunt circle in two ends, branch not, single or become the Eight characters to arrange, 0.7～0.9 micron * 1.0～2.5 microns of thalline, Gram-positive, without gemma, can be used for microbial fermentation engineering and produces glutamic acid also and then produce sodium glutamate (monosodium glutamate).The useless wine with dregs bacterium slag of glutamic acid fermentation is the main secondary product of monosodium glutamate industry, and annual output can reach tons up to a million, mainly as fertilizer and feed processing, cheap.The useless wine with dregs bacterium slag of the glutamic acid fermentation of take is raw material, detects and extract purifying muramic acid, can significantly promote comprehensive utilization value, improves the economic and social benefits.Have not yet to see and take the useless wine with dregs bacterium slag of glutamic acid fermentation and extract the report of purifying muramic acid as raw material.Summary of the invention
The invention provides a kind of method of extracting purifying muramic acid from corynebacterium glutamicum bacterium slag, the present invention also provides a kind of thin-layer chromatography detection method of muramic acid.
The present invention is achieved by the following technical solutions:
A method of extracting muramic acid from corynebacterium glutamicum bacterium slag, step is as follows:
1) get fresh corynebacterium glutamicum bacterium slag (glutamic acid fermentation give up wine with dregs bacterium slag, i.e. raw material bacterium slag), be placed in container, the physiological saline that is first 0.9% by mass concentration is at room temperature (2～5 times) washing bacterium slag repeatedly, to remove foreign odor taste and lyotrope matter, filter, obtain filter residue;
Further, with after physiological saline washing, if filter residue also has foreign odor taste, in filter residue, add distilled water, 55 ℃ of temperature and follow under stirring and wash bacterium slag (can realize by constant temperature blender with magnetic force), wash 2～5 times, filtration, obtains filter residue;
2) by step 1) gained filter residue puts into closed container, the trichloroacetic acid solution that the mass concentration that adds 3～5 times of raw material bacterium slag amounts is 10%, at 45～60 ℃ of temperature, stirring 15min～1h (can realize by constant temperature blender with magnetic force, the preferred middling speed of stirring rate), with cracking thalline, remove LTA, filter, obtain filter residue;
3) by step 2) distilled water washing 1～2 time for gained filter residue, to remove trichloroacetic acid, then use successively methyl alcohol, methyl alcohol-chloroform mixed liquor (methyl alcohol, chloroform by volume 1:1 mix), the chloroform washing degreasing of 2～10 times of raw material bacterium slag amounts, each 2 times; Obtain degreasing filter residue;
4) to step 3) to add concentration in gained degreasing filter residue be the hydrochloric acid of 5～8mol/L, by every gram of raw material bacterium slag, adds 4～10 milliliters of hydrochloric acid, is placed in water-bath, acidolysis 4～8h at 90～100 ℃ of temperature; After acidolysis, obtain the acid hydrolysis solution of brownish black, by acid hydrolysis solution concentrated evaporate to dryness (50～65 ℃) on vacuum rotary evaporator, to fling to hydrochloric acid as far as possible, obtain muramic acid crude product;
Preferably, the described temperature by acid hydrolysis solution concentrated evaporate to dryness on vacuum rotary evaporator is 50～65 ℃;
5) step 4) gained muramic acid crude product, dissolves with distilled water, adds chloroform, and concuss rocks, and standing separatory discards lower floor's organic phase (impurity that contains lipophilicity and both sexes in organic phase), then adds chloroform extraction 1～3 time, obtains upper solution;
Preferably, the consumption of described distilled water is 2～4 times of raw material bacterium slag amount, and chloroform consumption is 5～10 times of distilled water volume;
6) by step 5) gained upper solution evaporation volatilizes, and with methyl alcohol, dissolves, and adds activated charcoal mixing and absorption 1～3 time, becomes colorless transparent to solution, obtains muramic acid solution, and concentrate drying, obtains muramic acid;
Preferably, described activated charcoal is selected model 303 craboraffins, and each activated carbon dosage is 0.5～1.5 times of raw material bacterium slag amount.
