CN102875641A - Method for continuously preparing pumpkin protein, ethanol fuel and dietary fiber - Google Patents
Method for continuously preparing pumpkin protein, ethanol fuel and dietary fiber Download PDFInfo
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Abstract
The invention relates to a method for continuously preparing pumpkin protein, ethanol fuel and dietary fiber, belonging to a technique for preparing active substances from natural substances. The method comprises the following steps: oscillating a pumpkin pulverized material solution at constant temperature, centrifuging, and filtering to obtain a filtrate and a filter residue; regulating the pH value of the filtrate to a protein isoelectric point by using a buffer solution, carrying out centrifugal separation to obtain a precipitate and a supernatant, carrying out washing and freeze-drying on the precipitate to obtain the pumpkin protein, and carrying out concentration and alcohol precipitation on the supernatant to obtain the water-soluble dietary fiber; and washing the pumpkin filter residue with water, adding water, sequentially carrying out alpha-amylase liquification and saccharifying enzyme saccharification, carrying out centrifugal separation to obtain a precipitate and a supernatant, washing the precipitate with water, carrying out freeze-drying to obtain the water-insoluble dietary fiber, carrying out fermentation treatment on the supernatant with active dry yeast, filtering, distilling the filtrate, and dehydrating with a desiccant to obtain the ethanol fuel. The invention has the advantages of no mutual interference of various steps, high product yield, simple technological operation and low cost, and can easily implement industrial production.
Description
Technical field
The invention belongs to the technology of preparation active substance from crude substance, relate generally to a kind of method take the discarded pulp of seed-use pumpkin as raw material continuous production Cucurmosin, alcohol fuel and food fibre.
Background technology
Pumpkin (
Cucurbita moschataDuch) be the Curcurbitaceae vine pumpkin fruit of overgrowing.China's pumpkin aboundresources, the pumpkin planting area occupies the second place of the world, and ultimate production ranks first in the world, and wherein 20% is edible, and 80% is seed-use pumpkin.Pumpkin nutrient is worth very high, be rich in the multiple nutritional components such as amino acid, protein, Mierocrystalline cellulose, electrolytes and minerals, also contain polysaccharide, carotenoid etc. and have the nourishing function bioactive ingredients, be mainly used in antitumor, hypoglycemic, reducing blood-fat, prevention prostatosis and strengthen the medical field such as immunity.Along with to the going deep into of pumpkin nutrient composition and value medical health care research, the exploitation of pumpkin product is subject to the attention of countries in the world day by day, and very wide exploitation prospect is arranged.
Seed-use pumpkin is the important pumpkin Cultivar of China, and owing to the nutrient composition contents such as solubility sugar and starch in the pulp of seed-use pumpkin kind are on the low side, mouthfeel is bad, and pulp is basic all discarded, causes serious waste.How the discarded pulp of seed-use pumpkin kind is taken full advantage of to become and be badly in need of the subject matter that solves in the seed-use pumpkin variety production.Protein content owing to less to the research of pumpkin fruit albumen, therefore can be used as new protein resource research and development both at home and abroad in the seed-use pumpkin pulp about 1.5%; Food fibre and food thereof as food function factor, the world in recent years various countries have also obtained paying close attention to widely and rapidly development, dietary fiber content is the main component of dry-matter up to 12.4% in the seed-use pumpkin pulp, and development and use seed-use pumpkin food fibre has huge market; Starch content is lower in the seed-use pumpkin pulp, and content is about 3%, but discarded pulp cost is zero, can be used as the raw material of preparation alcohol fuel, alleviates because grain prepares grain that ethanol brings and the rise of oil price.Therefore, take the discarded pulp of seed-use pumpkin as raw material, continuous production Cucurmosin, alcohol fuel and food fibre have wide market, and its prospect is very considerable.
At present about the preparation of activeconstituents in the pumpkin fruit with utilize the aspect to have some Patents, such as domestic patent CN102180991A " a kind of method that fully utilizes pumpkin ", disclose a kind of substep extraction method and extracted and utilized the membrane separation technique concentrated method that removes the gred, produce pectin, squash polyoses and pumpkin powder, but do not relate to the continuous production of other active substances such as albumen, food fibre in the pumpkin fruit; Patent CN102524698A " a kind of preparation method of pumpkin dietary fiber " discloses a kind of method of utilizing ultrafiltration, steaming and decocting under high pressure to obtain pumpkin dietary fiber in conjunction with high-pressure homogeneous and vacuum microwave treatment process; Patent CN101195835A " utilizes pumpkin to be the preparation technology of fermenting raw materials ethanol ", relates to take pumpkin fruit as raw material, smashes the method that obtains ethanol behind the defibrination by enzymatic hydrolysis and fermentation and second distillation.But patent CN102524698A only can separate and utilize a kind of effective constituent with CN101195835A, and other activeconstituentss have to remove as impurity in purge process, has caused the very large wasting of resources.
Summary of the invention
The method that the purpose of this invention is to provide a kind of continuous production Cucurmosin, alcohol fuel and food fibre, the purpose that reach simple to operate, fully utilizes each active substance in the pumpkin.
