CN101455240A - Pumpkin seed oil extraction method using water enzyme method - Google Patents
Pumpkin seed oil extraction method using water enzyme method Download PDFInfo
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Abstract
A method for extracting pumpkin seed oil using a water-enzyme method pertains to the vegetable oil extraction technology, including: using pumpkin seed powders with the particle diameter between 0.3mm to 1.5mm after screening, performing buffering liquid enzymolysis under the enzymolysis condition of 15000-30000U of complex enzyme dosage, 1 to 7 of the solution pH value, 2-8h of extraction time, and 20-60 DEG C of extraction temperature; after demulsification by high-speed centrifugation, adding a water solution into the centrifugal liquid, and then adding a demulsifying agent, using a separating funnel for demixing, adding a desiccant, and then obtaining lipid after centrifugation; or adding an organic solvent in the centrifugate, rotarily evaporating the solvent to obtain lipid. The invention has advantages of high oil yield, good lipid quality, simple process equipment, low consumption, less machining programs, and capability of being easy for the industrial production use.
Description
Technical field
The invention belongs to the vegetable oil extractive technique, relate generally to the pumpkin seed oil extracting method.
Background technology
China's pumpkin planting is in extensive range, and the pumpkin source is abundant.But only have the small part pumpkin to be used to make dry fruit roasted seeds and nuts and health beverages, major part exports to areas such as Eastern Europe, Japan, the resources development and utilization deficiency, and the extraction and application technology of pumpkin seed oil is the difficult problem of restriction pumpkin industrialized developing always.Pumpkin seed oil is rich in unrighted acid, natural complex, phytosterol, the contour bioactive ingredients of linoleic acid, and wherein unsaturated fatty acid content is higher, and oleic acid on average contains 16.59%, and linoleic acid on average contains 39.53%.Utilize pumpkin seed oil effectively to remove interior free yl, have effect anti-ageing, antitumor, that prevent cardio-cerebrovascular diseases.At present perfect not enough for the extractive technique of pumpkin seed oil, majority also adopts methods such as tradition squeezing, organic solvent lixiviate, not only wasted time and energy but also recovery rate not high, also exist to be difficult to problems such as separation, purity be low.How to seek a safety, efficient, save time, labour-saving pumpkin seed oil extraction process is the problem that the present invention will solve.
The tradition squeezing process is its characteristics with high temperature high-moisture steaming stir-fry, the squeezing of high temperature low moisture, pigment dissolves in the oil under higher temperature, moisture in a large number in the oilseeds cell, the material embryo steams at high temperature local burnt the paste is taken place when frying, produce a large amount of melanin, deepened the color and luster of crude oil, cause difficulty to refining, albumen destruction is more in the cake of squeezing back, has reduced the nutritive value of oil cake; Though the ultrasonic extraction method grease can extract greases a large amount of in the oil plant, grease obtained nutrient composition content is relatively low; Supercritical liquid extraction technique, operating pressure is big, and the extraction time is long, and to the requirement height of equipment, energy consumption is big, and output is low, the production cost height.The aqueous enzymatic method oil extracting process is to utilize water as decentralized photo, various enzymes leach grease at this aqueous phase to the degradation of oil plant cell, at non-oil component (protein and carbohydrate) the affine difference of oil and water and profit proportion difference are separated oil with non-oil component.Aqueous enzymatic method is handled the grease obtained quality height of oil plant, mild condition, and capable of reducing energy consumption, minimizing chemical reagent use amount.At present, the research of aqueous enzymatic method oil extracting process is perfect not enough, and problems such as enzymolysis optimum condition, dissolvent residual, emulsification system breakdown of emulsion also need further to research and solve.
