AU2020101061A4 - A Method for Determination of Soluble Sugar Content in Stem Samples of Sweet Sorghum - Google Patents
A Method for Determination of Soluble Sugar Content in Stem Samples of Sweet Sorghum Download PDFInfo
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- AU2020101061A4 AU2020101061A4 AU2020101061A AU2020101061A AU2020101061A4 AU 2020101061 A4 AU2020101061 A4 AU 2020101061A4 AU 2020101061 A AU2020101061 A AU 2020101061A AU 2020101061 A AU2020101061 A AU 2020101061A AU 2020101061 A4 AU2020101061 A4 AU 2020101061A4
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- G—PHYSICS
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/84—Systems specially adapted for particular applications
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
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- G—PHYSICS
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
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Abstract
The invention provides a method for measuring soluble sugar content of sweet sorghum
stem dry samples, which comprises the following steps of: preparing reagents; Sample
determination; Using colorimetry to determine the absorbance of compounds to judge the sugar
content of sweet sorghum stalks. Phenol sulfuric acid method is a simple method for
determination of soluble total sugar, with cheap reagents, less experimental instruments and
higher sensitivity. It is basically not affected by the presence of protein in the experimental
process, and the generated color can be stabilized for about 3h, which is convenient for
experimentaloperation.
Description of Drawings
0.8
y 0.0056x + 0.0163
0.6 - R2=0.9929
0.4
0.2
0.0
15 35 55 75 95 115
Different Sucrose Concentrations (pg)
Figure 1
3.0 -
Dry Sample
Fresh Sample
2.5
2.0
0
1.1
0.
10/
Description
Description of Drawings
0.8
y 0.0056x + 0.0163 0.6 - R2=0.9929
0.4
0.2
0.0 15 35 55 75 95 115 Different Sucrose Concentrations (pg)
Figure 1
Dry Sample 3.0 - Fresh Sample
2.5
2.0
1.1
0.
10/
Descriptions
A Method for Determination of Soluble Sugar Content in Stem Samples of Sweet Sorghum
Technical Field The invention relates to the technical field of detection of soluble sugar content in sweet sorghum stem samples, in particular to a method for determining soluble sugar content in sweet sorghum stem samples.
Background Technology Sweet sorghum is widely used. It not only produces grain, but also sugar and syrup. It can also be used as wine, alcohol and monosodium glutamate. Fiber can also be used to make paper. Sweet sorghum has been planted for a long time as a forage and sugar crop. Sweet sorghum is stored in a large amount during harvest, which makes it difficult to process it all in a short time, while fresh samples are not easy to store, which easily results in loss of sweet sorghum. Therefore, fresh samples of sweet sorghum stalks drying can be stored for a long time. However, what is the change of sugar content in the stem after the sample is dried. It is necessary to establish a stable and reliable method for determination of soluble sugar content in the stem of sweet sorghum.
Invention Summary The invention aims to provide a method for measuring the content of soluble sugar in the stem sample of sweet sorghum so as to solve the technical problem.
In order to solve the technical problem, the invention adopts the following technical scheme to realize:
The invention relates to a method for measuring soluble sugar content of sweet sorghum stem dry samples, which is characterized by comprising the following steps:
I. Reagent Configuration
A. 9g/ml phenol solution: Dissolve phenol crystal with a density of 1.017g/ml in 60C water bath, and add 88.496ml phenol liquid into a volumetric flask containing 10ml distilled water;
B. 10mg/ml sucrose standard solution: Weigh lg of sucrose, add 5ml of distilled water into a 100ml volumetric flask for pre-dissolution, slowly add 0.25ml of concentrated sulfuric acid, shake well immediately, place at room temperature for 30s, continue to slowly add 0.25ml of
Descriptions
concentrated sulfuric acid, shake well immediately, place at room temperature for 1min, and fix the volume with distilled water for 100 ml;
C. 100 p g/ml sucrose solution: Suck 1ml of 10mg/ml sucrose standard solution and fix the volume to 100ml;
II. Sample Determination
A. Standard curve making:
Al. Preparation of substrate reaction solution: Prepare 7 branches 20ml calibration test tubes, numbered: 0-7 in sequence, with number 0 as control tube and number 1-6 as sample tube. Prepare test samples according to the following table, with a total volume of 2ml:
Scale tube number Reagent 0 1 2 3 4 5 6
100 g/ml sucrose 0.00 0.15 0.30 0.45 0.60 0.75 0.90 solution, unit ml
Distilled water, unit ml 2.00 1.85 1.70 1.55 1.40 1.25 1.10
Sucrose final concentration, unit 0 15 30 45 60 75 90 ptg/2ml
A2. Preparation of reaction solution: 9g/ml phenol solution 1ml is added to 7 branches test tubes in sequence, and after fully mixing, 5ml concentrated sulfuric acid is added to 7 branches test tubes in sequence and quickly, with time interval within 10s;
A3. Sample determination: The reaction solution was left at room temperature for 30min, absorbance was measured at 485nm wavelength, standard curve was drawn, and standard linear equation was obtained.
