All fruits contain glucose and fructose, most also contain sucrose, and a few fruits, such as grapes, do not contain sucrose. Among the three sugars, fructose is the sweetest, sucrose is the second, and glucose is the worst. The sweetness ratio is 100: 66: 45.
The analysis results of apples are as follows: soluble sugar 9% ~ 14% (generally 1 1% ~ 12%), reducing sugar 7% ~ 10% (generally 9% ~10) The soluble sugar of pear is 7% ~ 14% (generally 8% ~ 10%), reducing sugar is 6% ~ 9%, and sucrose is 0.6% ~ 5%. The content of fructose in stone fruit (apple and pear) is higher, and the glucose in stone fruit (peach, plum and apricot) is slightly higher than fructose.
Principle of (1) method
Potassium ferricyanide can oxidize reducing sugar in alkaline solution. When a certain amount of potassium ferricyanide solution with known concentration is titrated with sugar leaching solution, potassium ferricyanide is reduced to potassium ferrocyanide, and the reduced sugar is oxidized to sugar acid. When potassium ferricyanide exists in the solution, the solution shows the color of indicator. When potassium ferricyanide is completely reduced to potassium ferrocyanide, add a drop of sugar solution, and the indicator will be reduced to khaki triphenol methane compound, and the sugar content will be calculated according to the volume of consumed sugar solution.
(2) preparation of reagent
① 1% potassium ferricyanide solution: accurately weigh 100g of analytically pure K3Fe(CN)6 dried at 105℃, add deionized water to dissolve it in a 300ml beaker, transfer it to a 1l volumetric flask, dilute it to scale, shake it evenly and put it in a brown bottle for later use.
②2.50 mol/L NaOH solution: In a small beaker, weigh 105g NaOH (analytically pure), dissolve it in water, cool it, and dilute it with water to 1l (solution A). Shake well, then suck 5ml from solution A and add water to 100ml (solution B). Suck 10 ~ 15 ml solution b and calibrate its concentration with 0. 1 mol/L standard acid. The concentration of solution B multiplied by 20 is the concentration of solution A. Liquid A needs to be accurate to 20.50 mol/L. ..
③ Neutral lead acetate solution: weigh 300g of lead acetate [Pb (CH3, Chief Operating Officer) 2.3H2O] and 50g of lead oxide (PbO) and add 50ml of water, heat it to white or purplish red in a boiling water bath (about 2-3h), then add 950ml of water while stirring, pour the mixed solution into a bottle, cover it and put it in a warm place until the solution is clear. Filter and store in a sealed brown bottle for later use.
④ saturated sodium sulfate (Na2SO4. 10H2O) solution: weigh 165g Na2SO4. 10H2O and dissolve in 10H2O.
⑤ methylene blue indicator: 1.0g methylene blue is dissolved in 100ml water.
⑥ Methyl red indicator: dissolve 0.2g of methyl red in100 ml of 60% ethanol.
⑦8% sodium hydroxide solution: dissolve 8 g sodium hydroxide in water and dilute it to 100 ml.
8 Dilute sodium hydroxide solution: dilute 10 ml of 8% sodium hydroxide solution to 100 ml.
9 concentrated hydrochloric acid (HCl): specific gravity 1. 19, analytical purity.
(3) Instruments and equipment
One in ten thousand. 1% balance, constant temperature water bath pot, high-speed tissue masher, etc.
Sample collection: take several representative fruits (not less than 10) without pests and diseases in the middle of the orchard crown periphery according to different treatments, bring them back to the laboratory, wash and dry the fruit samples or wipe the peel with a wet towel. When analyzing apples, pears and grapes, it is necessary to bring the peel. Peaches that are easy to peel should be peeled, and citrus should be stripped of the exocarp and the white mesh inside. Watermelon and melon only take edible parts. The seeds of grapes, apples, peaches, plums, apricots, oranges and watermelons should be thrown away, and the skins of apples and pears should also be removed.
When sampling, apples, pears, peaches, watermelons, melons and other fresh fruits. Take it out from the middle of each fruit and chop it with a stainless steel knife. Citrus takes two petals on the diagonal of each fruit; Berries (grapes) take several fruits from the top, bottom, left and right sides of each ear. Grapes are sometimes analyzed with juice.
(4) Determination method
① Extraction of sugar solution: weigh 65438±050g chopped sample in a 400ml beaker, put it on a dish balance, add 65438±050ml deionized water, and stir it on a high-speed tissue masher for 3min. Weigh 50g slurry (equivalent to 25g sample) on an industrial balance with a small beaker, and rinse it into a 250ml volumetric flask with water, so that the liquid in the flask is about 150ml. Neutralize the organic acid in the solution with 8% sodium hydroxide solution, add 1 ~ 2 drops respectively, shake the solution evenly, and put a little red litmus test paper in the bottle until the red litmus test paper turns blue obviously in the solution.
