CN1474183A - Method for detecting starch content in ash tree flower polysaccharide - Google Patents
Method for detecting starch content in ash tree flower polysaccharide Download PDFInfo
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Abstract
The present invention is the method of detecting starch content in ash tree flower polysaccharide product. The detection includes the steps of eliminating liposoluble component, eliminating soluble saccharide as interference matter, enzyme decomposing starch, detecting starch, processing enzyme hydrolyzed starch liquid, Fehling's process of determining reducing sugar content, etc. The method of the present invention is suitable for operation, has stable and reliable determined result, small variation coefficient of determined result, less error, high repeatability and other advantages.
Description
(1) technical field
The present invention relates to the assay method of grifolan extract, relate in particular to the method for quantitatively determining of non-active ingredient content of starch in the grifola frondosus deep fermentation liquid polyoses extract.
(2) background technology
Active component polysaccharides and non-active ingredient polysaccharide-starch have mainly been comprised in the grifola frondosus deep fermentation liquid polyoses extract; Wherein, the quantitative measurement of non-active ingredient starch determines it is very necessary for the quality control of grifola frondosus active polysaccharide product and polysaccharide product producing process.Yet, still there is not desirable method at present about the mensuration of non-active ingredient content of starch in the grifolan, just adopt national standard " assay method of starch in the GB/T 5009.9-1985 food " to measure; Among its result, in grifolan the non-active ingredient contents of starch, the content that has also comprised the grifola frondosus active polysaccharide makes that the accurate detection of the content of grifola frondosus active polysaccharide has produced very big uncertainty in the quality control, production run of grifola frondosus active polysaccharide product.Therefore, the mensuration of starch has become key technical problem in its product development, production and the quality control in the grifola frondosus active polysaccharide product.And national standard method " assay method of starch in the GB/T 5009.9-1985 food " existing deficiency and how further improve and perfect has become in the present grifolan institute's problem demanding prompt solution in the content of starch assay method.
(3) summary of the invention
Purpose of the present invention promptly is at the existing deficiency of content of starch assay method in the above-mentioned grifolan, and the method for non-active ingredient content of starch in a kind of accurate mensuration grifola frondosus active polysaccharide is provided.The assay method of content of starch in the grifola frondosus active polysaccharide involved in the present invention has advantages such as workable, that measurement result is reliable and stable, error is little, reappearance is high, the Routine Test Lab condition can be measured.
The assay method of content of starch in the grifolan of the present invention, form by following steps:
1) removes liposoluble constituent: get the polysaccharide sample to be measured that oven dry grinds, divide another name 0.5～2 to restrain in 6 Centrifuge Cups, add the 30ml ether respectively, fully vibration is 5～15 minutes, centrifugal 5～10 minutes with 4000～5000 rev/mins, abandoning supernatant adds the 30ml ether again, and so repeated centrifugation is 3 times;
2) remove the soluble saccharide interfering material: the 80% ethanol 30ml that adds 50～60 ℃ in the Centrifuge Cup after above-mentioned abandoning supernatant, fully vibration is 5～15 minutes, with 4000～5000 rev/mins centrifugal 5～10 minutes, abandoning supernatant, so repeated centrifugation is 3 times; And then add 40～50 ℃ of distilled water 30ml in the cup after centrifugal, vibration is 60 minutes in 40～50 ℃ of waters bath with thermostatic control, with 4000～5000 rev/mins centrifugal 5～10 minutes, abandoning supernatant adds 30ml distilled water again, so repeated centrifugation is 3 times;
3) enzyme process starch-splitting: to step 2) add 5ml distilled water in the Centrifuge Cup after the abandoning supernatant, moistening and stir isolated sample, add 80～90 ℃ of distilled water 40ml again, after stirring evenly, put to handle in the boiling water bath and made starch gelatinization in 50～60 minutes; Be cooled to 50 ℃ afterwards, add 1% carbohydrase liquid 10ml, and add the phosphate buffer 25ml of pH4～6 immediately, add 5～7 of toluene again, mixing seals, and places to be incubated 20～24 hours under 45～48 ℃ of conditions and to carry out amylorrhexis;
4) amylorrhexis detects: get above-mentioned starch enzymolysis liquid 1 and be added dropwise to 1 of iodine solution and carry out hydrolysis result and detect, if show blueness, redness or purple, it is incomplete to be amylorrhexis, repeating step 3 again) the enzyme process starch-splitting, until add iodine solution detect do not show blue, redness or purple till, it is complete to be amylorrhexis;
5) starch enzymolysis liquid is handled: after the amylorrhexis liquid cooling with above-mentioned complete enzymolysis, use 6molL
-1NaOH is neutralized to neutrality, dropwise adds saturated Pb (0Ac) then while stirring
2Solution is not till starch enzymolysis liquid has white flocculent deposit; In starch enzymolysis liquid, dropwise add saturated metabisulfite solution again, till not producing white precipitate; With 4000-5000 rev/min of centrifugal 5-10 minute, supernatant is transferred to volumetric flask, regulating pH is 9.1～9.6, to measure the demand constant volume;
6) the Fehling method is measured content of reducing sugar: the starch enzymolysis liquid of getting after handling in the step 5) is a sample, presses GB5009.7-85 " assay method of reducing sugar in the food " direct titrimetric method, the content of soluble sugar in the working sample.
