[0038] The present invention will be further described below in conjunction with specific embodiments:
[0039] A method for extracting dietary fiber from sweet potato, comprising: removing starch, enzymatic hydrolysis, centrifuging and extracting dietary fiber, said extracting dietary fiber includes extracting insoluble dietary fiber and extracting soluble dietary fiber;
[0040] Extract insoluble dietary fiber: Centrifuge the mixed solution after enzymatic hydrolysis to obtain separated precipitate and supernatant, soak the obtained precipitate with ethanol solution, and then filter by suction to obtain solid and filtrate; the obtained solid It is insoluble dietary fiber;
[0041] Extraction of soluble dietary fiber: the supernatant and filtrate obtained in the above process are evaporated most of the water in a water bath, and then the 95% ethanol solution is added for extraction. At this time, flocculent precipitation appears in the supernatant and the filtrate. The supernatant and the filtrate are separated into a flocculent precipitate, and the obtained flocculent precipitate is soluble dietary fiber.
[0042] The present invention adds a pretreatment process between the starch removal and enzymatic hydrolysis processes, specifically adding NaOH solution, stirring uniformly, and soaking for 1 hour.
[0043] The enzymatic hydrolysis specifically includes adding phosphate buffer to adjust pH, hydrolyzing with alpha-amylase, saccharifying enzyme, adding disodium hydrogen phosphate buffer and adjusting pH, and enzymatic hydrolyzing with trypsin.
[0044] The specific operation process of the present invention is:
[0045] The first step is the preparation of sweet potato flour: wash fresh sweet potatoes, cut into pieces, soak them in baking soda solution for 30 minutes, then mash them with a high-speed tissue masher, wash them with water for 2 to 5 times, and use three layers of gauze for each wash Filter to obtain fresh sweet potato residue, dry and grind to obtain sweet potato powder.
[0046] The second step, pretreatment: take sweet potato powder and 0.2mol/L sodium hydroxide solution, make a ratio of 1:10 by weight, and stir evenly. After stirring evenly, soak for 1 hour to make the sodium hydroxide and sweet potato powder. mixture;
[0047] The third step, enzymolysis:
[0048] a. Add phosphate buffer to the mixed solution and adjust the pH of the mixed solution to about 4.5-5.0.
[0049] b. After obtaining a mixed solution from a, add α-amylase solution to it at a temperature of 60°C, and hydrolyze the enzyme for 40 minutes; kill the enzyme at 90°C;
[0050] c. Add the saccharification enzyme solution to the mixed solution enzymatically hydrolyzed with α-amylase solution in b at a pH of 4.5 to 5.0 and a temperature of 60°C, and hydrolyze in a water bath for 40 minutes; Enzyme
[0051] d. Add disodium hydrogen phosphate buffer and NaOH to the mixed solution hydrolyzed in a water bath with saccharification enzyme solution in c, adjust Ph7-9 or so, add trypsin solution to it at a temperature of 60℃, and hydrolyze in a water bath 40min; Inactivate enzyme at 90℃;
[0052] e. After the mixed solution hydrolyzed in a trypsin solution in d, bath in a boiling water bath for 5 minutes to inactivate all enzymes;
[0053] The fourth step is to extract dietary fiber:
[0054] f. Extract insoluble dietary fiber: The mixed solution that has passed through the boiling water bath in the third step is centrifuged through a centrifuge to obtain separated precipitate and supernatant. The obtained precipitate is soaked in 95% ethanol for 2 minutes, and then transferred Suction and filter on a suction filter to obtain solids and filtrate; the obtained solids are insoluble dietary fiber;
[0055] g. Extract soluble dietary fiber: evaporate most of the water in the supernatant and filtrate obtained in F in a water bath, and then add a 95% ethanol solution for extraction at a ratio of 4:1. At this time, the supernatant and filtrate The flocculent precipitation appears in the supernatant and the filtrate through the centrifuge to separate the flocculent precipitate, and the obtained flocculent precipitate is the soluble dietary fiber.