The domestic report of research of pumpkin seed oil is less at present, the pumpkin of speaking of among the Chinese invention patent CN1766075A " a kind of method of extracting pumpkin seed oil and pumpkin albumen " to germinate is that raw material carries out defibrination, enzymolysis and extraction pumpkin seed oil, and is different with this patent following points: (1) materials variance.What the CN1766075A patent was emphasized is to extract with the pumpkin of germinateing, germination has certain promotion to various component contents in the pumpkin, but the condition (bud length, temperature, humidity etc.) of germinateing is difficult to control, deal with improperly and can reduce wherein various component contents on the contrary, be not suitable for large-scale production.(2) extracting method difference.The extracting method that the CN1766075A patent adopts is conventional enzyme solution, does not possess the advantage of aqueous enzymatic extraction as previously mentioned.(3) not to grinding particle size, concentration of substrate, enzyme dosage, complex enzyme proportioning, enzymolysis pH value, temperature (℃), time conditions such as (h) carries out orthogonal experiment and determines optimal conditions.(4) also exist between the evaluation index of pumpkin seed oil than big-difference.The patent of external relevant pumpkin seed oil extraction method using water enzyme does not almost have, the more pumpkin seed oil that concentrates on is used for assisting diseases such as treatment benign prostate disease and breast cancer as the additive of food or medicine, U.S.'s patent of invention 09/259,857Dietary supplement containing saw palmetto, pumpkin seed, and nettle root has mentioned and has contained saw palmetto, and the dietary supplements of pumpkin seed oil and hempleaf nettle root can help to keep the health of prostate body of gland.U.S.'s patent of invention 09/921,018Anti-aromatase pharmaceutical composition forcontrolling testosterone/estrone ratios mentions contains hempleaf nettle root, pumpkin seed oil, the anti-fragrance ingredient medicine of extracts such as African cherry bark can be assisted treatment benign prostate disease and breast cancer.
Summary of the invention
Purpose of the present invention is exactly the problem that exists at above-mentioned prior art, studies a kind of method of pumpkin seed oil extraction method using water enzyme, the purpose that reach raising oil yield and oil quality, simplification technical process and equipment, cuts down the consumption of energy.
The object of the present invention is achieved like this:
A kind of method of pumpkin seed oil extraction method using water enzyme comprises pumpkin extraction, cleaning, airing from the melon capsule, pulverizes, sieves, and the back pumpkin powder that 1. sieves directly is controlled in 0.3～1.5mm; 2. at complex enzyme consumption 15000～30000U, pH value of solution value 1～7, extraction time 2～8h, extract buffer solution enzymolysis under the enzymatic hydrolysis condition of 20～60 ℃ of temperature; 3. high speed centrifugation breakdown of emulsion; 4. after adding the aqueous solution in centrifugate, add demulsifier again, the separatory funnel layering adds drier, obtains grease after centrifugal, perhaps in centrifugate, add organic solvent extraction after, the rotary evaporation solvent obtains grease.
The buffer solution that described enzymolysis uses is 0.1mol/L acetate-sodium acetate solution.
The ratio of described enzymolysis liquor capacity ml that is made up of enzyme and buffer solution and pumpkin opaque amount g is in 1～5 scope.
Described complex enzyme is made up of protease, cellulase and pectase, and its best proportioning is controlled in 2:13:22～2:17:30 scope.
Rotating speed is controlled in 3000～10000r/min scope during the high speed centrifugation breakdown of emulsion.
The solvent that the organic solvent extraction pumpkin seed oil is selected for use is a n-hexane.
Described demulsifier is saturated NaCl solution.
Described drier is anhydrous CaCl
2
The pumpkin seed oil extraction method using water enzyme nutrient composition content is abundant, and oil quality is good.Compare with water enzymolysis organic solvent extraction method, water enzymolysis water extracts pumpkin seed oil oil yield descend to some extent (less than 1%).This is because organic solvent has promoted the process of oil plant release grease, makes the easier organic facies that enters of grease, and oil content is separated more easily from albumen and aqueous phase.Adopting water enzymolysis water oil extracting process to extract grease is in order to explore the oil-producing technique of no solvent residue, even it is lower slightly to extract the grease oil yield with water, but to satisfy the health requirement, get rid of dissolvent residual hidden danger, can reduce energy consumption in addition, saving cost.At present, the breaking method of aqueous enzymatic extraction vegetable fat employing has: the centrifugation breakdown of emulsion; Heating demulsification type; Reconcile pH value breakdown of emulsion; The organic solvent extraction breakdown of emulsion.The three kinds of method demulsification efficiency in back are not high, to the nutritive value of grease and security causes a hidden trouble and operation is loaded down with trivial details.This patent adopts the high speed centrifugation breakdown of emulsion (after 3000～10000r/min), the liquid phase after the Separation of Solid and Liquid to be added the saturated NaCl solution of demulsifier assist breakdown of emulsion, can destroy oil, water, protein three's emulsification system fully, reach demulsification.In addition, saturated NaCl solution cost is low, reusable edible, protection environment.Present device is simple, and energy consumption is low, and technology is brief, is easy to industrialization, has broad application prospects.