Wherein, above-mentioned steps 1) or 2) described duration of oscillation is 8～12 minutes.
Wherein, above-mentioned steps 2) temperature of described 80% ethanol is 55～58 ℃.
Wherein, above-mentioned steps 2) distilled water of described adding and the temperature of water bath with thermostatic control be 45～50 ℃.
Wherein, above-mentioned steps 3) pH of described phosphate buffer is 4.8～5.6.
Wherein, above-mentioned steps 3) described temperature of carrying out amylorrhexis is 45 ℃, and the time is 20 hours.
Wherein, above-mentioned steps 5) starch enzymolysis liquid pH after the described processing is 9.3～9.5.
Wherein, above-mentioned steps 6) assay method of reducing sugar comprises the steps: in the described food
1. demarcate basic copper tartrate solution: draw fehling reagent first liquid and each 5.0ml of second liquid, place the 150ml conical flask, add water 10ml, add 2 of beaded glasses, drip about 9ml glucose standard solution from buret, be controlled to be heated in the 2min and boil, taking advantage of boils continues to drip the glucose standard solution with per two seconds 1 speed, just take off until the solution blueness and to be terminal point, the cumulative volume of record consumption of glucose standard solution, operation repetitive is three parts simultaneously, gets its mean value, calculates every 10ml (first, each 5ml of second liquid) basic copper tartrate solution is equivalent to the quality (mg) of glucose;
2. sample solution prediction: draw fehling reagent first liquid and each 5.0ml of second liquid, place the 150ml conical flask, add water 10ml, add 2 of beaded glasses, be controlled to be heated in the 2min and boil, take advantage of the speed of boiling, from buret, drip sample solution, and keep the solution fluidized state with first quick and back slow, when treating that solution colour shoals, with 1 speed titration in per two seconds, just take off until the solution blueness and to be terminal point, record sample liquid consumes volume;
3. sample solution is measured: draw fehling reagent first liquid and each 5.0ml of second liquid, place the 150ml conical flask, add water 10ml, add 2 of beaded glasses, drip the sample solution that lacks 1ml than the prediction volume from buret, make in 2min, to be heated to and boil, taking advantage of boils continues with 1 speed titration in per two seconds, just takes off until blueness to be terminal point, and record sample liquid consumes volume, with three parts of method operation repetitives, draw the mean consumption volume.
Adopt the present invention to carry out the mensuration of content of starch in the grifolan product, have workable, measurement result is reliable and stable, the coefficient of variation of measurement result is little, error is little, the reappearance advantages of higher.Experiment shows, middle per sample content of starch difference, and the coefficient of variation is generally in 5%～10% scope.Can satisfy Routine Test Lab to the requirement of analytical test and relevant enterprise to the controllable quality requirement.
The present invention is further illustrated below in conjunction with embodiment.