A thin-layer chromatography scanning detection method for muramic acid, step is as follows:
1) get detected sample liquid (being the muramic acid solution that said method prepares), point sample on efficient thin-layer silicon offset plate, meanwhile, by the titer point sample of variable concentrations in the efficient thin-layer silicon offset plate of same; Developping agent adds and in expansion cylinder, carries out pre-equilibration, after efficient thin-layer silicon offset plate put into expansion cylinder carry out second outspread, taking-up is dried;
The efficient thin-layer silicon offset plate of GF254 that described efficient thin-layer silicon offset plate is selected 10cm * 10cm;
The point sample amount of described detected sample liquid is 10 μ L/ points;
Described titer is the titer of 5 kinds of variable concentrations, and concentration is respectively 2.0,3.0,4.0,5.0,6.0mg/ml, and point sample amount is 1 μ L/ point;
Described developping agent is methanol-acetone-acetic acid-water-isopropyl alcohol mixed liquor, and wherein, the volume ratio of methyl alcohol, acetone, acetic acid, water, isopropyl alcohol is 4～6:2～4:0.5～1:0.5～1:0.5～2;
The temperature of described developping agent pre-equilibration and silica gel plate expansion process is 25 ℃, and 20 minutes, second outspread was opened up apart from 8cm (being all from initial point meter) at every turn at every turn.
2) ethanol solution of ninhydrin that is 1% by mass concentration is sprayed onto on efficient thin-layer silicon offset plate and toasts dyeing;
Further, the baking temperature of described dyeing is 110 ℃, time 5min;
3) utilize thin-layer chromatogram scanner to carry out thin-layer chromatography scanning, obtain muramic acid Rf (Rf value) and peak area integrated value; Meanwhile, calculate the regression equation (calculating by thin layer chromatograph management software, is conventional means) of the relation of titer quality and peak area integrated value, then the peak area integrated value of liquid calculates the mass concentration of muramic acid in sample liquid per sample.
The present invention, when carrying out thin-layer chromatography, for obtaining effective effect, has carried out many explorations to developping agent, and utilizes different agent combination: acetone-water; Isopropyl alcohol-glacial acetic acid-water; Normal butyl alcohol-toluene-glacial acetic acid-water; Normal butyl alcohol-glacial acetic acid-isopropyl alcohol; Normal butyl alcohol-toluene-glacial acetic acid-water; Isopropyl alcohol: acetic acid: water: methyl alcohol and methyl alcohol: acetone: acetic acid: water: isopropyl alcohol etc.Each combination, all carried out ratio debugging repeatedly, finally, discovery is with methanol-acetone-acetic acid-water-isopropyl alcohol, volume ratio is the developping agent of 4～6:2～4:0.5～1:0.5～1:0.5～2 proportioning and to launch effect while launching 25 ℃ of temperature better, and the volume ratio of methyl alcohol, acetone, acetic acid, water, isopropyl alcohol of take is best during as 5:3:1:1:1.
The method of extracting muramic acid from corynebacterium glutamicum bacterium slag of the present invention, has removed the impurity in bacterium slag, has obtained the muramic acid of purifying.The thin-layer chromatography detection method of muramic acid of the present invention, has the advantages such as simple, quick, accurate, reliable, stable.
Embodiment
Below in conjunction with embodiment, the present invention is further illustrated.
Embodiment 1 extracts muramic acid and carries out thin-layer chromatography detection
Extract: get 5 grams of fresh corynebacterium glutamicum bacterium slags (raw material bacterium slag), be placed in 150ml beaker, first with 0.9% physiological saline, at room temperature repeatedly wash bacterium slag, remove foreign odor taste, filter, obtain filter residue, proceed in 200ml triangular flask, the trichloroacetic acid solution that adds 25ml10%, by constant temperature blender with magnetic force at 55 ℃, suitably under rotating speed, stir 30min, with cracking thalline, remove LTA, filter, remove filtrate, with distilled water, wash residue to wash away trichloroacetic acid, then use successively 50ml methyl alcohol, 50ml methyl alcohol-chloroform mixed liquor (methyl alcohol and chloroform by volume 1:1 mix), 50ml chloroform washing degreasing, each 2 times, obtain degreasing filter residue, the hydrochloric acid 50ml that adds 6mol/L, at 95 ℃ of heating acidolysis 5h of water-bath, obtains the acid hydrolysis solution of brownish black, the acid hydrolysis solution of brownish black, at the concentrated 50 ℃ of evaporates to dryness of Rotary Evaporators, to fling to hydrochloric acid as far as possible, is dissolved with 20ml tri-distilled water, add 100ml chloroform, concuss rocks, and standing separatory, except the impurity of sub-cloud lipophilicity and both sexes, add chloroform extraction 2 times, each chloroform consumption is 100ml, obtains upper solution again, evaporation volatilizes, and with the methyl alcohol of 30ml, dissolves, and adds 5g303 craboraffin mixing and absorption 3 times, becomes colorless transparent to solution, obtains muramic acid solution.