[0056] The trypsin solution is prepared by solid powder trypsin and water at a ratio of 0.5-0.7 mL/g.
[0057] The saccharification enzyme liquid is a solid powder of saccharification enzyme and water prepared at a ratio of 4.0-5.0 mL/g.
[0058] The α-amylase liquid is a solid powder of α-amylase and water prepared at a ratio of 1.0-1.4 mL/g.
[0059] The sweet potato flour can also be replaced by sweet potato dregs, a by-product produced by a sweet potato starch production enterprise.
[0060] The trypsin solution, saccharification enzyme solution, and α-amylase solution should be stored at a temperature of 3 to 5°C.
[0061] The glucoamylase dosage is 5.0 mL/g, the alpha-amylase dosage is 1.4 mL/g, and the trypsin dosage is 0.5 mL/g.
[0062] In order to further illustrate the present invention, the present invention will be specifically described in combination with experiments as follows:
[0063] Preliminary process of the experiment: The first step: preparation of sweet potato powder, specifically: wash 2Kg of fresh sweet potato, cut into small pieces, soak in sodium bicarbonate solution for 30 minutes, then mash with a high-speed tissue masher for 20 seconds, and wash with water 3 times After washing, filter with three layers of gauze to remove the starch and polysaccharides in the sweet potato dregs to obtain fresh sweet potato dregs, weigh, and blast dry for 8 hours in a blast drying box at 60°C (stirring several times in the middle), and dry thoroughly Then take it out, cool and weigh, dry and seal it for later use.
[0064] The second step is the preparation of enzyme solution. The most important medicines used in the third step of the present invention are three enzymes, namely α-amylase, trypsin and saccharification enzyme, of which α-amylase and saccharification enzyme are both Food grade, trypsin is chemically pure (250U/mg), and all three enzymes are solid powders.
[0065] Since the enzyme reagent to be added in the third step of the present invention is liquid, the first thing to do is to prepare the enzyme liquid. Among them, the ratio of enzyme to water in the preparation of α-amylase solution, saccharification enzyme solution and trypsin solution is 1:8 (g/mL), 1:10 (g/mL), and 1:1 (mg/mL). The prepared enzyme solution is stored in a refrigerator at about 4°C for later use. To maintain better activity, the α-amylase solution and saccharification enzyme solution are stored for 3 to 5 days, and the trypsin solution is stored for 1 to 2 weeks.
[0066] Test design
[0067] 1. Prepare sweet potato powder according to the first step of the present invention, and then accurately weigh 7 portions of 1.000±0.005g sweet potato powder into 7 50mL conical flasks.
[0068] 2. In 7 conical flasks containing sweet potatoes, add 0.2mol/L NaOH solution at a ratio of 1:10 by weight, stir evenly, and soak for 1 hour.
[0069] 3. Then add 50 mL of phosphate buffer (pH 4.92) to the 7 mixed solutions soaked in 2 and stir well and adjust the pH to about 4.5-5.0 with hydrochloric acid.
[0070] 4. Then add α-amylase to the 7 mixed solutions soaked in 3, in a constant temperature water bath at 55°C for 40 minutes, and inactivate the enzyme at 90°C for 5 minutes.
[0071] 5. Then add glucoamylase to the 7 mixed solutions soaked in 4, in a constant temperature water bath at 55°C for 40 minutes, and inactivate the enzyme at 90°C for 5 minutes.
[0072] 6. Then add the disodium hydrogen phosphate buffer solution to the 7 mixed solutions soaked in 5, and adjust the Ph to about 8 with NaOH.
[0073] 7. Then, add trypsin to the 7 mixed solutions in 6 in a constant temperature water bath at 55°C for 40 minutes and inactivate the enzyme at 90°C for 5 minutes.
[0074] 8. Centrifuge the mixed solution obtained in 7 for 10 minutes at 4000 r/min.