Put the bottle in a water bath at 80℃ for 30 minutes and shake it every 5 minutes. Pay attention to adjust the temperature of the water bath. Take out the bottle from the water bath, and add neutral lead acetate while it is hot, so as to precipitate protein and pigment. A few drops each, shake the bottle well, and then let it stand for a while to see if the upper solution is clear. If it is clear, don't add lead acetate to prevent excessive, but it will make the solution turbid, and it is impossible to filter out the clear night (according to experience, 25 grams of Marshal apple needs about 1.6 ~ 65438+ neutral lead acetate. 25 grams of pears need 1.5 ~ 3.5 ml) to be placed for a few minutes, and then saturated sodium sulfate solution with 3 ~ 4 times the amount of lead acetate is added to make excess lead generate lead sulfate precipitation. Cool the volumetric flask, add water to 250 ml, shake well, and filter with filter paper into a 250 ml triangular flask. This sugar solution can be used to determine reducing sugar.
② Determination of reducing sugar: Before formal determination, it should be predicted once. Add 20 ml of 1% potassium ferricyanide solution, 5 ml of 2.50 mol/L sodium hydroxide solution and 3 ml of sugar solution into a 100 ml triangular flask with three burettes, put the triangular flask into a 300-watt electric furnace with asbestos mesh and boil for 1 min, add a drop of methylene blue indicator, and then continue with sugar solution while changing heating. The color change of the solution: green-red-blue-purple-purple-khaki, and the mutation from purple to khaki will reach the end. When the total amount of sugar consumed (including 3 ml added first) is between 4 and 6 ml, the result is the most accurate. If the consumption of sugar solution exceeds 8 ml, the concentration of sugar is too weak. It is appropriate to titrate with 10 ml potassium ferricyanide solution and 2.5 ml 2.50 mol/L sodium hydroxide solution in sugar solution. If boiling 1 min, the solution turns blue immediately after adding methylene blue indicator, indicating that the sugar solution is too strong and needs to be diluted twice before titration.
Formal titration: From the above prediction, we can roughly get the consumption of sugar when it reacts with 20 ml of potassium ferricyanide solution. For formal titration, first add 0.5 ml of sugar solution less than the above sugar consumption into 20 ml of potassium ferricyanide solution and 5 ml of sodium hydroxide solution (for example, if the predicted consumption is 5.00 ml, add 4.5 ml first), boil 1 min, and then add 1 drop indicator.
③ Determination of soluble sugar: In the titration of reducing sugar, although there is sucrose in the solution, it cannot be oxidized by potassium ferricyanide, so only the content of reducing sugar can be titrated. If you want to titrate sucrose, take another sugar solution and add acid to hydrolyze it, then convert it into the same amount of glucose and fructose, and then titrate it according to the above method.
Determination steps: Suck 50 ml of sugar filtrate, put it in a 1 volumetric flask, take another 100 ml volumetric flask, add 50 ml of water, insert a thermometer to observe the temperature change, and put it in a water bath preheated to 70℃. When the temperature in the flask rises to 60℃, take out the volumetric flask and add 3 ml hydrochloric acid (specific gravity 650) accurately with a burette. Put the volumetric flask into the water bath again (be careful to shake it frequently). When the temperature in the flask rises to 70℃, keep it at 68 ~ 70℃ for 8 minutes accurately. Take out the volumetric flask, quickly cool it, add 2 drops of methyl red indicator, neutralize it with 8% sodium hydroxide solution, and use dilute sodium hydroxide solution until the solution changes from red to orange. If the solution turns yellow and turbid, it means that too much alkali is added, and it should be turned back to orange-red with dilute hydrochloric acid. Add water to the hydrolyzed sugar solution to 100 ml, and calibrate it with 20 ml potassium ferricyanide solution according to the determination of reducing sugar.
Determination of concentration of potassium ferricyanide solution: accurately weigh 0.2000 g of analytically pure glucose baked at 1 10℃ for 2 hours, dissolve it in water, and dilute it to 100 ml. Titrate this solution with 100 ml potassium ferricyanide solution in the same way as above, and record the amount of sugar solution.
(5) Calculation of results
10 ml potassium ferricyanide solution is equivalent to the number of grams of glucose (g) as follows:
G=M×V/ 100
Reducing sugar (%) = 2g× v/kloc-0 /×100/w× v2.
Soluble sugar (%) = 2g× v/kloc-0 /× 2×100/w× v3.
Sucrose (%) = soluble sugar (%)-reducing sugar (%)] × 95%
Where: m-weight of glucose (g);
V refers to the volume (ml) of standard glucose solution used for titration of 10 ml potassium ferricyanide solution;
V 1- constant volume of sugar leaching solution (ml);
V2—— the volume (ml) of sugar solution to be measured consumed by 20 ml of potassium ferricyanide solution when measuring reducing sugar;
V3—— the volume (ml) of the hydrolyzed sugar solution to be measured consumed by 20 ml of potassium ferricyanide solution when determining soluble sugar;
W—— fresh weight of sample (g);
2—— In the determination of soluble sugar, 50 ml of filtrate was sucked out, and after hydrolysis, the volume was constant to 100 ml, which doubled the raw sugar solution.