Detect: get muramic acid solution 1ml, dilute 15 times, obtain sample liquid.The titer of preparation muramic acid, concentration is respectively: 0.2,0.3,0.4,0.5,0.6mg/ml.Sample liquid (10 μ L/ select) and titer (1 μ L/ point) are put on the efficient thin-layer silicon offset plate of same GF254 (10cm * 10cm).Developping agent is methanol-acetone-glacial acetic acid-water-isopropyl alcohol mixed liquor, and wherein, the volume ratio of methyl alcohol, acetone, acetic acid, water, isopropyl alcohol is 5:3:1:1:1.Developping agent is added to expansion cylinder (10cm * 12cm * 15cm) pre-equilibration 20 minutes at 25 ℃.Efficient thin layer plate is put into expansion cylinder, at 25 ℃, launches, and when exhibition distance reaches 8cm, silica gel plate plate is taken out and is dried.And then put back to expansion cylinder secondary (from initial point meter) and launch 8cm, after launching to finish, silica gel plate plate is taken out and dried.The dyeing of the ethanol solution of ninhydrin that is 1% by mass concentration spray plate, is then placed at 110 ℃ and toasts 5 minutes.The point that sample liquid and standard items are corresponding is dyed redness.With camag3 type thin-layer chromatogram scanner, absorbing wavelength scanning with 492nm, obtain peak area integrated value and muramic acid migration value Rf is 0.84, the regression equation of the relation of thin-layer chromatogram scanner software winCATS1.4.1 statistical computation peak area integrated value and titer quality, correlativity Y=16.854X+4572.2, r=0.9991RSD=1.53%.After testing and calculate, obtaining muramic acid content in corynebacterium glutamicum bacterium slag is 1.10g/100g.
Embodiment 2 extracts muramic acid and carries out thin-layer chromatography detection
Extract: get 5 grams of fresh corynebacterium glutamicum bacterium slags (raw material bacterium slag), be placed in 150ml beaker, first with 0.9% physiological saline, at room temperature repeatedly wash bacterium slag, remove foreign odor taste, filter, obtain filter residue, proceed in 200ml triangular flask, the trichloroacetic acid solution that adds 15ml10%, by constant temperature blender with magnetic force at 45 ℃, suitably under rotating speed, stir 15 minutes, with cracking thalline, remove LTA, filter, remove filtrate, with distilled water, wash residue to wash away trichloroacetic acid, then use successively 10ml methyl alcohol 10ml methyl alcohol-chloroform mixed liquor (methyl alcohol and chloroform by volume 1:1 mix), 10ml chloroform washing degreasing, each 2 times, obtain degreasing filter residue, the hydrochloric acid 20ml that adds 5mol/L, at 90 ℃ of heating acidolysis 4h of water-bath, obtains the acid hydrolysis solution of brownish black, the acid hydrolysis solution of brownish black, at the concentrated 50 ℃ of evaporates to dryness of Rotary Evaporators, to fling to hydrochloric acid as far as possible, is dissolved with 10ml tri-distilled water, add 50ml chloroform, concuss rocks, and standing separatory, except the impurity of sub-cloud lipophilicity and both sexes, add chloroform extraction 2 times, each chloroform consumption is 50ml, obtains upper solution again, evaporation volatilizes, and with the methyl alcohol of 30ml, dissolves, and adds 2.5g303 craboraffin mixing and absorption 3 times, becomes colorless transparent to solution, obtains muramic acid solution.