Description of drawings
Fig. 1 is the process chart that water enzymolysis organic solvent extracts pumpkin seed oil.
Fig. 2 is the process chart that water enzymolysis water extracts pumpkin seed oil.
Fig. 3 n-hexane standard items gas-chromatography collection of illustrative plates.
The residual gas chromatogram spectrum of n-hexane in Fig. 4 sample.
The specific embodiment
Below further specify the present invention with embodiment.
Embodiment 1: the determining of aqueous enzymatic method optimum enzymolysis condition
Determine that aqueous enzymatic method produces the pumpkin granularity of pumpkin seed oil:
Adopt the omnipotent cell pulverization machine of high speed that pumpkin is pulverized 5s (noticing that degree of grinding is not too high), filter out the pumpkin powder that particle diameter is respectively 0.325mm, 0.5mm, 1.0mm, 1.5mm with standard test.Get the raw material 7g of above-mentioned granularity respectively, add the 0.1mol/L acetate-sodium acetate buffer solution of pH value 4.8 with the solid-to-liquid ratio of 1:2.Add 0.5mL cellulase stoste, 50 ℃ of enzymolysis 2h, centrifugal after, add the solvent extraction of 56mL n-hexane, rotary evaporation goes out oil sample, is index with the oil yield after the constant weight, determines that pumpkin carries the suitableeest grinding particle size of oil and be controlled in 0.3～1.5mm.
Determine that aqueous enzymatic method produces the solid-to-liquid ratio of pumpkin seed oil:
0.1mol/L acetate-the sodium acetate buffer solution that in the pumpkin that 7g sieves, adds pH value 4.8 with the solid-to-liquid ratio of 1:1,1:1.5,1:2,1:2.5,1:3.Add 0.5mL cellulase stoste, 50 ℃ of enzymolysis 2h, centrifugal after, add the solvent extraction of 56mL n-hexane, rotary evaporation goes out oil sample.With the oil yield is index, determines that the 0.1mol/L acetate-sodium acetate buffer solution of pH value 4.8 and the best liquid-solid ratio of pumpkin powder (mL:g) are controlled at 1～5 scope.
Determine that aqueous enzymatic method produces the hydrolase of pumpkin seed oil:
Be respectively through testing main nutrient composition and the content of determining pumpkin: crude fat content 49.32 ± 0.11%, protein content 31.42 ± 0.24%, pectin content 1.36 ± 0.14%, content of cellulose 1.73 ± 0.19%, content of starch 0.74 ± 0.28%.Because content of starch is less relatively in the pumpkin, so select protease, pectase and the cellulase enzyme as hydrolysis oil plant cell for use.
Determining of each single enzyme dosage:
Determining of acidic protein enzyme dosage dilutes 1g acid protease enzyme powder and is settled in the volumetric flask of 100mL with the phosphate buffer of 0.02mol/L pH value 5.The enzyme liquid of getting respectively after 1mL, 2mL, 3mL, 4mL, the 5mL dilution is crossed in the pumpkin of 0.5mm aperture sieve in 7g, adds this buffer solution to 21mL, 45 ℃ of enzymolysis 2h, centrifugal after, add the solvent extraction of 56mL n-hexane, rotary evaporation goes out oil sample.With the oil yield is index, and the optimum dose of determining acid protease under this concentration is 1000～8000U scope.
Determining of cellulase consumption, with the 0.1mol/L acetate of pH value 4.8-sodium acetate buffer solution dilution 1.0mL cellulase stoste and constant volume in the 100mL volumetric flask, get 1mL, 2mL, 3mL, 4mL, 5mL, 6mL respectively, 7mL crosses in the pumpkin of 0.5mm aperture sieve in 7g, add this cushioning liquid to 21mL, 50 ℃ of enzymolysis 2h, after centrifugal, add the solvent extraction of 56mL n-hexane, rotary evaporation goes out oil sample.With the oil yield is index, and the optimum dose of determining cellulase under this concentration is 5000～15500U scope.