The object of the present invention is achieved like this:
The method of a kind of continuous production Cucurmosin, alcohol fuel and food fibre comprises the discarded pulp of seed-use pumpkin is cleaned, removes the peel, dries, pulverized; 1. with the discarded pulp comminuting matter of pumpkin with 1～5% NaOH solution, constant temperature oscillation, centrifugation, filtration under liquid ratio 20:1～30:1,60～80 ℃ of temperature of extraction, extraction times 50～70 min conditions, get filtrate and filter residue; 2. add damping fluid and regulate the pH value to isoelectric points of proteins in gained filtrate, centrifugation must precipitate and supernatant liquor, and washing of precipitate, lyophilize are obtained Cucurmosin; 3. step 2. in centrifugation gained supernatant liquor obtain water-soluble dietary fibre through concentrated, alcohol precipitation; 4. after 1. middle filtration gained filter residue washes with water step, 3:1～8:1 adds entry by liquid ratio, successively with α-amylase insulation liquefaction and saccharifying enzyme insulation saccharification, centrifugation, must precipitate and supernatant liquor, with precipitation wash with water, lyophilize obtains water insoluble dietary fiber; 5. 4. centrifugation gained supernatant liquor active dry yeast constant temperature oscillation fermentation of step, obtain alcohol fuel behind the distillation of gained fermented liquid process, the desiccant dehydration.
Beneficial effect of the present invention is, selects 1～5% NaOH solution that Cucurmosin is had good lixiviate ability, and to insoluble dietary fibre in the pumpkin powder and starch lixiviate ability; In addition, by Cucurmosin and alcohol fuel preparation condition are carried out response surface optimization, protein extracting ratio and ethanol fermentation rate all reach the highest, under top condition, Cucurmosin product protein content reaches 70～80%, under optimum enzymolysis condition, the ethanol fermentation rate reaches 65～75%, is 90～99% alcohol fuel through obtaining content behind the single flash, desiccant dehydration again.The present invention is take the discarded pulp of seed-use pumpkin as raw material, and continuous, easy production has high value-added product, can spur economy, drives farmer richness.This method is simple to operate, does not need complicated equipment, is easy to industrialization, has broad application prospects.
Embodiment
The below is described in detail the embodiment of the invention.
The method of a kind of continuous production Cucurmosin, alcohol fuel and food fibre comprises the discarded pulp of seed-use pumpkin is cleaned, removes the peel, dries, pulverized; 1. with the discarded pulp comminuting matter of pumpkin with 1～5% NaOH solution, constant temperature oscillation, centrifugation, filtration under liquid ratio 20:1～30:1,60～80 ℃ of temperature of extraction, extraction times 50～70 min conditions, get filtrate and filter residue; 2. add damping fluid and regulate the pH value to isoelectric points of proteins in gained filtrate, centrifugation must precipitate and supernatant liquor, and washing of precipitate, lyophilize are obtained Cucurmosin; 3. step 2. in centrifugation gained supernatant liquor obtain water-soluble dietary fibre through concentrated, alcohol precipitation; 4. after 1. middle filtration gained filter residue washes with water step, 3:1～8:1 adds entry by liquid ratio, successively with α-amylase insulation liquefaction and saccharifying enzyme insulation saccharification, centrifugation, must precipitate and supernatant liquor, with precipitation wash with water, lyophilize obtains water insoluble dietary fiber; 5. 4. centrifugation gained supernatant liquor active dry yeast constant temperature oscillation fermentation of step, obtain alcohol fuel behind the distillation of gained fermented liquid process, the desiccant dehydration.
Step is regulated the damping fluid that the pH value uses to isoelectric points of proteins in 2. and is 0.1mol/L acetic acid-sodium acetate solution, and the Cucurmosin isoelectric pH value is in 3 ~ 6 scopes.
Step 2. middle centrifugation rotating speed is 5000～10000r/min, in centrifugation time 10～40min scope.
Step 2. middle washing precipitation adopts acetone and 95% ethanol continuous washing.
Step 3. in the supernatant liquor condition for preparing water-soluble dietary fibre be pH value 6～8,2～6 times of supernatant liquor volumes of dehydrated alcohol consumption, alcohol precipitation times 30～70 min.
Step 4. middle α-amylase insulation liquefaction top condition is pH value 4～6, α-amylase addition 0.05～0.15%, 60～80 ℃ of liquefaction temperatures, liquefying time 50～70 min.
Step 4. middle saccharifying enzyme insulation saccharification top condition is pH value 3～4, saccharifying enzyme addition 0.05～0.15%, 55～65 ℃ of saccharification temperatures, saccharification time 10～20 min.
Step 5. middle supernatant liquor is pH value 4～5, inoculum of dry yeast 0.05～0.10%, 28～30 ℃ of leavening temperatures, fermentation time 65～75 h with active dry yeast constant temperature oscillation fermentation top condition.
The dehydration siccative that uses was unslaked lime when step was prepared into alcohol fuel with fermented liquid in 5..
Determining of Cucurmosin optimum extraction condition
Determining of seed-use pumpkin pulp albumen iso-electric point:
Albumen after extracting is dissolved in the HAc-NaAc buffer system, regulate pH value 2 ~ 8, centrifugal 10～the 40min of 5000～10000r/min, obtain protein precipitation, with each washing precipitation of ethanol of acetone and 95% 3 times, take by weighing quality after the lyophilize, according to precipitation quality judging pumpkin fruit albumen iso-electric point, determine that seed-use pumpkin pulp albumen iso-electric point is 3 ~ 6 in the pH value.