Detect: get muramic acid solution 1ml, dilute 15 times, obtain sample liquid.The titer of preparation muramic acid, concentration is respectively: 0.2,0.3,0.4,0.5,0.6mg/ml.Sample liquid (10 μ L/ select) and titer (1 μ L/ point) are put on the efficient thin-layer silicon offset plate of same GF254 (10cm * 10cm).Developping agent is methanol-acetone-glacial acetic acid-water-isopropyl alcohol mixed liquor, and wherein, the volume ratio of methyl alcohol, acetone, acetic acid, water, isopropyl alcohol is 4:2:0.5:0.5:0.5.Developping agent is added to expansion cylinder (10cm * 12cm * 15cm) pre-equilibration 20 minutes at 25 ℃.Efficient thin layer plate is put into expansion cylinder, at 25 ℃, launches, and when exhibition distance reaches 8cm, silica gel plate plate is taken out and is dried.And then put back to expansion cylinder secondary (from initial point meter) and launch 8cm, after launching to finish, silica gel plate plate is taken out and dried.The dyeing of the ethanol solution of ninhydrin that is 1% by mass concentration spray plate, is then placed at 110 ℃ and toasts 5 minutes.The point that sample liquid and standard items are corresponding is dyed redness.With camag3 type thin-layer chromatogram scanner, absorbing wavelength scanning with 492nm, obtain peak area integrated value and muramic acid migration value Rf is 0.84, the regression equation of the relation of thin-layer chromatogram scanner software winCATS1.4.1 statistical computation peak area integrated value and titer quality, correlativity Y=4113.9915+30.0484*X, r=0.99953RSD=3.71%.After testing and calculate, obtaining muramic acid content in corynebacterium glutamicum bacterium slag is 1.16g/100g.
Embodiment 3 extracts muramic acid and carries out thin-layer chromatography detection
Extract: get 5 grams of fresh corynebacterium glutamicum bacterium slags (raw material bacterium slag), be placed in 150ml beaker, first with 0.9% physiological saline, at room temperature repeatedly wash bacterium slag, remove foreign odor taste, filter, obtain filter residue, proceed in 200ml triangular flask, the trichloroacetic acid solution that adds 20ml10%, by constant temperature blender with magnetic force at 60 ℃, suitably under rotating speed, stir 1 hour, with cracking thalline, remove LTA, filter, remove filtrate, with distilled water washing residue 2 times to wash away trichloroacetic acid, then use successively 30ml methyl alcohol, 30ml methyl alcohol-chloroform mixed liquor (methyl alcohol and chloroform by volume 1:1 mix), 30ml chloroform washing degreasing, each 2 times, obtain degreasing filter residue, the hydrochloric acid 30ml that adds 8mol/L, at 100 ℃ of heating acidolysis 8h of water-bath, obtains the acid hydrolysis solution of brownish black, the acid hydrolysis solution of brownish black, at the concentrated 65 ℃ of evaporates to dryness of Rotary Evaporators, to fling to hydrochloric acid as far as possible, is dissolved with 15ml tri-distilled water, add 90ml chloroform, concuss rocks, and standing separatory, except the impurity of sub-cloud lipophilicity and both sexes, add chloroform extraction 2 times, each chloroform consumption is 90ml, obtains upper solution again, evaporation volatilizes, and with the methyl alcohol of 20ml, dissolves, and adds 7.5g303 craboraffin mixing and absorption 3 times, becomes colorless transparent to solution, obtains muramic acid solution.
Detect: get muramic acid solution 1ml, dilute 15 times, obtain sample liquid.The titer of preparation muramic acid, concentration is respectively: 0.2,0.3,0.4,0.5,0.6mg/ml.Sample liquid (10 μ L/ select) and titer (1 μ L/ point) are put on the efficient thin-layer silicon offset plate of same GF254 (10cm * 10cm).Developping agent is methanol-acetone-glacial acetic acid-water-isopropyl alcohol mixed liquor, and wherein, the volume ratio of methyl alcohol, acetone, acetic acid, water, isopropyl alcohol is 6:4:0.9:0.8:2.Developping agent is added to expansion cylinder (10cm * 12cm * 15cm) pre-equilibration 20 minutes at 25 ℃.Efficient thin layer plate is put into expansion cylinder, at 25 ℃, launches, and when exhibition distance reaches 8cm, silica gel plate plate is taken out and is dried.And then put back to expansion cylinder secondary (from initial point meter) and launch 8cm, after launching to finish, silica gel plate plate is taken out and dried.The dyeing of the ninhydrin ethanolic solution that is 1% by mass concentration spray plate, is then placed at 110 ℃ and toasts 5 minutes.The point that sample liquid and standard items are corresponding is dyed redness.With camag3 type thin-layer chromatogram scanner, with the absorbing wavelength scanning of 492nm, obtain peak area integrated value and muramic acid migration value Rf is 0.84, use gained in embodiment 1 to be related to regression equation, after testing and calculate, obtaining muramic acid content in corynebacterium glutamicum bacterium slag is 1.13g/100g.