Determining of pectase consumption, with the 0.1mol/L citric acid of pH value 4-PBS dilution 1.0g pectase powder and constant volume in the 100mL volumetric flask, the enzyme liquid of getting respectively after 1mL, 2mL, 3mL, 4mL, 5mL, the 6mL dilution is crossed in the pumpkin of 0.5mm aperture sieve in 7g, add this cushioning liquid to 21mL, 50 ℃ of enzymolysis 2h, after centrifugal, add the solvent extraction of 56mL n-hexane, rotary evaporation goes out oil sample.With the oil yield is index, and the optimum dose of determining pectase under this concentration is 3000～15500U scope.
Determining of the best proportioning of complex enzyme:
Respectively the experiment of single factor result of three kinds of enzymes being made as the horizontal upper limit of enzyme dosage, is 4 in the pH value, and temperature is enzymolysis 3h under 50 ℃ of conditions, is index with the oil yield, determines that complex enzyme extracts the best proportioning of grease.
The best proportioning orthogonal test of table 1 complex enzyme factor level coding schedule
Table 2 orthogonal test analysis of variance table
By orthogonal experiments, variance analysis as can be known, the hydrolysis of different types of enzyme extraction pumpkin seed oil influences primary and secondary and is pectase, protease, cellulase in proper order, and wherein pectase influences significantly putting forward oily result.By comparing the more excellent horizontal k of each factor
1, k
2, k
3, can get the best proportioning of complex enzyme extraction pumpkin seed oil and be combined as A
1B
1C
3, promptly recovery rate is the highest when protease consumption 1000U, cellulase consumption 8500U, pectase consumption 14500U.To best complex enzyme proportioning combination A
1B
1C
3Carry out demonstration test, the pumpkin oil yield is 37.04%.By the composite hydrolysis effect of enzyme, improved the oil yield of single enzyme hydrolysis extraction pumpkin.
Determining of complex enzyme optimum enzymolysis condition:
For factor designs the experiment of four factors, three horizontal quadratures, is index with the oil yield with complex enzyme consumption, enzymolysis time, temperature, pH value, determines the optimum enzymolysis condition of pumpkin seed oil extraction method using water enzyme.
Table 3 complex enzyme extracts pumpkin seed oil enzymatic hydrolysis condition quadrature result
Table 4 quadrature is analysis of variance table as a result
By orthogonal experiments and extreme difference, variance analysis as can be known, complex enzyme extracts the primary and secondary of pumpkin seed oil hydrolysising condition influence and is in proper order: the pH value of hydrolysis, complex enzyme consumption, hydrolysis temperature, hydrolysis time.By comparing the more excellent horizontal k of each factor
1, k
2, k
3, can get complex enzyme and extract the pumpkin seed oil optimum hydrolysising condition and be: complex enzyme consumption 25500U, be 4 in the pH value, operative temperature is under 40 ℃ the condition, hydrolysis 4h.
Embodiment 2: water enzymolysis organic solvent extractionprocess extracts pumpkin seed oil
Adopt the omnipotent cell pulverization machine of high speed that pumpkin is pulverized 5s (noticing that degree of grinding is not too high), get the pumpkin powder that 7g crosses the 0.5mm aperture sieve, add complex enzyme 25500U, add 0.1mol/L acetate-sodium acetate buffer solution again to 21mL, in the pH value is 4,40 ℃ of enzymolysis 4h, with gained solution high speed centrifugation (3000～10000r/min) breakdowns of emulsion behind the enzymolysis, in centrifugate, add 56mL n-hexane solvent, extract 3～4 times, combining extraction liquid, rotary evaporation goes out oil sample, and the pumpkin oil yield is 38.34% after the constant weight.
The residual mensuration of n-hexane in the check water enzymolysis organic solvent oil extracting methods.