(4) embodiment
Embodiment 1:
1) removes liposoluble constituent: get the polysaccharide sample to be measured (being numbered No. 1) that oven dry grinds, claim 1.0727 grams, 1.1028 grams, 1.1509 grams, 0.948 gram, 1.1813 grams, 1.0481 grams to be put in respectively in 6 Centrifuge Cups, add the 30ml ether respectively, fully vibration is 12 minutes, centrifugal 8 minutes with 4500 rev/mins, abandoning supernatant adds the 30ml ether again, and so repeated centrifugation is 3 times;
2) remove the soluble saccharide interfering material: add 57 ℃ 80% ethanol 30ml in the Centrifuge Cup after above-mentioned abandoning supernatant, fully vibrated 15 minutes, with 5000 rev/mins centrifugal 10 minutes, abandoning supernatant, so repeated centrifugation is 3 times; And then add 50 ℃ of distilled water 30ml in the Centrifuge Cup after abandoning supernatant, vibration is 60 minutes in 50 ℃ of waters bath with thermostatic control, with 5000 rev/mins centrifugal 5 minutes, abandoning supernatant adds 30ml distilled water again, so repeated centrifugation is 3 times;
3) enzyme process starch-splitting: to step 2) add 5ml distilled water in the Centrifuge Cup after the abandoning supernatant, moistening and stir isolated sample, add 90 ℃ of distilled water 40ml again, after stirring evenly, put to handle in the boiling water bath and made starch gelatinization in 55 minutes; Be cooled to 50 ℃ afterwards, add 1% carbohydrase liquid 10ml, and add the phosphate buffer 25ml of pH 5.2 immediately, add 5 of toluene again, mixing seals, and places under 45 ℃ of conditions insulation to carry out amylorrhexis in 20 hours;
4) amylorrhexis detects: get above-mentioned starch enzymolysis liquid 1 and be added dropwise to 1 of iodine solution and carry out hydrolysis result and detect, to the result for add iodine solution detect do not show blueness, redness or purple till, it is complete to be amylorrhexis;
5) starch enzymolysis liquid is handled: after the amylorrhexis liquid cooling with above-mentioned complete enzymolysis, use 6molL
-1NaOH is neutralized to neutrality, dropwise adds saturated Pb (OAc) then while stirring
2Solution is not till starch enzymolysis liquid has white flocculent deposit; In starch enzymolysis liquid, dropwise add saturated metabisulfite solution again, till not producing white precipitate; With 4000 rev/mins centrifugal 10 minutes, get supernatant and be transferred to the 250ml volumetric flask, regulating pH is 9.3, constant volume is made as V
2
6) the Fehling method is measured content of reducing sugar: the starch enzymolysis liquid of getting after handling in the step 5) is a sample, presses GB5009.7-85 " assay method of reducing sugar in the food " direct titrimetric method, the content of soluble sugar in the working sample.
1. demarcate basic copper tartrate solution: draw fehling reagent first liquid and each 5.0ml of second liquid, place the 150ml conical flask, add water 10ml, add 2 of beaded glasses, drip about 9ml glucose standard solution from buret, be controlled to be heated in the 2min and boil, taking advantage of boils continues to drip the glucose standard solution with per two seconds 1 speed, just take off until the solution blueness and to be terminal point, the cumulative volume of record consumption of glucose standard solution, operation repetitive is three parts simultaneously, gets its mean value V0, calculates every 10ml (first, each 5ml of second liquid) basic copper tartrate solution is equivalent to the quality (mg) of glucose;
2. sample solution prediction: draw fehling reagent first liquid and each 5.0ml of second liquid, place the 150ml conical flask, add water 10ml, add 2 of beaded glasses, be controlled to be heated in the 2min and boil, take advantage of the speed of boiling, from buret, drip sample solution, and keep the solution fluidized state with first quick and back slow, when treating that solution colour shoals, with 1 speed titration in per two seconds, just take off until the solution blueness and to be terminal point, record sample liquid consumes volume;
3. sample solution is measured: draw fehling reagent first liquid and each 5.0ml of second liquid, place the 150ml conical flask, add water 10ml, add 2 of beaded glasses, drip the sample solution that lacks 1ml than the prediction volume from buret, make in 2min, to be heated to and boil, taking advantage of boils continues with 1 speed titration in per two seconds, just takes off until blueness to be terminal point, and record sample liquid consumes volume, with three parts of method operation repetitives, draw mean consumption volume V
1
Simultaneously, to do the reagent blank test with method.
The result calculates: starch %=V
0* C/ (1000mV
1) the mensuration embodiment analysis result of content of starch sees attached list 1 in * 0.9*100% (in the formula: content of starch is with glucose meter) the grifola frondosus active polysaccharide.
Wherein:
m
1---10ml basic copper tartrate solution (each 5ml of first, second liquid) is equivalent to the quality of reducing sugar (with glucose meter), mg;
M---sample quality, g;
V
0---the standard glucose liquor capacity that 10ml basic copper tartrate solution (each 5ml of first, second liquid) consumes, ml;
V
2---constant volume, ml
V
1---mean consumption sample solution volume during working sample, ml.