Claims (9)
1. a thin-layer chromatography scanning detection method for muramic acid, is characterized in that: step is as follows:
1) get detected sample liquid, point sample on efficient thin-layer silicon offset plate, meanwhile, by the titer point sample of variable concentrations in the efficient thin-layer silicon offset plate of same; Developping agent adds and in expansion cylinder, carries out pre-equilibration, after efficient thin-layer silicon offset plate put into expansion cylinder carry out second outspread, taking-up is dried;
Described developping agent is methanol-acetone-acetic acid-water-isopropyl alcohol mixed liquor, and wherein, the volume ratio of methyl alcohol, acetone, acetic acid, water, isopropyl alcohol is 4～6:2～4:0.5～1:0.5～1:0.5～2;
2) ethanol solution of ninhydrin that is 1% by mass concentration is sprayed onto on efficient thin-layer silicon offset plate and toasts dyeing;
3) utilize thin-layer chromatogram scanner to carry out thin-layer chromatography scanning, obtain muramic acid Rf (Rf value) and peak area integrated value; Meanwhile, calculate the regression equation of the relation of titer quality and peak area integrated value, then the peak area integrated value of liquid calculates the mass concentration of muramic acid in sample liquid per sample.
2. the thin-layer chromatography scanning detection method of muramic acid according to claim 1, is characterized in that: described efficient thin-layer silicon offset plate is selected the efficient thin-layer silicon offset plate of GF254.
3. the thin-layer chromatography scanning detection method of muramic acid according to claim 1, is characterized in that: described titer is the titer of 5 kinds of variable concentrations, and concentration is respectively 2.0,3.0,4.0,5.0,6.0mg/ml.
4. the thin-layer chromatography scanning detection method of muramic acid according to claim 1, is characterized in that: the volume ratio of described methyl alcohol, acetone, acetic acid, water, isopropyl alcohol is 5:3:1:1:1.
5. the thin-layer chromatography scanning detection method of muramic acid according to claim 1, is characterized in that: the temperature of described developping agent pre-equilibration and silica gel plate expansion process is 25 ℃, time 20min.
6. the thin-layer chromatography scanning detection method of muramic acid according to claim 1, is characterized in that: the baking temperature of described baking dyeing is 110 ℃, time 5min.
7. the thin-layer chromatography scanning detection method of muramic acid according to claim 1, is characterized in that: described sample liquid prepares by the following method:
1) get fresh corynebacterium glutamicum bacterium slag (raw material bacterium slag), the physiological saline that is first 0.9% by mass concentration at room temperature repeatedly washs bacterium slag, to remove foreign odor taste and lyotrope matter, filters, and obtains filter residue;
2) step 1) gained filter residue, the trichloroacetic acid solution that the mass concentration that adds 3～5 times of raw material bacterium slag amounts is 10%, at 45～60 ℃ of temperature, stirs 15min～1h, filters, and obtains filter residue;
3) by step 2) distilled water washing 1～2 time for gained filter residue, to remove trichloroacetic acid, then, successively with methyl alcohol, methyl alcohol-chloroform mixed liquor, the chloroform washing degreasing of 2～10 times of filter residue quality, each 2 times; Obtain degreasing filter residue;
4) to step 3) to add concentration in gained degreasing filter residue be the hydrochloric acid of 5～8mol/L, by every gram of raw material filter residue, adds 4～10 milliliters of hydrochloric acid, is placed in water-bath, acidolysis 4～8h at 90～100 ℃ of temperature; After acidolysis, obtain the acid hydrolysis solution of brownish black, by the concentrated evaporate to dryness of acid hydrolysis solution, obtain muramic acid crude product;
5) step 4) gained muramic acid crude product, dissolves with distilled water, adds chloroform, and concuss rocks, and standing separatory discards lower floor's organic phase, then adds chloroform extraction 1～3 time, obtains upper solution;
6) by step 5) gained upper solution evaporation volatilizes, and with methyl alcohol, dissolves, and adds activated charcoal mixing and absorption 1～3 time, becomes colorless transparent to solution, obtains sample liquid.