B. Determination of Soluble Sugar in Stem Samples of Sweet Sorghum
Bl. Sample treatment
Descriptions
Taking one stem of sweet sorghum and drying to constant weight; using plant sample pulverizer to pulverize stem dry sample, sieving with 50 mesh sieve, and packaging in plastic bag for later use;
B2. Extraction of soluble total sugar
Weighing 3 parts of sweet sorghum stem powder, respectively placing in 3 graduated test tubes at 0.3g/ part, 3 graduated test tubes respectively adding 8ml distilled water, sealing with cork, boiling in boiling water for 30min to extract total sugar, filtering the extractive solution with filter paper into a 25ml volumetric flask, continuously rinsing the test tubes 2-3 times, and keeping constant volume for later use;
B3. Determination
( Absorb 0.5ml to 3 Piece graduated test tubes of the above extractive solution, and add 1.5ml distilled water respectively;
( Quickly add 1ml of 9g/ml phenol solution, immediately shake and mix well, and place on the test tube rack for 20 s;
a Slowly add 5ml concentrated sulfuric acid, immediately shake slightly and mix evenly to prevent liquid splashing;
) 10mim is placed in a refrigerator at 4°C for color development.
S The absorbance was measured at 485nm wavelength, and the sugar content was obtained according to the standard curve.
III. Results Calculation
Calculate the sugar content of the sample according to the following formula mx Vtx N Soluble sugar content (mg/g)= 106
Where, m: Sugar content found from standard curve, unit g; Vt is the volume of extractive solution in ml; Vs is the volume of sample taken for determination, in ml; N is the dilution multiple; mo is the sample quality.
Further, in the B2 small step of step 2, during the heating process, the test tube shall be
Descriptions
immersed in boiling water, and the test tube shall not be close to the beaker wall to ensure uniform heating, and the heating time of each test tube shall be kept consistent.
Soluble sugar in sweet sorghum mainly refers to monosaccharide and Oligosaccharide soluble in water. Furfural or methylfurfural produced by dehydration of sugar under the action of concentrated sulfuric acid is easily condensed with phenol to form an orange-red compound (in the range of 10-100mg), the color depth of which is proportional to the content of sugar. Therefore, colorimetry (485nm wavelength, the compound has the maximum absorption peak) can be used to determine the absorbance value of the compound to judge the sugar content of sweet sorghum stalks.
The invention has the beneficial effects that:
The phenol sulfuric acid method determination of soluble total sugar provided by the invention has the advantages of simple method, cheap reagent, fewer experimental instruments and higher sensitivity, is basically not affected by the presence of protein in the experimental process, and the generated color can be stable for about 3h, thus being convenient for experimental operation. It can provide important theoretical basis for the storage of sweet sorghum stalks during production and processing.
Brief Description of Drawings Fig. 1 is a schematic diagram of absorbance standard curve of the invention;
Fig. 2 is a schematic diagram of soluble sugar content of sweet sorghum in different growth stages.
Detailed Description of the Preferred Embodiments
In order to make the technical means, creative features, objectives and effects of the present invention easy to understand, the present invention will be further described with reference to the following specific embodiments, but the following embodiments are only the preferred embodiments of the present invention, not all of them. Based on the embodiments in the embodiments, other embodiments obtained by those skilled in the art under the premise of not making creative work all belong to the protection scope of the present invention. The experimental methods in the following examples are conventional methods unless otherwise specified, and the materials, reagents, etc. used in the following examples can be obtained from commercial channels unless otherwise specified.
Embodiment 1
Descriptions
Instrumentation and equipments
Spectrophotometer, water bath pot, calibration test tube, calibration pipette, 10ml acid burette, scissors, test tube rack, iron rack table, funnel, small beaker, 80tm filter paper.