Embodiment 2:
| Subordinate list 1: the mensuration embodiment analysis result of content of starch in the grifola frondosus active polysaccharide | ||||||
| Embodiment (C=1mg/ml) | ||||||
| Sample number into spectrum | Repeat | ? ?????m(g) | ? ??V 2(ml) | ? ???V 1(ml) | ? ???V 0(ml) | Starch % |
| No. 1 (grifola frondosus crude polysaccharides 2N) | ??1 | ????1.0727 | ??100 | ???11.6 | ???11.86 | ?????8.58 |
| ??2 | ????1.1028 | ??100 | ???12.6 | ???11.86 | ?????7.68 | |
| ??3 | ????1.1509 | ??100 | ???11.6 | ???11.86 | ?????8.00 | |
| ??4 | ????0.948 | ??100 | ???17.29 | ???11.86 | ?????6.51 | |
| ??5 | ????1.1813 | ??100 | ???10.92 | ???11.86 | ?????8.27 | |
| ??6 | ????1.0481 | ??100 | ???12.56 | ???11.86 | ?????8.11 | |
| On average | ?????7.86 | |||||
| Standard deviation STDEV | ?????0.723 | |||||
| Coefficient of variation CV% | ?????9.205 | |||||
| Mean standard deviation | ?????0.2953 | |||||
| Average ± mean standard deviation | ????7.86±0.30 | |||||
1) removes liposoluble constituent: get the polysaccharide sample to be measured (being numbered No. 2) that oven dry grinds, claim 0.943 gram, 1.0392 grams, 0.85 gram, 0.9977 gram, 0.9023 gram, 0.9383 gram to be put in respectively in 6 Centrifuge Cups, add the 30ml ether respectively, fully vibration is 15 minutes, centrifugal 10 minutes with 4000 rev/mins, abandoning supernatant adds the 30ml ether again, and so repeated centrifugation is 3 times;
2) remove the soluble saccharide interfering material: add 50 ℃ 80% ethanol 30ml in the Centrifuge Cup after above-mentioned abandoning supernatant, fully vibrated 10 minutes, with 5000 rev/mins centrifugal 5 minutes, abandoning supernatant, so repeated centrifugation is 3 times; And then add 40 ℃ of distilled water 30ml in the Centrifuge Cup after abandoning supernatant, vibration is 60 minutes in 45 ℃ of waters bath with thermostatic control, with 4000 rev/mins centrifugal 10 minutes, abandoning supernatant adds 30ml distilled water again, so repeated centrifugation is 3 times;
3) enzyme process starch-splitting; To step 2) add 5ml distilled water in the Centrifuge Cup after the abandoning supernatant, moistening and stir isolated sample, add 80 ℃ of distilled water 40ml again, after stirring evenly, put to handle in the boiling water bath and made starch gelatinization in 60 minutes; Be cooled to 50 ℃ afterwards, add 1% carbohydrase liquid 10ml, and add the phosphate buffer 25ml of pH 4.8 immediately, add 7 of toluene again, mixing seals, and places under 48 ℃ of conditions insulation to carry out amylorrhexis in 24 hours;
4) amylorrhexis detects: get above-mentioned starch enzymolysis liquid 1 and be added dropwise to 1 of iodine solution and carry out hydrolysis result and detect, to the result for add iodine solution detect do not show blueness, redness or purple till, it is complete to be amylorrhexis;
5) starch enzymolysis liquid is handled: after the amylorrhexis liquid cooling with above-mentioned complete enzymolysis, use 6molL
-1NaOH is neutralized to neutrality, dropwise adds saturated Pb (OAc) then while stirring
2Solution is not till starch enzymolysis liquid has white flocculent deposit; In starch enzymolysis liquid, dropwise add saturated metabisulfite solution again, till not producing white precipitate; With 4800 rev/mins centrifugal 7 minutes, get supernatant and be transferred to the 250ml volumetric flask, regulating pH is 9.5, constant volume is made as V
2
6) the Fehling method is measured content of reducing sugar: the starch enzymolysis liquid of getting after handling in the step 5) is a sample, presses GB5009.7-85 " assay method of reducing sugar in the food " direct titrimetric method, the content of soluble sugar in the working sample.