8. from corynebacterium glutamicum bacterium slag, extract a method for muramic acid, it is characterized in that: step is as follows:
1) get fresh corynebacterium glutamicum bacterium slag (raw material bacterium slag), the physiological saline that is first 0.9% by mass concentration at room temperature repeatedly washs bacterium slag, to remove foreign odor taste and lyotrope matter, filters, and obtains filter residue;
2) step 1) gained filter residue, the trichloroacetic acid solution that the mass concentration that adds 3～5 times of raw material filter residue quality is 10%, at 45～60 ℃ of temperature, stirs 15min～1h, filters, and obtains filter residue;
3) by step 2) distilled water washing 1～2 time for gained filter residue, to remove trichloroacetic acid, then, successively with methyl alcohol, methyl alcohol-chloroform mixed liquor, the chloroform washing degreasing of 2～10 times of raw material bacterium slag amounts, each 2 times; Obtain degreasing filter residue;
4) to step 3) to add concentration in gained degreasing filter residue be the hydrochloric acid of 5～8mol/L, by every 1 gram of raw material bacterium slag, adds 4～10 milliliters of hydrochloric acid, is placed in water-bath, acidolysis 4～8h at 90～100 ℃ of temperature; After acidolysis, obtain the acid hydrolysis solution of brownish black, by the concentrated evaporate to dryness of acid hydrolysis solution, obtain muramic acid crude product;
5) step 4) gained muramic acid crude product, dissolves with distilled water, adds chloroform, and concuss rocks, and standing separatory discards lower floor's organic phase, then adds chloroform extraction 1～3 time, obtains upper solution;
6) by step 5) gained upper solution evaporation volatilizes, and with methyl alcohol, dissolves, and adds activated charcoal mixing and absorption 1～3 time, becomes colorless transparent to solution, obtains muramic acid solution, and concentrate drying, obtains muramic acid.
9. according to the method for muramic acid of extracting from corynebacterium glutamicum bacterium slag described in right pressure ball 8, it is characterized in that: described activated charcoal is selected model 303 craboraffins.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410188150.XA CN103954726B (en) | 2014-05-06 | 2014-05-06 | Method and the detection method of muramic acid is extracted from corynebacterium glutamicum bacterium slag |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410188150.XA CN103954726B (en) | 2014-05-06 | 2014-05-06 | Method and the detection method of muramic acid is extracted from corynebacterium glutamicum bacterium slag |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN103954726A true CN103954726A (en) | 2014-07-30 |
| CN103954726B CN103954726B (en) | 2015-11-18 |
Family
ID=51332026
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201410188150.XA Expired - Fee Related CN103954726B (en) | 2014-05-06 | 2014-05-06 | Method and the detection method of muramic acid is extracted from corynebacterium glutamicum bacterium slag |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103954726B (en) |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106188168A (en) * | 2016-07-18 | 2016-12-07 | 山东师范大学 | Utilize the method that the dreg producing glutamic acid prepares glucosamine hydrochloride |
| CN106188171A (en) * | 2016-07-18 | 2016-12-07 | 山东师范大学 | A kind of method utilizing the dreg producing glutamic acid to prepare 3-O-.alpha.-carboxyethyl-D-glucosamine. |
| CN106279308A (en) * | 2016-07-18 | 2017-01-04 | 山东师范大学 | The method utilizing the dreg separation 3-O-.alpha.-carboxyethyl-D-glucosamine. producing glutamic acid |
| CN110250319A (en) * | 2019-07-10 | 2019-09-20 | 福州大学 | A kind of preparation of novel corynebacterium glutamicum waste residue probiotics |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20050191732A1 (en) * | 1999-06-25 | 2005-09-01 | Basf Aktiengesellschaft | Corynebacterium glutamicum genes encoding proteins involved in homeostasis and adaptation |
-
2014
- 2014-05-06 CN CN201410188150.XA patent/CN103954726B/en not_active Expired - Fee Related
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20050191732A1 (en) * | 1999-06-25 | 2005-09-01 | Basf Aktiengesellschaft | Corynebacterium glutamicum genes encoding proteins involved in homeostasis and adaptation |
Non-Patent Citations (3)
| Title |
|---|
| ARSOLD S.BLEIWEIS等: "Improved Separation of Components of Streptococcal Cell", 《ANALYTICAL BIOCHEMISTRY》 * |