Liquid ratio is on the impact of extraction rate of protein:
Get the 10.00g pumpkin powder, add 3%NaOH solution by different liquid ratios respectively, 80 ℃ of water-bath 1h, centrifugal suction filtration, supernatant liquor transfer pH to isoelectric points of proteins, protein precipitation is separated out, centrifugal, the precipitation washing with alcohol of acetone and 95%, last lyophilize, get pumpkin fruit albumen, and calculate the extraction yield of protein.Determine that liquid ratio is in 10:1～30:1 scope.
The NaOH massfraction is on the impact of extraction rate of protein:
Get the 10.00g pumpkin powder, be that the 20:1 ratio adds respectively different concns NaOH solution in liquid ratio, 80 ℃ of water-bath 1h, centrifugal suction filtration, supernatant liquor transfer pH to isoelectric points of proteins, protein precipitation is separated out, centrifugal, the precipitation washing with alcohol of acetone and 95%, last lyophilize, get pumpkin fruit albumen, and calculate the extraction yield of protein.Determine that the NaOH liquid quality fraction is 1～5%.
The alkali solubility temperature is on the impact of extraction rate of protein:
Get the 10.00g pumpkin powder, be that the 20:1 ratio adds 3%NaOH solution in liquid ratio, water-bath 1h under differing temps respectively, centrifugal suction filtration, supernatant liquor transfer pH to isoelectric points of proteins, protein precipitation is separated out, centrifugal, the precipitation washing with alcohol of acetone and 95%, last lyophilize, get pumpkin fruit albumen, and calculate the extraction yield of protein.Determine that extracting temperature is 50～90 ℃.
The molten time of alkali is on the impact of extraction rate of protein:
Get the 10.00g pumpkin powder, be that the 20:1 ratio adds 3%NaOH solution in liquid ratio, at 80 ℃ of lower respectively water-bath different times, centrifugal suction filtration, supernatant liquor transfer pH to isoelectric points of proteins, protein precipitation is separated out, centrifugal, the precipitation washing with alcohol of acetone and 95%, last lyophilize, get pumpkin fruit albumen, and calculate the extraction yield of protein.Determine that extraction time is 15～75 min.
Second-order regression for rotatable orthogonal design (1/2 implements):
Take extraction rate of protein mean value as examination index (establishing 3 repetitions), according to the single factor experiment result, select liquid ratio, NaOH concentration, temperature, these four factors of time are carried out the quadratic regression orthogonal rotational test, utilize SAS software that test-results is set up regression equation, and experimental factor is carried out the pairwise interaction analysis.
Table 1 quadratic regression rotary combination design table and extraction yield (1/2 implements)
The every variance analysis of table 2 regression equation
| The regression variance source | Degree of freedom | Sum of squares | Inequality | The F value | Prob﹥F | Significance |
| Once | 4 | 227.503591 | 0.5877 | 56.20 | <.0001 | ** |
| Quadratic term | 4 | 92.336065 | 0.2385 | 22.81 | 0.0009 | ** |
| Mutual | 6 | 61.182868 | 0.1581 | 10.08 | 0.0064 | * |
| Regression model | 14 | 381.022524 | 0.9843 | 26.89 | 0.0003 | ** |
| Lose and intend item | 2 | 4.428377 | 2.214189 | 5.39 | 0.0733 | ? |
| Pure error | 4 | 1.643680 | 0.410920 | ? | ? | ? |
| Total error | 6 | 6.072057 | 1.012010 | ? | ? | ? |
| Population variance | 20 | 387.094581 | ? | ? | ? | ? |
Table 3 regression analysis coefficient value and analytical results
| Parameter | Regression coefficient | Standard deviation | The T value | Prob﹥T | Significance |
| Constant | -257.199519 | 73.484971 | -3.50 | 0.0128 | * |
| b | 3.900360 | 2.367987 | 1.65 | 0.1506 | ? |
| c | -47.330732 | 11.909348 | -3.97 | 0.0073 | * |
| t | 5.746751 | 1.306707 | 4.40 | 0.0046 | ** |
| h | 4.547766 | 0.793957 | 5.73 | 0.0012 | ** |
| b*b | -0.118520 | 0.029300 | -4.05 | 0.0068 | ** |
| c*b | 1.457917 | 0.308652 | 4.72 | 0.0032 | ** |
| c*c | -2.363003 | 0.732488 | -3.23 | 0.0180 | * |
| t*b | -0.004236 | 0.019759 | -0.21 | 0.8373 | ? |
| t*c | 0.265625 | 0.098797 | 2.69 | 0.0361 | * |
| t*t | -0.032430 | 0.007325 | -4.43 | 0.0044 | ** |
| h*b | -0.059454 | 0.020577 | -2.89 | 0.0277 | ** |
| h*c | 0.338565 | 0.102884 | 3.29 | 0.0166 | * |
| h*t | -0.022523 | 0.006586 | -3.42 | 0.0142 | * |
| h*h | -0.024747 | 0.003256 | -7.60 | 0.0003 | ** |
Extraction yield under table 4 optimal conditions under optimal value and the optimal conditions
(liquid ratio 20:1～30:1, NaOH concentration 1～5%, 60～80 ℃ of temperature, time 50～70min) are carried out proof test, triplicate at optimum extraction conditions according to physical condition.It is 70～80% that the result gets extraction rate of protein, and trial value and model theory value are very approaching.And the revision test relative deviation is no more than 1～2%, illustrates that proteins extraction condition circulation ratio is good.