Reagent configuration
1. 9g/ml phenol solution: Dissolve phenol crystal (density 1.017g/ml) in a water bath at °C, and add 88.496ml phenol liquid into a volumetric flask containing 10ml distilled water, which can be stored for 45 days at room temperature.
2. 10mg/ml sucrose standard solution: Weigh Ig of sucrose (analytically pure), add 5ml of distilled water into a 100ml volumetric flask for pre-dissolution, slowly add 0.25ml of concentrated sulfuric acid, shake well immediately, place at room temperature for 30s, continue to slowly add 0.25ml of concentrated sulfuric acid, shake well immediately, place at room temperature for 1min, and use distilled water for constant volume of100ml.
3. 100[tg/ml sucrose solution: Suck 1 ml of sucrose standard solution (10 mg/ml) and fix the volume to 100ml.
Sample Determination
1. Standard curve making:
1.1 Preparation of substrate reaction solution: prepare 7 branches 20ml calibration test tubes, serial numbers: 0 (control tube), 1-6 (sample tube, repeated twice), and configure test samples (total volume 2ml) according to the following table:
Scale tube number Reagent 0 1 2 3 4 5 6
100jag/ml sucrose 0.00 0.15 0.30 0.45 0.60 0.75 0.90 solution, unit ml
Distilled water, unit ml 2.00 1.85 1.70 1.55 1.40 1.25 1.10
Sucrose final concentration, unit 0 15 30 45 60 75 90 ptg/2ml
Descriptions
1.2 Preparation of reaction solution: 9g/ml phenol solution 1ml is added to 7 branches test tubes in sequence, and after fully mixing, 5ml concentrated sulfuric acid is added to 7 branches test tubes in sequence and quickly (about 10s, time interval should not be too long).
1.3 Sample determination: The reaction solution was left at room temperature for 30min, absorbance was measured at 485nm wavelength, standard curve was drawn, and standard linear equation was obtained (fig. 1).
2. Determination of Soluble Sugar in Stem Samples of Sweet Sorghum
2.1 Sample treatment
Take a stem of sweet sorghum and dry it to constant weight. Use plant sample pulverizer to pulverize stem dry sample, sieve with 50 mesh sieve, and put into plastic bag for later use.
2.2 Extraction of Soluble Total Sugar
Weigh 3 parts (0.3g/ part) of sweet sorghum stalk powder, place it in three graduated test tubes (3 repetitions), respectively add 8ml distilled water, cork seal, boil in boiling water for min to extract total sugar, filter the extract with filter paper to a 25ml volumetric flask, continue rinsing the test tubes for 2-3 times, fix the volume, and set aside.
Remarks: During the heating process, ensure that the test tube is immersed in boiling water, the test tube cannot be close to the beaker wall, ensure uniform heating, and keep the heating time consistent.
2.3 Determination
(D Absorb 0.5ml to 3 branches graduated test tubes of the above treatment extract (repeat for 3 times), and add 1.5ml distilled water respectively;
( Quickly add 1ml phenol solution (9g/ml), shake and mix immediately, and place on the test tube rack for 20s;
a Slowly add 5ml concentrated sulfuric acid, immediately shake slightly and mix evenly to prevent liquid splashing;
@ Put in refrigerator at 4°Cfor 10mim, with color development.
Descriptions
@The absorbance (485nm wavelength) was measured, and the sugar content was obtained according to the standard curve.
Results calculation
Calculate the sugar content of the sample according to the following formula mx Vtx N Soluble sugar content (mg/g)= mx Vsxl1 6
In the formula, m: Sugar content found from standard curve (g); Vt is the volume of the extract (ml); Vs is the sample volume (ml) taken for determination; N is the dilution multiple; mO is the sample quality.
Data Result Analysis
As shown in fig. 2, the stem samples of sweet sorghum in three periods were selected in this experiment. Fresh and dry samples were used respectively to determine the soluble sugar content in the stem of sweet sorghum by the improved phenol sulfuric acid method. The results show (fig. 2): Under this determination method, the soluble sugar content indexes of the dry and fresh sweet sorghum stalks are very close, which indicates that the improved phenol sulfuric acid method can be used to determine soluble sugar in the dry sweet sorghum stalks, which provides an important theoretical basis for the storage of sweet sorghum stalks during production and processing.
The above shows and describes the basic principles, main features and advantages of the present invention. Those skilled in the art should understand that the present invention is not limited by the above-mentioned embodiments. The above-mentioned embodiments and descriptions are only the preferred embodiments of the present invention and are not intended to limit the present invention. Without departing from the spirit and scope of the present invention, various changes and improvements can be made to the present invention, which fall within the scope of the claimed invention. The scope of protection claim by that present invention is defined by the append claims and their equivalents.