| GUNNAR KLEPPE等: "The Specificity Requirements of Bacteriophage T4 Lysozyme Involvement of N-Acetamido Groups", 《EUR J BIOCHEM.》 * |
| I KAWAMOTO等: "Cell wall composition of Micromonospora olivoasterospora, Micromonospora sagamiensis, and related organisms", 《JOURNAL OF BACTERIOLOGY》 * |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106188168A (en) * | 2016-07-18 | 2016-12-07 | 山东师范大学 | Utilize the method that the dreg producing glutamic acid prepares glucosamine hydrochloride |
| CN106188171A (en) * | 2016-07-18 | 2016-12-07 | 山东师范大学 | A kind of method utilizing the dreg producing glutamic acid to prepare 3-O-.alpha.-carboxyethyl-D-glucosamine. |
| CN106279308A (en) * | 2016-07-18 | 2017-01-04 | 山东师范大学 | The method utilizing the dreg separation 3-O-.alpha.-carboxyethyl-D-glucosamine. producing glutamic acid |
| CN106188171B (en) * | 2016-07-18 | 2018-06-08 | 山东师范大学 | A kind of utilize produces the method that the bacteria residue of glutamic acid prepares muramic acid |
| CN106188168B (en) * | 2016-07-18 | 2018-10-02 | 山东师范大学 | The method for preparing aminoglucose hydrochloride using the bacteria residue of production glutamic acid |
| CN106279308B (en) * | 2016-07-18 | 2018-10-26 | 山东师范大学 | The method for detaching muramic acid using the bacteria residue of production glutamic acid |
| CN110250319A (en) * | 2019-07-10 | 2019-09-20 | 福州大学 | A kind of preparation of novel corynebacterium glutamicum waste residue probiotics |
Also Published As
| Publication number | Publication date |
|---|---|
| CN103954726B (en) | 2015-11-18 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN103472160B (en) | Liquid chromatography-mass spectrometry method for rapidly detecting content of beta-agonists and sample pretreatment method | |
| CN103954726B (en) | Method and the detection method of muramic acid is extracted from corynebacterium glutamicum bacterium slag | |
| CN103512984B (en) | Sample pretreatment method for detecting different clenbuterol residuals | |
| CN102898566B (en) | Preparation method of metal organic framework molecularly imprinted polymer used for enriching trace amounts of Metolcarb | |
| CN104749262B (en) | Method of rapidly determining fluoroquinolone type medicines in faeces of livestock and poultry | |
| CN108760935B (en) | Method for extracting and determining sulfonamide antibiotics in plants | |
| CN103936837A (en) | Method used for high-efficient purification of pneumocandins B0 | |
| CN102503879A (en) | Preparation method for fullerene amino acid derivative | |
| CN106198832A (en) | A kind of quality of production control method of intensified loquet distillate | |
| CN102735512B (en) | Method for extracting biotin from Antarctic krill and high-efficiency thin layer scanning analysis method | |
| CN104098465A (en) | Technological method for extraction of protocatechuic acid from Blumea riparia (Bl.) DC | |
| CN104977367B (en) | Method for detecting 20 kinds of beta-stimulant drug residues in livestock urine | |
| CN100388940C (en) | Quality control method of Chinese medicinal preparation for treating child hyperpyrexia | |
| CN110551137B (en) | Method for extracting and purifying glabridin and application of glabridin in cosmetics | |
| CN100355775C (en) | Polypeptide of ascidicea and preparation thereof | |
| CN101869630A (en) | Measurement method for content of atropine sulfate in traditional Chinese medicine suppository containing belladonna liquid extract | |
| CN104165952A (en) | Method for detecting banned azo dyes in printing ink, coatings and paint | |
| CN102830183A (en) | Sample pre-treatment method for determining endogenesis hormone content of towel gourd seeds by adopting HPLC (High Performance Liquid Chromatography) | |
| CN102178698A (en) | Method for controlling quality of Chinese patent medicine | |
| CN101433824A (en) | Method for extracting sulfabenzpyrazine from animal sample and special immune affinity sorbent thereof | |
| CN103611157A (en) | Panax japonicus saponin as well as preparation method and application thereof | |
| CN107505417A (en) | The detection method of clenbuterol hydrochloride in fresh meat and meat products | |
| CN104833564A (en) | Sample pretreatment method suitable for detection of basic orange II in bean product | |
| CN103344726B (en) | Method for extraction of dicyandiamide component in dairy product | |
| CN105181875B (en) | Method and sample pretreatment method for detecting content of chloramphenicol in silkworm faeces |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20151118 Termination date: 20160506 |