The alcohol fuel preparation condition is optimized
Liquefaction condition is optimized:
In the molten condition of the suitableeest alkali, taking by weighing the 20.00g pumpkin powder, to carry out alkali molten, and centrifugation washes the molten residue that obtains of alkali with water, then adds suitable quantity of water, adjust the temperature to liquefaction temperature after, add α-amylase, insulation is liquefied.Solution after the liquefaction is cooled to 60 ℃, adds the activation saccharifying enzyme by 0.1% of raw materials quality, water bath heat preservation 10min carries out saccharification.Centrifugation after saccharification finishes is cooled to 30 ℃ with supernatant liquor, adds the activation yeast by 0.1% of raw materials quality, the jam-pack fermentation bung, and 48h ferments in constant-temperature shaking incubator.Measure CO
2Weight loss and alcoholic strength are with how many definite each factor optimum values of fermentation rate.
PH affects the ethanol fermentation rate:
Determine that the α-amylase consumption is 0.1% of raw material, 70 ℃ of liquefaction temperatures, liquefying time are 60min, regulate respectively different pH.Take the ethanol fermentation rate as index, determine that the pH value is 4～8.
The α-amylase addition is on the impact of ethanol fermentation rate:
Determine that liquefying time is 60min, pH is 6, and liquefaction temperature is 70 ℃, presses respectively raw material and adds the different mass α-amylase.Take the ethanol fermentation rate as index, determine that the α-amylase addition is 0.01～0.2%.
Liquefaction temperature affects the ethanol fermentation rate:
Determine that liquefying time is 60min, pH is 6, and the α-amylase consumption is 0.1% of raw material, regulates respectively different liquefaction temperatures 5.Take the ethanol fermentation rate as index, determine that liquefaction temperature is 50～90 ℃.
Liquefying time affects the ethanol fermentation rate:
Determine that pH is 6, liquefaction temperature is 70 ℃, and the α-amylase consumption is 0.1% of raw material, regulates respectively different liquefying times.Take the ethanol fermentation rate as index, determine that liquefying time is 30～70 min.
Second-order regression for rotatable orthogonal design:
According to the single factor experiment result, take fermentation rate mean value as examination index (establishing 3 repetitions), select pH, α-amylase addition, liquefaction temperature, liquefying time to carry out the test of four factors quadratic regression orthogonal rotational, experimental factor has been carried out the pairwise interaction analysis.
According to the single factor experiment result, take fermentation rate mean value as examination index (establishing 3 repetitions), select pH, α-amylase addition, liquefaction temperature, liquefying time to carry out the test of four factors quadratic regression orthogonal rotational, experimental factor has been carried out the pairwise interaction analysis.
Table 5 quadratic regression rotary combination design table and fermentation rate (1/2 implements)
The every variance analysis of table 6 regression equation
| The regression variance source | Degree of freedom | Sum of squares | Inequality | The F value | Prob﹥F | Significance |
| Once | 4 | 176.004811 | 0.6547 | 135.43 | <.0001 | ** |
| Quadratic term | 4 | 76.992190 | 0.2864 | 59.24 | <.0001 | ** |
| Mutual | 6 | 13.897576 | 0.0517 | 7.13 | 0.0153 | * |
| Regression model | 14 | 266.894577 | 0.9927 | 58.68 | <.0001 | ** |
| Lose and intend item | 2 | 1.111423 | 0.555712 | 2.65 | 0.1848 | ? |
| Pure error | 4 | 0.838000 | 0.209500 | ? | ? | ? |
| Total error | 6 | 1.949423 | 0.324904 | ? | ? | ? |
| Population variance | 20 | 268.844 | ? | ? | ? | ? |
Table 7 regression analysis coefficient value and analytical results
| Parameter | Regression coefficient | Standard deviation | The T value | Prob﹥T | Significance |
| Constant | -368.134753 | 51.094621 | -7.20 | 0.0004 | ** |
| p | 43.243843 | 7.905475 | 5.47 | 0.0016 | ** |
| c | 354.277472 | 161.140931 | 2.20 | 0.0702 | ? |
| t | 3.379120 | 0.736191 | 4.59 | 0.0037 | ** |
| h | 5.688788 | 0.795790 | 7.15 | 0.0004 | ** |
| p*p | -4.025413 | 0.415036 | -9.70 | <.0001 | ** |
| c*p | 44.319444 | 17.488592 | 2.53 | 0.0444 | * |
| c*c | -1402.165138 | 166.014296 | -8.45 | 0.0002 | ** |
| t*p | 0.026736 | 0.055980 | 0.48 | 0.6498 | ? |
| t*c | 2.909722 | 1.119592 | 2.60 | 0.0407 | * |
| t*t | -0.025154 | 0.004150 | -6.06 | 0.0009 | ** |
| h*p | -0.080347 | 0.087443 | -0.92 | 0.3936 | ? |
| h*c | -9.270833 | 1.748859 | -5.30 | 0.0018 | ** |
| h*t | -0.003646 | 0.005598 | -0.65 | 0.5390 | ? |
| h*h | -0.035604 | 0.004150 | -8.58 | 0.0001 | ** |
Fermentation rate under table 8 optimal conditions under optimal value and the optimal conditions
(pH value 4～6, α-amylase addition 0.05～0.15%, 60～80 ℃ of temperature, time 50～70min) are carried out proof test, triplicate at optimum extraction conditions according to physical condition.It is 50～60% that the result obtains fermentation rate, and trial value and model theory value are very approaching.And the revision test relative deviation is no more than 0～1 %, illustrates that the liquefaction condition circulation ratio is good.
