CN102680424A - Determination method for trehalose contentin activated sludge - Google Patents
Determination method for trehalose contentin activated sludge Download PDFInfo
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- CN102680424A CN102680424A CN2011100639079A CN201110063907A CN102680424A CN 102680424 A CN102680424 A CN 102680424A CN 2011100639079 A CN2011100639079 A CN 2011100639079A CN 201110063907 A CN201110063907 A CN 201110063907A CN 102680424 A CN102680424 A CN 102680424A
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Abstract
The invention relates to a determination method for the trehalose content in activated sludge, belonging to the technical field of biological sewage treatment in environmental engineering. In order to realize the application of the trehalose in the technical field of biological sewage treatment, the trehalose in activated sludge microbial cells is subjected to combined extraction of trichloroacetic acid and ultrasound on the basis of an anthrone colorimetry method, and the trehalose is dewatered to generate furfural and a derivative thereof while encountering with concentrated sulfuric acid, wherein the derivative has a reaction with anthranone, a solution is blue after the reaction, and the content of the trehalose in the activated sludge is measured at a 610nm point by colorimetric estimation. The method has the advantages of safety, reliability, simplicity, easy application, lower cost and high measurement precision and can be used for realizing accurate and quick measurement of the trehalose in the activated sludge.
Description
Technical field
The invention belongs to the dirty water living creature processing technique field in the environmental engineering subject, be specifically related to a kind of assay method of active sludge content of trehalose.
Background technology
Trehalose (Trehalose) is a kind of non-reducing disaccharide, by 2 glucose molecules with α, α, 1, the 1-glycosidic bond is formed by connecting, molecular formula is C
12H
22O
112H
2O, relative molecular weight are 378.33, white crystals, and chemical property is extremely stable, and is nontoxic, can caramelization.Research shows that trehalose has magical protective effect to biosome, is a kind of important metabolic product that biosome reply external environment stimulates.The content of trehalose that exists in the degeneration-resistant tolerance that biosome shows under rugged surroundings and their bodies is closely related, and biosome can be resisted extraneous injury through regulating the compound body intracellular trehalose.At present, biosome trehalose analysis determining technology has been widely used in industrial fields such as food, medicine, cosmetics, but in field of environment engineering, is not applied so far.
Because active sludge is bearing the impact that brings because of load variations, variation of ambient temperature, the variation of pH value etc. constantly in the sewage biological treatment system of actual motion; This impact also can be reflected on the content of trehalose level of active sludge in the biologic process for treating sewage; Therefore through assay determination active sludge microorganism cell intracellular trehalose content; Can characterize out active sludge exactly and receive the degree that external environment stimulates, and can seek the technical method that utilizes foreign aid's trehalose to strengthen the active sludge stress ability.
Summary of the invention
Technical matters to be solved by this invention is; Owing to still do not measure the technical method of active sludge trehalose at present in the biological sewage treatment field; Can't research and analyse the internal relation between trehalose and the active sludge; Therefore the present invention has designed the assay method of trehalose in the active sludge, to realize the application of trehalose in field of environment engineering.
The principle of this method is to adopt the anthrone colourimetry to be the basis; The intracellular trehalose of active sludge microorganism is carried out trichloroacetic acid-ultrasonic in combination to be extracted; Trehalose is met concentrated sulphuric acid dehydration and is generated furfural and derivant thereof, and this derivant and anthrone are reacted, and reaction back solution is blue-green; In the 610nm place absorption maximum is arranged, but the content of trehalose in the colorimetric estimation active sludge herein.
To achieve these goals, the present invention takes following technical step to carry out:
The active sludge intermixture that in the 10mL centrifuge tube, adds 0.8mL places hydro-extractor, centrifugal 5min under 4500r/min; With distilled water flushing, the centrifugal gained active sludge of stirring, centrifugal 5min under 4500r/min, 3 times repeatedly; Remove trichloroacetic acid (TCA) solution that adds 6mL, 0.4mol/L after the centrifuged supernatant, be placed on sonicated 80min in the ultrasonication machine, centrifugal 8min under 4500r/min then, supernatant is the trehalose extract; Getting anthrone reagent 10mL joins in the 50mL fermentation tube; Add gained trehalose extract 5mL in the aforementioned centrifugal process, and immerse in the cooling bath rapidly and cool off, place boiling water bath 6min then; Be cooled to room temperature after the taking-up rapidly, under the 610nm wavelength, measure absorbance with the 1cm cuvette; Simultaneously the aforementioned mud that in ultrasonic centrifugal treating process, produces is weighed behind (105 ± 1) ℃ following oven dry 1h; At last according to absorbance and mud weighing results calculated activity mud cell intracellular trehalose content.
The computing formula of active sludge cell intracellular trehalose content is:
C---the content (mgL of trehalose in every milligram of active sludge in the mixed liquor
-1/ mgMLSS);
K---slope of standard curve;
W---mud net weight (mg);
D
610---the absorbance of wavelength 610nm place's trehalose and anthrone reagent chromophoric solution.
Good effect of the present invention is:
1, this method is safe and reliable, simple, can realize the fast measuring of trehalose in the active sludge;
2, the used instrument and equipment of this method is the conventional instrument in laboratory, need not large-scale checkout equipment, has the characteristics of inexpensive easy realization, and analysis cost is lower;
3, this method has reduced because of sludge content changes the error that is caused significantly, has improved the mensuration computational accuracy, and it measures the result and the national standard sample carries out the quadrature comparison, and the data homing rate reaches 99.5%.