Claims (2)
1. The invention relates to a method for measuring soluble sugar content of sweet sorghum stem dry samples, which is characterized by comprising the following steps:
I. Reagent configuration
A. 9g/ml phenol solution: dissolve phenol crystal with density of 1.017g/ml in 60°C water bath, and add 88.496ml phenol liquid into a volumetric flask containing 10ml distilled water;
B. l0mg/ml sucrose standard solution: weigh Ig of sucrose, add 5ml of distilled water into a 100ml volumetric flask for pre-dissolution, slowly add 0.25ml of concentrated sulfuric acid, shake well immediately, place at room temperature for 30s, continue to slowly add 0.25ml of concentrated sulfuric acid, shake well immediately, place at room temperature for 1min, and fix the volume with distilled water for 100 ml;
C. 100 g/ml sucrose solution: suck 1ml of 10mg/ml sucrose standard solution and fix the volume to 100ml;
II. Sample determination
A. Standard curve making:
Al. Preparation of substrate reaction solution: Prepare 7 20ml calibration test tubes, numbered 0-7 in sequence, with number 0 as control tube and number 1-6 as sample tube. Prepare test samples according to the following table, with a total volume of 2ml:
Scale tube number Reagent 0 1 2 3 4 5 6
100 g/ml sucrose 0.00 0.15 0.30 0.45 0.60 0.75 0.90 solution, unit ml
Distilled water, unit ml 2.00 1.85 1.70 1.55 1.40 1.25 1.10
Sucrose final concentration, unit 0 15 30 45 60 75 90 ptg/2ml
A2. Preparation of reaction solution: 9g/ml phenol solution 1ml is added to 7 test tubes in sequence, and after fully mixing, 5ml concentrated sulfuric acid is added to 7 test tubes in
Claims
sequence and rapidly, with time interval within 10s;
A3. Sample determination: the reaction solution was left at room temperature for 30min, absorbance was measured at 485nm wavelength, standard curve was drawn, and standard linear equation was obtained;
B. Determination of Soluble Sugar in Stem Samples of Sweet Sorghum
B1. Sample treatment
Taking one stem of sweet sorghum and drying to constant weight; Pulverizing stem dry samples with a plant sample pulverizer, sieving with 50 mesh sieve, and packaging in plastic bag for later use;
B2. Extraction of Soluble Total Sugar
Weighing 3 parts of sweet sorghum stem powder, respectively placing in 3 graduated test tubes at 0.3g/ part, 3 graduated test tubes respectively adding 8ml distilled water, sealing with cork, boiling in boiling water for 30min to extract total sugar, filtering the extractive solution with filter paper into a 25ml volumetric flask, continuously rinsing the test tubes 2-3 times, and keeping constant volume for later use;
B3. Determination
( Absorb 0.5ml to 3 graduated test tubes of the above extractive solution, and add 1.5ml distilled water respectively;
@ Quickly add 9g/ml phenol solution 1ml, immediately shake and mix well, and place on the test tube rack for 20s;
@ Slowly add 5ml concentrated sulfuric acid, immediately shake slightly and mix evenly to prevent liquid splashing;
@ Put in refrigerator at 4°C for 10mim, with color development,
@ The absorbance was measured at 485nm wavelength, and the sugar content was obtained according to the standard curve.
Claims
III. Results calculation
Calculate the sugar content of the sample according to the following formula
m x Vt x N Soluble sugar content (mg/g)= 106 m,12 x V3 X 10
Where m is the sugar content found from the standard curve, in tg; Vt is the volume of extractive solution, unit ml; Vs is the volume of sample taken for determination, in ml; N is the dilution multiple; mo is the sample quality.
2. The method for measuring soluble sugar content of sweet sorghum stem dry sample according to claim 1 is characterized in that in the B2 small step of step 2, the test tube is immersed in boiling water during the heating process, the test tube cannot be close to the beaker wall, uniform heating is ensured, and the heating time of each test tube is kept consistent.
Description of Drawings 19 Jun 2020
Light Absorption Value 2020101061
Different Sucrose Concentrations (μg)
Figure 1
Dry Sample
Fresh Sample Soluble Sugar Content (mg/g)
Anthesis Pustulation Period Autumn
Different Birth Periods
Figure 2
1/1
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