The saccharification condition optimizing:
According to the molten condition of the suitableeest alkali and the suitableeest liquefaction condition, take by weighing the 20.00g pumpkin powder and carry out molten, the liquefaction of alkali, behind the saccharification temperature that the karusen that obtains is cooled to set after the liquefaction, add the saccharifying enzyme after the activation, water-bath vibration certain hour carries out saccharification.Centrifugation after saccharification finishes is cooled to 30 ℃ with supernatant liquor, adds the activation yeast by 0.1% of raw materials quality, the jam-pack fermentation bung, and 48h ferments in constant-temperature shaking incubator.Measure CO
2Weight loss and alcoholic strength are with how many definite each factor optimum values of fermentation rate.
Saccharification pH is on the impact of alcohol getting rate:
Determine that saccharification temperature is 60 ℃, saccharification 20min presses 0.1% of raw material and adds saccharifying enzyme, regulates respectively different pH.Take the ethanol fermentation rate as index, determine that the pH value is 3～5.
The saccharifying enzyme addition is on the impact of ethanol fermentation rate:
Determine that pH is 4, saccharification temperature is 60 ℃, and saccharification 20min presses respectively raw material and adds the different mass saccharifying enzyme.Take the ethanol fermentation rate as index, determine that the saccharifying enzyme addition is 0.05～0.25%.
Saccharification temperature is on the impact of ethanol fermentation rate:
Determine that pH is 4, press 0.1% of raw material and add saccharifying enzyme that saccharification 20min regulates respectively different saccharification temperatures.Take the ethanol fermentation rate as index, determine that saccharification temperature is 50～70 ℃.
Saccharification time is on the impact of alcohol getting rate:
Determine that pH is 4, press 0.1% of raw material and add saccharifying enzyme that saccharification temperature is 60 ℃, respectively the saccharification different time.Take the ethanol fermentation rate as index, determine that saccharification time is 5～30 min.
Quadratic regression rotation composite test
According to the single factor experiment result, take fermentation rate mean value as examination index (establishing 3 repetitions), select pH, saccharifying enzyme addition, saccharification temperature, saccharification time to carry out the test of four factors quadratic regression orthogonal rotational, experimental factor has been carried out the pairwise interaction analysis.
Table 9 quadratic regression rotary combination design table and fermentation rate
The every variance analysis of table 10 regression equation
| The regression variance source | Degree of freedom | Sum of squares | Inequality | The F value | Prob﹥F | Significance |
| Once | 4 | 270.964299 | 0.7567 | 247.27 | <.0001 | ** |
| Quadratic term | 4 | 77.266882 | 0.2158 | 70.51 | <.0001 | ** |
| Mutual | 6 | 8.225415 | 0.0230 | 5.00 | 0.0354 | * |
| Regression model | 14 | 356.456597 | 0.9954 | 92.94 | <.0001 | ** |
| Lose and intend item | 2 | 0.510927 | 0.255464 | 0.90 | 0.4749 | ? |
| Pure error | 4 | 1.132800 | 0.283200 | ? | ? | ? |
| Total error | 6 | 1.643727 | 0.273955 | ? | ? | ? |
| Population variance | 20 | 358.100324 | ? | ? | ? | ? |
Table 11 regression analysis coefficient value and analytical results
| Parameter | Regression coefficient | Standard deviation | The T value | Prob﹥T | Significance |
| Constant | -642.810507 | 82.634686 | -7.78 | 0.0002 | ** |
| p | 75.939927 | 17.947273 | 4.23 | 0.0055 | ** |
| c | 787.577595 | 183.685900 | 4.29 | 0.0052 | ** |
| t | 16.137279 | 2.027221 | 7.96 | 0.0002 | ** |
| h | 2.886443 | 1.836859 | 1.57 | 0.1671 | ? |
| p*p | -11.493046 | 1.524429 | -7.54 | 0.0003 | ** |
| c*p | 62.305556 | 32.117862 | 1.94 | 0.1005 | ? |
| c*c | -1747.304644 | 152.442926 | -11.46 | <.0001 | ** |
| t*p | 0.118056 | 0.205614 | 0.57 | 0.5867 | ? |
| t*c | -8.597222 | 2.056135 | -4.18 | 0.0058 | ** |
| t*t | -0.132330 | 0.015244 | -8.68 | 0.0001 | ** |
| h*p | 0.033056 | 0.321179 | 0.10 | 0.9214 | ? |
| h*c | -8.380556 | 3.211786 | -2.61 | 0.0402 | ** |
| h*t | 0.026250 | 0.020561 | 1.28 | 0.2489 | ? |
| h*h | -0.122330 | 0.015244 | -8.02 | 0.0002 | ** |
Fermentation rate under table 12 optimal conditions under optimal value and the optimal conditions
Carry out proof test, triplicate according to physical condition at optimum extraction conditions (pH value 3～4, saccharifying enzyme addition 0.05～0.15%, 55～65 ℃ of temperature, times 10～20 min).It is 60～70% that the result gets fermentation rate, and trial value and model theory value are very approaching.And the revision test relative deviation is no more than 0～1%, illustrates that saccharification condition circulation ratio is good.