Embodiment
1, typical curve is drawn
0.1g trehalose standard items are water-soluble, and constant volume is configured to standard solution to 100mL.Accurately absorption standard aqueous trehalose 0,0.5,1.0,1.5,2.0,2.5,3.0mL place the 100mL volumetric flask, and constant volume shakes up (be equivalent to respectively 0,5.0,10.0,15.0,20.0,25.0,30.0mg/L aqueous trehalose).Getting anthrone reagent 10mL joins in the 50mL fermentation tube; Respectively add the trehalose standard solution 5mL of variable concentrations from the bottle wall carefully then, acutely shake up, and immerse in the cooling bath rapidly and cool off; Place boiling water bath 6min then; Be cooled to room temperature after the taking-up rapidly, use the 1cm cuvette, under the 610nm wavelength, measure absorbance.With the absorbance is horizontal ordinate, and sugar concentration (mg/L) is an ordinate, the drawing standard curve.
2, the extraction of active sludge trehalose
Trehalose in the active sludge is synthetic to be stored in the microbial cell, must after extracting, can measure.Consider the feasibility and the singularity of microbial cell broken wall, can take the ultrasonic disruption machine of certain ultrasound intensity to extract.
3, the mensuration of content of trehalose in the active sludge
The active sludge intermixture that in the 10mL centrifuge tube, adds 0.8mL places hydro-extractor, centrifugal 5min under 4500r/min; With distilled water flushing, the centrifugal gained active sludge of stirring, centrifugal 5min under 4500r/min, 3 times repeatedly; Remove trichloroacetic acid (TCA) solution that adds 6mL, 0.4mol/L after the centrifuged supernatant, be placed on sonicated 80min in the ultrasonication machine, centrifugal 8min under 4500r/min then, supernatant is the trehalose extract; Get anthrone reagent 10mL and join in the 50mL fermentation tube, add gained trehalose extract 5mL in the aforementioned centrifugal process along tube wall carefully, acutely shake up; And immerse in the cooling bath rapidly and cool off; Place boiling water bath 6min then, be cooled to room temperature after the taking-up rapidly, use the 1cm cuvette; Under the 610nm wavelength, measure absorbance, simultaneously the mud that produces in the aforementioned centrifugal process is weighed behind (105 ± 1) ℃ following oven dry 1h; At last according to absorbance and mud weighing results calculated activity mud cell intracellular trehalose content.
Claims (1)
1. use characteristic of the present invention is: in the mensuration process of active sludge content of trehalose; The active sludge intermixture of getting 0.8mL places hydro-extractor centrifugal, with distilled water flushing, to stir centrifugal gained active sludge centrifugal again, adds trichloroacetic acid solution after removing supernatant; Through centrifugal after the sonicated; Get its supernatant and add anthrone reagent, cooling is placed on 6min in the boiling water bath, again cooling; Under the 610nm wavelength, measure absorbance with the 1cm cuvette, at last according to absorbance and mud weighing results calculated activity mud cell intracellular trehalose content.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
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| CN2011100639079A CN102680424A (en) | 2011-03-17 | 2011-03-17 | Determination method for trehalose contentin activated sludge |
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|---|---|---|---|
| CN2011100639079A CN102680424A (en) | 2011-03-17 | 2011-03-17 | Determination method for trehalose contentin activated sludge |
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| CN102680424A true CN102680424A (en) | 2012-09-19 |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109596550A (en) * | 2018-12-24 | 2019-04-09 | 苏州科铭生物技术有限公司 | A kind of content of trehalose assay kit and its method based on spectrophotometry |
| CN112167617A (en) * | 2020-06-29 | 2021-01-05 | 武汉林宝莱生物科技有限公司 | Female anti-aging whitening pill and preparation method thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH11197686A (en) * | 1998-01-20 | 1999-07-27 | Mitsubishi Heavy Ind Ltd | Method for storing activated sludge |
| CN101650303A (en) * | 2009-09-17 | 2010-02-17 | 无锡健特药业有限公司 | Method for measuring general ginsenoside by anthrone colorimetric method |
-
2011
- 2011-03-17 CN CN2011100639079A patent/CN102680424A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH11197686A (en) * | 1998-01-20 | 1999-07-27 | Mitsubishi Heavy Ind Ltd | Method for storing activated sludge |
| CN101650303A (en) * | 2009-09-17 | 2010-02-17 | 无锡健特药业有限公司 | Method for measuring general ginsenoside by anthrone colorimetric method |
Non-Patent Citations (2)
| Title |
|---|
| 李艳玲 等: "蒽酮-硫酸法测定海藻糖含量显色条件的改进", 《食品工业科技》 * |
| 毛若琳 等: "活性污泥中脱氧核糖核酸的提取和测定", 《建筑技术通讯(给水排水)》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109596550A (en) * | 2018-12-24 | 2019-04-09 | 苏州科铭生物技术有限公司 | A kind of content of trehalose assay kit and its method based on spectrophotometry |
| CN112167617A (en) * | 2020-06-29 | 2021-01-05 | 武汉林宝莱生物科技有限公司 | Female anti-aging whitening pill and preparation method thereof |
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Application publication date: 20120919 |