[0075] 9. Separate the supernatant and the precipitate: add 95% ethanol to the precipitate and stir, then use a vacuum pump to filter, and dry in a blast drying oven at 60°C for about 6 hours to obtain insoluble dietary fiber; In an Erlenmeyer flask with 4 times the volume of 95% ethanol, the mixture is stirred and left to stand overnight. Then, it is filtered with a vacuum pump, dried and precipitated to obtain soluble dietary fiber.
[0076] The above is the dietary fiber extraction method adopted in this experiment. Among them, the specific amount of each substance added in steps 4, 5, and 7 depends on the test type (will be given in the following operation), and it needs to be stirred every 10 minutes during the water bath. In operation 9, if the obtained clear liquid containing soluble dietary fiber is relatively large, it can be evaporated and concentrated with a rotary evaporator to reduce the volume, and then add four times the volume of absolute ethanol for precipitation. The final obtained insoluble and soluble dietary fiber needs to be weighed separately in dry and wet weights to calculate and compare the extraction rate of insoluble dietary fiber and soluble dietary fiber under different conditions.
[0077] Test results and analysis
[0078] Determination of α-amylase dosage
[0079] Weigh 7 portions of 1.000±0.005g sweet potato residue powder accurately respectively, add NaOH solution to soak for 1h, add 50mL phosphate buffer, adjust pH 4.5~5.0, press 0.6, 0.8, 1.0, 1.2, 1.4, 1.6, 1.8mL/ Add g of α-amylase solution in a constant temperature water bath at 55°C for 40 minutes to inactivate the enzyme, then add 4.5 mL of the saccharification enzyme solution for enzymatic hydrolysis for 40 minutes, then inactivate the enzyme at a high temperature, add disodium hydrogen phosphate buffer, and adjust the pH of the solution to 8. 0.7mL trypsin solution (250U/mL) was added, and the enzymes were digested in a constant temperature water bath at 55°C for 40 minutes, and then the enzymes were inactivated. Investigate the effect of α-amylase dosage on the extraction effect of dietary fiber.
[0080] According to the above-mentioned α-amylase dosage determination, the dietary fiber extraction test was carried out by enzymatic hydrolysis, the results were recorded and analyzed, and the dietary fiber extraction rate and the changes that occurred with the increase of the α-amylase addition amount were drawn from the analyzed data.
[0081] by figure 1 It can be seen that the extraction rate of dietary fiber increases first and then decreases with the increase in the amount of α-amylase. When the amount of α-amylase increases to about 1.4mL/g, the yield of dietary fiber is the largest; among them, the soluble The extraction rate of dietary fiber increases firstly and then tends to be flat with the increase in the amount of α-amylase. When the amount of α-amylase is less than 1.4mL/g, the extraction rate of soluble dietary fiber increases slowly, and is added at 1.4mL/g When the amount of α-amylase exceeds 1.4mL/g, the extraction rate of soluble dietary fiber increases slightly, but the increase tends to be stable; according to the comprehensive evaluation value, the orthogonal test α-amylase amount is determined The most suitable range is 1.0~1.4mL/g.
[0082] Determination of the amount of glucoamylase added. Weigh 7 portions of 1.000g sweet potato residue powder accurately, add 0.2M/L NaOH solution and soak for 1h, add 50mL phosphate buffer, adjust pH 4.5~5.0, and 1.4mL α-amylase solution , Add saccharification enzyme solution in the amount of 3.5, 4.0, 4.5, 5.0, 5.5, 6.0, 6.5mL/g, hydrolyze the enzyme in a constant temperature water bath at 55℃ for 40min, add disodium hydrogen phosphate buffer, adjust the pH of the solution to 8 , Then add 0.7mL trypsin solution for enzymolysis, and then inactivate the enzyme. Investigate the effect of the amount of glucoamylase on the effect of dietary fiber extraction.