Fermentation condition optimization:
According to the molten condition of the suitableeest alkali, the suitableeest liquefaction condition and the suitableeest saccharification condition, take by weighing that the 20.00g pumpkin powder carries out that alkali is molten, liquefaction, saccharification, with the karusen centrifugation after the saccharification, behind the leavening temperature that supernatant liquor is cooled to set, add the active dry yeast after activating, the jam-pack fermentation bung ferments in constant-temperature shaking incubator.After the fermentation ends, measure CO
2Weight loss and alcoholic strength are with how many definite each factor optimum values of fermentation rate.
Fermentation pH is on the impact of ethanol fermentation rate:
Determine that leavening temperature is 30 ℃, fermentation 48h presses 0.1% of raw material and adds yeast, regulates respectively different pH.Take the ethanol fermentation rate as index, determine that pH is 3.8～5.8.
Inoculum of dry yeast is on the impact of ethanol fermentation rate:
Determine that fermentation pH is 4.8, leavening temperature is 30 ℃, and fermentation 48h presses respectively raw material and adds the different mass yeast.Take the ethanol fermentation rate as index, determine that inoculum of dry yeast is 0.01～0.2%.
Leavening temperature is on the impact of ethanol fermentation rate:
Determine that fermentation pH is 4.8, fermentation 48h presses 0.1% of raw material and adds yeast, regulates respectively the different fermentations temperature.Take the ethanol fermentation rate as index, determine that leavening temperature is 28～32 ℃.
Fermentation time is on the impact of ethanol fermentation rate:
Determine that fermentation pH is 4.8, leavening temperature is 30 ℃, presses 0.1% of raw material and adds yeast, and different time ferments respectively.Take the ethanol fermentation rate as index, determine that fermentation time is 40～80 h.
Second-order regression for rotatable orthogonal design
According to the single factor experiment result, take fermentation rate mean value as examination index (establishing 3 repetitions), select pH, inoculum of dry yeast, leavening temperature, fermentation time to carry out the test of four factors quadratic regression orthogonal rotational, experimental factor has been carried out the pairwise interaction analysis.
Table 13 quadratic regression rotary combination design table and fermentation rate
The every variance analysis of table 14 regression equation
| The regression variance source | Degree of freedom | Sum of squares | Inequality | The F value | Prob﹥F | Significance |
| Once | 4 | 312.202406 | 0.7322 | 243.30 | <.0001 | ** |
| Quadratic term | 4 | 95.753794 | 0.2246 | 74.62 | <.0001 | ** |
| Mutual | 6 | 16.508011 | 0.0387 | 8.58 | 0.0097 | * |
| Regression model | 14 | 424.464211 | 0.9955 | 94.51 | <.0001 | ** |
| Lose and intend item | 2 | 0.762449 | 0.381225 | 1.31 | 0.3647 | ? |
| Pure error | 4 | 1.162320 | 0.290580 | ? | ? | ? |
| Total error | 6 | 1.924769 | 0.320795 | ? | ? | ? |
| Population variance | 20 | 426.38898 | ? | ? | ? | ? |
Table 15 regression analysis coefficient value and analytical results
| Parameter | Regression coefficient | Standard deviation | The T value | Prob﹥T | Significance |
| Constant | -3031.604245 | 417.247599 | -7.27 | 0.0003 | ** |
| p | 96.347120 | 37.776469 | 2.55 | 0.0435 | * |
| c | 773.463429 | 380.640319 | 2.03 | 0.0884 | ? |
| t | 175.109986 | 25.448957 | 6.88 | 0.0005 | ** |
| h | 7.423527 | 1.607719 | 4.62 | 0.0036 | ** |
| p*p | -19.253003 | 1.650239 | -11.67 | <.0001 | ** |
| c*p | 92.768519 | 34.471246 | 2.69 | 0.0360 | * |
| c*c | -1677.300322 | 165.023897 | -10.16 | <.0001 | ** |
| t*p | 2.812500 | 1.112490 | 2.53 | 0.0448 | * |
| t*c | -13.541667 | 11.124903 | -1.22 | 0.2692 | ? |
| t*t | -3.028251 | 0.412560 | -7.34 | 0.0003 | ** |
| h*p | -0.243545 | 0.146853 | -1.66 | 0.1483 | ? |
| h*c | -7.980047 | 1.468534 | -5.43 | 0.0016 | ** |
| h*t | -0.094777 | 0.047007 | -2.02 | 0.0904 | ? |
| h*h | -0.021457 | 0.002882 | -7.45 | 0.0003 | ** |
Fermentation rate under table 16 optimal conditions under optimal value and the optimal conditions
Carry out proof test, triplicate according to physical condition at optimum extraction conditions (pH value 4～5, inoculum of dry yeast 0.05～0.10%, 28～30 ℃ of temperature, times 65～75 h).It is 65～75% that the result obtains fermentation rate, and trial value and model theory value are very approaching, and the revision test relative deviation is no more than 0～1 %, illustrates that the fermentation condition circulation ratio is good.