1. demarcate basic copper tartrate solution: draw fehling reagent first liquid and each 5.0ml of second liquid, place the 150ml conical flask, add water 10ml, add 2 of beaded glasses, drip about 9ml glucose standard solution from buret, be controlled to be heated in the 2min and boil, taking advantage of boils continues to drip the glucose standard solution with per two seconds 1 speed, just takes off until the solution blueness to be terminal point, the cumulative volume of record consumption of glucose standard solution, operation repetitive is three parts simultaneously, gets its mean value V
0, calculate the quality (mg) that every 10ml (each 5ml of first, second liquid) basic copper tartrate solution is equivalent to glucose:
2. sample solution prediction: draw fehling reagent first liquid and each 5.0ml of second liquid, place the 150ml conical flask, add water 10ml, add 2 of beaded glasses, be controlled to be heated in the 2min and boil, take advantage of the speed of boiling, from buret, drip sample solution, and keep the solution fluidized state with first quick and back slow, when treating that solution colour shoals, with 1 speed titration in per two seconds, just take off until the solution blueness and to be terminal point, record sample liquid consumes volume;
3. sample solution is measured: draw fehling reagent first liquid and each 5.0ml of second liquid, place the 150ml conical flask, add water 10ml, add 2 of beaded glasses, drip the sample solution that lacks 1ml than the prediction volume from buret, make in 2min, to be heated to and boil, taking advantage of boils continues with 1 speed titration in per two seconds, just takes off until blueness to be terminal point, and record sample liquid consumes volume, with three parts of method operation repetitives, draw mean consumption volume V
1
Simultaneously, to do the reagent blank test with method.
The result calculates: starch %=V
0* C/ (1000mV
1) the mensuration embodiment analysis result of content of starch sees attached list 2 in * 0.9*100% (in the formula: content of starch is with glucose meter) the grifola frondosus active polysaccharide.
| Subordinate list 2: the mensuration embodiment analysis result of content of starch in the grifola frondosus active polysaccharide | ||||||
| Embodiment 2 (C=1mg/ml) | ||||||
| Sample number into spectrum | Repeat | ??m(g) | ?V 2(ml) | ???V 1(ml) | ????V 0(ml) | Starch % |
| No. 2 (grifola frondosus crude polysaccharides) | ??1 | ??0.943 | ??500 | ???10.23 | ????11.81 | ????55.09 |
| ??2 | ??1.0392 | ??500 | ???9.36 | ????11.81 | ????54.64 | |
| ??3 | ??0.85 | ??500 | ???10.08 | ????11.81 | ????62.03 | |
| ??4 | ??0.9977 | ??500 | ???9.83 | ????11.81 | ????54.19 | |
| ??5 | ??0.9023 | ??500 | ???10.73 | ????11.81 | ????54.89 | |
| ??6 | ??0.9383 | ??500 | ???10.14 | ????11.81 | ????55.86 | |
| On average | ????56.12 | |||||
| The STDEV standard deviation | ????2.948 | |||||
| Coefficient of variation CV% | ????5.254 | |||||
| Mean standard deviation | ????1.2036 | |||||
| Average ± mean standard deviation | ????55.12±1.20 | |||||
Claims (8)
1. the assay method of content of starch in the grifolan, form by following steps:
1) removes liposoluble constituent: get the polysaccharide sample to be measured that oven dry grinds, divide another name 0.5～2 to restrain in 6 Centrifuge Cups, add the 30ml ether respectively, fully vibration is 5～15 minutes, centrifugal 5～10 minutes with 4000～5000 rev/mins, abandoning supernatant adds the 30ml ether again, and so repeated centrifugation is 3 times;
2) remove the soluble saccharide interfering material: the 80% ethanol 30ml that adds 50～60 ℃ in the Centrifuge Cup after above-mentioned abandoning supernatant, fully vibration is 5～15 minutes, with 4000～5000 rev/mins centrifugal 5～10 minutes, abandoning supernatant, so repeated centrifugation is 3 times; And then add 40～50 ℃ of distilled water 30ml in the Centrifuge Cup after abandoning supernatant, in 40～50 ℃ of waters bath with thermostatic control the vibration 60 minutes, with 4000～5000 rev/mins centrifugal 5～10 minutes, abandoning supernatant, add 30ml distilled water again, so repeated centrifugation is 3 times;
3) enzyme process starch-splitting: to step 2) add 5ml distilled water in the cup after the centrifugal abandoning supernatant, moistening and stir isolated sample, add 80～90 ℃ of distilled water 40ml again, after stirring evenly, put to handle in the boiling water bath and made starch gelatinization in 50～60 minutes; Be cooled to 50 ℃ afterwards, add 1% carbohydrase liquid 10ml, and add the phosphate buffer 25ml of pH4～6 immediately, add 5～7 of toluene again, mixing seals, and places to be incubated 20～24 hours under 45～48 ℃ of conditions and to carry out amylorrhexis;