[0083] According to the determination of the amount of glucoamylase described above, the dietary fiber extraction test was carried out by enzymatic hydrolysis, the results were recorded and analyzed, and the dietary fiber extraction rate and the changes that occurred with the increase in the amount of glucoamylase added were drawn from the analyzed data. by figure 2 It can be seen that the total yield of dietary fiber increases slowly and then decreases slowly with the increase of the amount of glucoamylase. When the amount of glucoamylase increases to 5.0mL/g, the yield of dietary fiber is the largest; the extraction rate of soluble dietary fiber increases with The increase in the amount of glucoamylase showed a trend of first increasing and then decreasing. When the amount of glucoamylase was less than 4.5mL/g, the extraction rate of soluble dietary fiber increased slowly. When the dosage of glucoamylase was 4.5-5.5mL/g, the soluble dietary fiber The extraction rate stabilized and then dropped sharply. According to the comprehensive evaluation value, the optimal range of the orthogonal test glucoamylase dosage is 4.0~5.0mL/g.
[0084] Determine the amount of trypsin added. Weigh exactly 7 portions of 1.000±0.005g sweet potato residue powder, add NaOH solution and soak for 1 hour, add 50mL phosphate buffer, adjust pH 4.5~5.0, α-amylase solution 1.4mL, glucoamylase Enzymatic hydrolysis of 4.5 mL of the solution was performed in a constant temperature water bath at 55°C for 40 minutes to inactivate the enzyme. Add disodium hydrogen phosphate buffer, adjust the pH of the solution to 8, add trypsin solution in the amount of 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0 mL/g, and inactivate the enzyme in a constant temperature water bath at 55°C for 40 minutes. Investigate the effect of trypsin dosage on the effect of dietary fiber extraction.
[0085] According to the above-mentioned trypsin dosage determination, the dietary fiber extraction test was carried out by enzymatic hydrolysis, the results obtained were recorded and analyzed, and the dietary fiber extraction rate and the changes that occurred with the increase of the trypsin addition amount were drawn from the analyzed data.
[0086] by image 3 It can be seen that the extraction rate of dietary fiber shows an increasing trend as the amount of trypsin increases. When the amount exceeds 0.7mL/g, the extraction rate begins to slowly decrease. Because the soluble dietary fiber in dietary fiber has its main physiological function and is the main indicator of dietary fiber quality, it is necessary to focus on the influence of the extraction rate of soluble dietary fiber. It can be seen from the figure that the extraction rate of soluble dietary fiber decreases with the increase of trypsin dosage, then increases and then decreases, and when the trypsin dosage reaches 0.7~0.8mL/g, the soluble dietary fiber The extraction rate reaches the maximum, which may be due to the protein in glycoprotein being degraded by the action of high concentration trypsin; therefore, according to the comprehensive evaluation value, the range of trypsin addition in the orthogonal test is determined to be 0.5-0.7mL/g.
[0087] Experimental Design of Optimum Technological Conditions for Extracting Dietary Fiber from Sweet Potato Residue
[0088] In the present invention, three kinds of enzymes are mainly used for processing: α-amylase, trypsin and glucoamylase. In the above experiments, the effects of the three kinds of enzymes on the extraction rate of total dietary fiber were analyzed, and the optimal dosages of various enzymes were determined under the condition that other conditions remained unchanged. On this basis, the orthogonal test of enzyme dosage for extracting dietary fiber from sweet potato residue by enzymatic hydrolysis can be carried out.
[0089] The data records obtained by the orthogonal experiment are shown in Table 1. Since it is known that there is no interaction between the three enzyme addition factors based on the existing experimental experience, the fourth column (ie D blank) is regarded as the error column. The comprehensive evaluation in the table refers to the comprehensive evaluation of the total extraction rate of dietary fiber and the extraction rate of soluble dietary fiber (and the average of this sum). K and k at the bottom of the table represent the sum of the comprehensive evaluation of each factor and each level. Average. R represents the range of the comprehensive evaluation average of the three levels under each factor (that is, the maximum value minus the minimum value).
[0090] Table 1 Orthogonal test results of enzyme dosage in the enzymatic extraction process of dietary fiber from sweet potato residue
[0091]
[0092] By analyzing the orthogonal test results shown in Table 1, it can be seen intuitively that the order of the three factors listed in the test is C→A→B, that is, the amount of glucoamylase→α-amylase Dosage → Trypsin dosage; the optimal combination is (A 3 B 1 C 3 ), that is, the dosage of glucoamylase is 5.0 mL/g, the dosage of α-amylase is 1.4 mL/g, and the dosage of trypsin is 0.5 mL/g.