Continuous production gained Cucurmosin and food fibre part nutritive ingredient and physico-chemical property
Cucurmosin main nutrient composition and part physical and chemical index:
The Cucurmosin that obtains after extraction, freeze-drying is faint yellow, and quality is better, and wherein protein content reaches 70～80%, compares with the raw material pumpkin powder, and food fibre and starch content obviously reduce.
Table 17 preparation gained Cucurmosin and raw material main nutrient composition be (%) relatively
| Sample | Protein | Food fibre | Starch |
| Pumpkin powder | 8.85 | 41.98 | 38.64 |
| Cucurmosin | 70.61±0.65 | 10.28±0.54 | 3.77±0.28 |
Table 18 preparation gained pumpkin protein isolate and isolated soybean protein part physical and chemical index are relatively
| Sample | Retention ability (g/g) | Oil absorbency (g/g) | Viscosity (mPas) |
| Pumpkin | 2.38±0.04 | 2.07±0.05 | 4.2±0.1 |
| Soybean | 4 | 1.54 | 4.5 |
By contrast as can be known, Cucurmosin has good physico-chemical property.At present, soybean protein isolate and soybean protein concentrate are used for meat product in a large number, mainly are exactly the characteristics of utilizing its retentiveness good, reach the purpose of improving quality product, improving the product yield.The oil absorbency of protein plays very important effect in the food formulations such as meat product, milk-product and biscuit sandwich and processing.In the such fluid foodstuffs of beverage, soup, sauce and cream, protein system viscosity and denseness are critical function character, and viscosity is an important indicator of product, and the viscosity of protein is at the characteristic important role of adjusting these food.
Food fibre main nutrient composition and part physical and chemical index:
With water-soluble dietary fibre (SDF) and the water insoluble dietary fiber (IDF) that extraction obtains, freeze-drying is pulverized to merge and is total dietary fiber (TDF).SDF is pale, and it is faint yellow that IDF is.Compare with the raw material pumpkin powder, preparation gained total dietary fiber content reaches 87.36%, compares with the raw material pumpkin powder, and Protein and starch contents obviously reduces.
Table 19 preparation gained food fibre and raw material main nutrient composition be (%) relatively
| Sample | Protein | Food fibre | Starch |
| Pumpkin powder | 8.85 | 41.98 | 38.64 |
| Pumpkin dietary fiber | 0.34±0.02 | 87.36±0.73 | 2.47±0.16 |
Table 20 preparation gained pumpkin dietary fiber and Rhizoma Dioscoreae esculentae dietary fiber physico-chemical property are relatively
| Sample | Retention ability (g/g) | Bulging force (mL/g) | In conjunction with waterpower (g/g) | Oil absorbency (g/g) | Cation exchange capacity (CEC) (mmol/g) | Viscosity (mPas) |
| Pumpkin | 5.00±0.01 | 13±1 | 15.66±0.1 | 1.36±0.02 | 0.58±0.02 | 0.7±0.1 |
| Sweet potato | 8 | 8.5 | 10.4 | 2.4 | - | 1.3 |
By with Rhizoma Dioscoreae esculentae dietary fiber contrast, pumpkin dietary fiber has higher bulging force and in conjunction with waterpower, and retention ability is lower than Rhizoma Dioscoreae esculentae dietary fiber.Food fibre can improve the quality of bakery product by being combined with moisture; The high retentiveness of food fibre can increase volume and the speed of human body defecation, and toxic substance is excreted rapidly.Food fibre becomes large in conjunction with volume after the water, causes satiety, and affects body and can utilize digesting and assimilating of carbohydrate etc. to other compositions of food, and obesity prevention benefits.Pumpkin dietary fiber absorbing grease ability is lower than sweet potato, but also can side direction reduces the secretion of human body bile acide and steroid.
Claims (9)
1. the method for a continuous production Cucurmosin, alcohol fuel and food fibre comprises the discarded pulp of seed-use pumpkin is cleaned, removes the peel, dries, pulverized; It is characterized in that: 1. with the discarded pulp comminuting matter of pumpkin with 1～5% NaOH solution, constant temperature oscillation, centrifugation, filtration under liquid ratio 20:1～30:1,60～80 ℃ of temperature of extraction, extraction times 50～70 min conditions, get filtrate and filter residue; 2. add damping fluid and regulate the pH value to isoelectric points of proteins in gained filtrate, centrifugation must precipitate and supernatant liquor, and washing of precipitate, lyophilize are obtained Cucurmosin; 3. step 2. in centrifugation gained supernatant liquor obtain water-soluble dietary fibre through concentrated, alcohol precipitation; 4. after 1. middle filtration gained filter residue washes with water step, 3:1～8:1 adds entry by liquid ratio, successively with α-amylase insulation liquefaction and saccharifying enzyme insulation saccharification, centrifugation, must precipitate and supernatant liquor, with precipitation wash with water, lyophilize obtains water insoluble dietary fiber; 5. 4. centrifugation gained supernatant liquor active dry yeast constant temperature oscillation fermentation of step, obtain alcohol fuel behind the distillation of gained fermented liquid process, the desiccant dehydration.