4) amylorrhexis detects: get above-mentioned starch enzymolysis liquid 1 and be added dropwise to 1 of iodine solution and carry out hydrolysis result and detect, if show blueness, redness or purple, it is incomplete to be amylorrhexis, repeating step 3 again) the enzyme process starch-splitting, until add iodine solution detect do not show blue, redness or purple till, it is complete to be amylorrhexis;
5) starch enzymolysis liquid is handled: after the amylorrhexis liquid cooling with above-mentioned complete enzymolysis, use 6molL
-1NaOH is neutralized to neutrality, dropwise adds saturated Pb (OAc) then while stirring
2Solution is not till starch enzymolysis liquid has white flocculent deposit; In starch enzymolysis liquid, dropwise add saturated metabisulfite solution again, till not producing white bag precipitation; With 4000-5000 rev/min of centrifugal 5-10 minute, supernatant is transferred to volumetric flask, regulating pH is 9.1～9.6, to measure the demand constant volume;
6) the Fehling method is measured content of reducing sugar: the starch enzymolysis liquid of getting after handling in the step 5) is a sample, presses GB5009.7-85 " assay method of reducing sugar in the food " direct titrimetric method, the content of soluble sugar in the working sample.
2. the assay method of content of starch is characterized in that in the grifolan as claimed in claim 1, step 1) or 2) described duration of oscillation is 8～12 minutes.
3. the assay method of content of starch is characterized in that step 2 in the grifolan as claimed in claim 1) temperature of described 80% ethanol is 55～58 ℃.
4. the assay method of content of starch is characterized in that step 2 in the grifolan as claimed in claim 1) distilled water of described adding and the temperature of water bath with thermostatic control be 45～50 ℃.
5. the assay method of content of starch is characterized in that in the grifolan as claimed in claim 1, and the pH of the described phosphate buffer of step 3) is 4.8～5.6.
6. the assay method of content of starch is characterized in that in the grifolan as claimed in claim 1, and the described temperature of carrying out amylorrhexis of step 3) is 45 ℃, and the time is 20 hours.
7. the assay method of content of starch is characterized in that in the grifolan as claimed in claim 1, and the starch enzymolysis liquid pH after the described processing of step 5) is 9.3～9.5.
8. the assay method of content of starch is characterized in that in the grifolan as claimed in claim 1, step 6) institute
The assay method of reducing sugar comprises the steps: in the food of stating
1. demarcate basic copper tartrate solution: draw fehling reagent first liquid and each 5.0ml of second liquid, place the 150ml conical flask, add water 10ml, add 2 of beaded glasses, drip about 9ml glucose standard solution from buret, be controlled to be heated in the 2min and boil, taking advantage of boils continues to drip the glucose standard solution with per two seconds 1 speed, just take off until the solution blueness and to be terminal point, the cumulative volume of record consumption of glucose standard solution, operation repetitive is three parts simultaneously, gets its mean value, calculates every 10ml (first, each 5ml of second liquid) basic copper tartrate solution is equivalent to the quality (mg) of glucose;
2. sample solution prediction: draw fehling reagent first liquid and each 5.0ml of second liquid,, place the 150ml conical flask, add water 10ml, add 2 of beaded glasses, be controlled to be heated in the 2min and boil, take advantage of the speed of boiling with first quick and back slow, from buret, drip sample solution, and keep the solution fluidized state, when treating that solution colour shoals, with 1 speed titration in per two seconds, just take off until the solution blueness and to be terminal point, record sample liquid consumes volume;
3. sample solution is measured: draw fehling reagent first liquid and each 5.0ml of second liquid, place the 150ml conical flask, add water 10ml, add 2 of beaded glasses, drip the sample solution that lacks 1ml than the prediction volume from buret, make in 2min, to be heated to and boil, taking advantage of boils continues with 1 speed titration in per two seconds, just takes off until blueness to be terminal point, and record sample liquid consumes volume, with three parts of method operation repetitives, draw the mean consumption volume.
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2003
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