The 20g pumpkin is after water enzymolysis organic solvent extracts grease, and rotary evaporation gets grease 8.7837g.Be placed in 50 ℃ of baking ovens, every the 30min weighing, the constant weight time of determining residual solvent is 4h.
The result of the test that the table 5 evaporate residual solvent time is determined
The n-hexane peak area is 7.5 in the sample, and substitution n-hexane calibration curve draws water enzymolysis organic solvent method and extracts the residual 46.60mg/kg of being of pumpkin seed oil solvent n-hexane.Regulation edible vegetable oil leaches oil solvent in the State Standard of the People's Republic of China GB2716-2005 edible vegetable oil sanitary standard residual≤50mg/kg.So wood experiment water enzymolysis organic solvent extracts pumpkin seed oil dissolvent residual index and meets national standard.
Embodiment 3: water enzymolysis water law is extracted pumpkin seed oil
Adopt the omnipotent cell pulverization machine of high speed that pumpkin is pulverized 5s (noticing that degree of grinding is not too high), get the pumpkin powder that 7g crosses the 0.5mm aperture sieve, add complex enzyme 25500U, add 0.1mol/L acetate-sodium acetate buffer solution again to 21mL, in the pH value is 4,40 ℃ of enzymolysis 4h, with gained solution high speed centrifugation (3000～10000r/min) breakdowns of emulsion behind the enzymolysis, in centrifugate, add the extraction with aqueous solution grease, add the saturated NaCl solution of demulsifier 5～15mL, extract 3～4 times, in the gained oil phase, add the anhydrous CaCl of 0.1～0.3g
2Drying obtains clarification, yellow green grease after centrifugal, and the pumpkin oil yield is 37.40%.Take by weighing the pretreated pumpkin of 100g, enlarge experiment by above-mentioned water enzymolysis water oil extracting process condition, grease obtained quality is constant, and oil yield is 37.98%.
Pumpkin seed oil extraction method using water enzyme, the measurement result of part nutritive index and physical and chemical index is as shown in table 6:
The measurement result of table 6 pumpkin seed oil part nutritive index and physical and chemical index
Claims (8)
1. the method for a pumpkin seed oil extraction method using water enzyme comprises pumpkin extraction, cleaning, airing from the melon capsule, pulverizes, sieves, and it is characterized in that: the back pumpkin powder that 1. sieves directly is controlled in 0.3～1.5mm; 2. at complex enzyme consumption 15000～30000U, pH value of solution value 1～7, extraction time 2～8h, extract buffer solution enzymolysis under the enzymatic hydrolysis condition of 20～60 ℃ of temperature; 3. high speed centrifugation breakdown of emulsion; 4. after adding the aqueous solution in centrifugate, add demulsifier again, the separatory funnel layering adds drier, obtains grease after centrifugal, perhaps in centrifugate, add organic solvent extraction after, the rotary evaporation solvent obtains grease.
2. according to the method for the described pumpkin seed oil extraction method using water enzyme of claim 1, it is characterized in that the buffer solution that described enzymolysis uses is 0.1mol/L acetate-sodium acetate solution.
3. according to the method for the described pumpkin seed oil extraction method using water enzyme of claim 1, the ratio that it is characterized in that described enzymolysis liquor capacity ml that is made up of enzyme and buffer solution and pumpkin opaque amount g is in 1～5 scope.
4. according to the method for the described pumpkin seed oil extraction method using water enzyme of claim 1, it is characterized in that described complex enzyme is made up of protease, cellulase and pectase, its best proportioning is controlled in 2:13:22～2:17:30 scope.
5. according to the method for the described pumpkin seed oil extraction method using water enzyme of claim 1, rotating speed is controlled in 3000～10000r/min scope when it is characterized in that the high speed centrifugation breakdown of emulsion.
6. according to the method for the described pumpkin seed oil extraction method using water enzyme of claim 1, it is characterized in that the solvent that the organic solvent extraction pumpkin seed oil is selected for use is a n-hexane.
7. according to the method for the described pumpkin seed oil extraction method using water enzyme of claim 1, it is characterized in that described demulsifier is saturated NaCl solution.
8. according to the method for the described pumpkin seed oil extraction method using water enzyme of claim 1, it is characterized in that described drier is anhydrous CaCl
2
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