[0093] Table 2 Variance Analysis of Fruits in the Orthogonal Test of Enzyme Dosage in the Extraction Process of Dietary Fiber from Sweet Potato Residue
[0094]
[0095] It can be seen from the variance analysis of the orthogonal test results in Table 2 that the variation from the three factors from large to small is: C→B→A (that is, the amount of glucoamylase → the amount of α-amylase → the amount of trypsin) , The F value corresponding to factor C is greater than F 0.05 (2, 2) but less than F 0.01 (2, 2), indicating that the amount of glucoamylase has a significant impact on the comprehensive evaluation value of dietary fiber.
[0096] Orthogonal test optimization of saccharification enzyme hydrolysis technology
[0097] It can be concluded from the analysis of variance in Table 2 that glucoamylase has a significant effect on the extraction effect of dietary fiber. Therefore, the orthogonal test is carried out with the enzymatic hydrolysis conditions of glucoamylase, namely temperature, time and pH as factors, and the analysis results are recorded as shown in Table 3. Show.
[0098] Since there is no interaction between the above three factors, we can use the D (blank) column as the error column. The comprehensive evaluation is the average of the corresponding TDF (total dietary fiber) extraction rate and SDF (soluble dietary fiber) extraction rate. K and k represent each factor (temperature, time, pH, and error column, respectively) ) The sum and average of comprehensive evaluation at each level. R represents the range of the average of the three levels of comprehensive evaluation under each factor (that is, the maximum value minus the minimum value).
[0099] Table 3 Orthogonal test results of the amount of glucoamylase in the enzymatic extraction process of dietary fiber from sweet potato residue
[0100]
[0101] From the analysis results listed in Table 3, it can be seen that the primary and secondary order of the test factors affecting the extraction rate of sweet potato dietary fiber is A→C→B, that is, temperature→pH→time, and the optimal combination is A 3 B 2 C 1 That is, the enzymatic hydrolysis temperature of the glucoamylase is 60℃, the enzymatic hydrolysis time is 40min, and the pH is 4.5. Therefore, it can be determined that this condition is the optimal combination of the process conditions for the enzymatic hydrolysis of the sweet potato dregs by the glucoamylase. Under this optimal technological condition, the total extraction rate of sweet potato dietary fiber by enzymatic hydrolysis can reach 0.879g, and the extraction rate of soluble dietary fiber can reach 0.238g.
[0102] Analysis on the characteristics of dietary fiber from sweet potato residue
[0103] Taking the optimal extraction process of the sweet potato residue dietary fiber as the test piece, the sweet potato residue dietary fiber product was prepared, and the water holding capacity and swelling power of the prepared dietary fiber product were measured. The results are as follows As shown in Table 4.
[0104] Table 4 Analysis of quality index of sweet potato residue dietary fiber products
[0105]
[0106] It can be seen from Table 4 that the total extraction rate of dietary fiber from sweet potato residue is 81.6%, and the extraction rate of soluble dietary fiber can reach 25.7%. Recently, more and more studies have found that the physiological and biochemical characteristics of soluble dietary fiber are significantly better than that of insoluble dietary fiber, and soluble dietary fiber should account for 30-50% of the total dietary fiber. At present, almost all cereal fiber does not meet this standard. The swelling power and water holding capacity of the dietary fiber of sweet potato residue reached 3.42mL/g and 665%, respectively, and its various functional indexes performed well.
[0107] Modern studies have shown that dietary fiber's water holding capacity can increase the volume and speed of human defecation, reduce rectal pressure, and prevent constipation and colon cancer. At the same time, the active groups on the surface can chelate and adsorb cholesterol and bile, thereby reducing the incidence of coronary heart disease. In addition, the volume of dietary fiber increases after rehydration, which can easily cause satiety, thereby preventing obesity caused by overeating.