2. the method for described a kind of continuous production Cucurmosin, alcohol fuel and food fibre according to claim 1, it is characterized in that the damping fluid that adjusting pH value was used to isoelectric points of proteins during described step 2. is 0.1mol/L acetic acid-sodium acetate solution, the Cucurmosin isoelectric pH value is in 3 ~ 6 scopes.
3. the method for described a kind of continuous production Cucurmosin, alcohol fuel and food fibre according to claim 1 is characterized in that the centrifugation rotating speed is 5000～10000r/min during step 2., in centrifugation time 10～40min scope.
4. the method for described a kind of continuous production Cucurmosin, alcohol fuel and food fibre according to claim 1 is characterized in that acetone and 95% ethanol continuous washing are adopted in washing precipitation during step 2..
5. the method for described a kind of continuous production Cucurmosin, alcohol fuel and food fibre according to claim 1 is characterized in that the condition that supernatant liquor during step 3. prepares water-soluble dietary fibre is pH value 6～8,2～6 times of supernatant liquor volumes of dehydrated alcohol consumption, alcohol precipitation times 30～70 min.
6. the method for described a kind of continuous production Cucurmosin, alcohol fuel and food fibre according to claim 1 is characterized in that α-amylase insulation liquefaction top condition is pH value 4～6, α-amylase addition 0.05～0.15%, 60～80 ℃ of liquefaction temperatures, liquefying time 50～70 min during step 4..
7. the method for described a kind of continuous production Cucurmosin, alcohol fuel and food fibre according to claim 1 is characterized in that saccharifying enzyme insulation saccharification top condition is pH value 3～4, saccharifying enzyme addition 0.05～0.15%, 55～65 ℃ of saccharification temperatures, saccharification time 10～20 min during step 4..
8. the method for described a kind of continuous production Cucurmosin, alcohol fuel and food fibre according to claim 1 is characterized in that during step 5. that supernatant liquor is pH value 4～5, inoculum of dry yeast 0.05～0.10%, 28～30 ℃ of leavening temperatures, fermentation time 65～75 h with the active dry yeast constant temperature oscillation top condition of fermenting.
9. the method for described a kind of continuous production Cucurmosin, alcohol fuel and food fibre according to claim 1, the dehydration siccative that uses is unslaked lime when it is characterized in that during step 5. that fermented liquid is prepared into alcohol fuel.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN104762327A (en) * | 2014-08-11 | 2015-07-08 | 四川安益生物科技有限公司 | Pumpkin yellow pigment yeast product and preparation method thereof |
| CN111184123A (en) * | 2019-12-31 | 2020-05-22 | 西昌市正中食品有限公司 | Process for continuously preparing protein, flavone, syrup and dietary fiber from tartary buckwheat |
| CN113367227A (en) * | 2021-03-23 | 2021-09-10 | 江苏大学 | Method for simultaneously extracting nannochloropsis oculata protein and dietary fiber by using ultrasonic wave assistance |
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| CN1830486A (en) * | 2004-11-05 | 2006-09-13 | 福建医科大学 | Application of pumpkin protein in preparation of medicine |
| CN101195835A (en) * | 2007-12-07 | 2008-06-11 | 李传生 | Production technique for fermenting ethyl alcohol with squash as raw material |
| CN101455240A (en) * | 2008-12-29 | 2009-06-17 | 东北农业大学 | Pumpkin seed oil extraction method using water enzyme method |
| CN102180991A (en) * | 2011-04-06 | 2011-09-14 | 武汉普赛特膜技术循环利用有限公司 | Comprehensive utilization method of pumpkin |
| CN102524698A (en) * | 2012-01-19 | 2012-07-04 | 中国计量学院 | Method for preparing pumpkin dietary fiber |
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| CN1830486A (en) * | 2004-11-05 | 2006-09-13 | 福建医科大学 | Application of pumpkin protein in preparation of medicine |
| CN101195835A (en) * | 2007-12-07 | 2008-06-11 | 李传生 | Production technique for fermenting ethyl alcohol with squash as raw material |
| CN101455240A (en) * | 2008-12-29 | 2009-06-17 | 东北农业大学 | Pumpkin seed oil extraction method using water enzyme method |
| CN102180991A (en) * | 2011-04-06 | 2011-09-14 | 武汉普赛特膜技术循环利用有限公司 | Comprehensive utilization method of pumpkin |
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| CN104762327A (en) * | 2014-08-11 | 2015-07-08 | 四川安益生物科技有限公司 | Pumpkin yellow pigment yeast product and preparation method thereof |
| CN111184123A (en) * | 2019-12-31 | 2020-05-22 | 西昌市正中食品有限公司 | Process for continuously preparing protein, flavone, syrup and dietary fiber from tartary buckwheat |
| CN113367227A (en) * | 2021-03-23 | 2021-09-10 | 江苏大学 | Method for simultaneously extracting nannochloropsis oculata protein and dietary fiber by using ultrasonic wave assistance |
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