CN103408964B - Method for improving extraction yield of melanin in Vaccinium bracteatum Thunb by using polyphenol oxidase and tea polyphenol - Google Patents
Method for improving extraction yield of melanin in Vaccinium bracteatum Thunb by using polyphenol oxidase and tea polyphenol Download PDFInfo
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Abstract
The invention provides a method for improving extraction yield of melanin in Vaccinium bracteatum Thunb by using polyphenol oxidase and tea polyphenol, which relates to a preparation and extraction method for a natural pigment. The method comprises the following steps: with leaves of Vaccinium bracteatum Thunb as a main raw material, carrying out beating, releasing enzymes like endogenous polyphenol oxidase and peroxidase which are beneficial for formation of melanin and releasing precursor substances like phenolic compounds which form melanin at the same time; then adding exogenous polyphenol oxidase or further adding tea polyphenol to accelerate the process of enzymatic oxidation and adjust chroma so as to realize quicker obtainment of more melanin; and finally heating and extracting melanin by using an ethanol solution with a certain concentration and carrying out filtration, condensation and drying so as to obtain a melanin product. According to the invention, the method is simple; natural melanin obtained by using the method has stable quality and good security; the method enables the extraction rate of melanin to be increased by 30 to 90% compared with a water extraction method; and based on changes of the usage amount of tea polyphenol, the color of a finished pigment product covers a tonal range consisting of a blue black color, a pure black color and a reddish black color, greatly broadening the application scope of natural melanin in Vaccinium bracteatum Thunb.
Description
Technical field
The invention provides a kind of method of utilizing polyphenoloxidase and tea-polyphenol to improve melanochrome extraction efficiency yield in Fructus Vaccinii Bracteati, is a kind of preparation and extracting method of natural pigment.
Background technology
Fructus Vaccinii Bracteati belongs to Ericaceae genus vaccinium plant, and China south of the River one is with the custom that has the black meal that edible its tender leaf dipping glutinous rice is cooked among the people.Owing to can forming stable black, therefore myrtillin leave is also subject to the attention of academic and industry member.The research of the chemical composition in relevant myrtillin leave and pigment About Coloration Mechanisms of Fancy thereof has had many decades, although study to such an extent that be not very thorough, basal component, pigment synthesis approach and pigments have had some preliminary conclusions: the chemical composition containing in Vaccinium bracteatum Thunb. Leaves has hentriacontane, friedelin (ketone), epifriedelanol, β-sitosterol, Quercetin, isorientin, Hydroxycinnamic acid, meso-inositol, ursolic acid, Oleanolic Acid and abundant amino acid and organic acid etc.Wherein quercetin content is up to 3.52% of extract, and flavones total amount accounts for the more than 11.64% of extract.Myrtillin leave pigment extract all has absorption under 286nm and 309nm, has and infers to think it may is anthocyanin class material; Also someone thinks that the main component of its pigment is cyanogen indigo-3-glucose glycosides or iridoid.More stable when the pigment extract of leaf and foodstuff additive coexist as NaCl, vitamins C and phenylformic acid, and can generate brown precipitation while coexisting with FeCl3, prompting cannot be stored with ferrous products.This pigment is stable under acidic conditions, and the next black that can make of condition of alkaline condition and heating is deepened.
The extracting method of this pigment is mainly solvent-extraction process at present.The patent of Chen Shaojun etc. 2005 the application method of extraction of natural melanin " a kind of from myrtillin leave " (200510104210.6), take natural phant myrtillin leave as main raw material, adopt the zymin assisted extraction such as polygalacturonase, cellulase, finally adopt ethanolic soln extraction of natural melanin.Patent " the extraction of flavonoid monomer in oriental blueberry melanin of the application in 2006 such as Wang Li, separated, purifying and evaluation " (application number: 200610040413.8), take Vaccinium bracteatum Thunb. Leaves as raw material, also be to adopt extraction using alcohol, melanochrome solution will be obtained after extraction using alcohol, through macroporous adsorbent resin, the flavonoid compound in melanochrome is carried out to roughing out, by polyamide column and HW-40 post, flavonoid compound monomer is carried out to purifying again, use again High Performance Liquid Chromatography/Mass Spectrometry, nuclear magnetic resonance technique carries out Structural Identification to purified components, therefrom separation has obtained 7 kinds of flavonoid compound monomers, comprise Quercetin, chrysin, 4',5,7-trihydroxyflavone, kaempferol and luteolin, its relative content is respectively: 37.51%, 2.26%, 9.57%, 1.72% and 15.16%.The patent " a kind of method of extracting pigments from bracteatum leaves " (application number 201110264827.X) of the application in 2011 such as Liu Dongfeng, after dry myrtillin leave is pulverized, add 2-5 doubly to measure 5% acetum and soak 2-4 hour, extract 2-4 time, after centrifugation, further adopt ultrafiltration, nanofiltration, obtain concentrated solution, add water-dispersion, upper polyamide column absorption, first wash decon with water, use 5-7BV50% ethanolic soln wash-out, elutriant is concentrated again, and lyophilize obtains myrtillin leave pigment.
Zhou Zhaoxiang etc. just reported that water is that extraction agent extracts the melanochrome of myrtillin leave as far back as 1989 in addition, obtained water miscible myrtillin leave melanochrome (food and fermentation industries, 1989(6), 54-59).Gu Wenxiu etc. (2005) are at < < chemistry of forest product and industrial > > (2005,25(4), 74-78) reported and adopted 62% ethanol to obtain myrtillin leave melanochrome with ultrasonic auxiliary extraction method.
Yet Wei state China waits 2011 at < < Food Additives Used in China > > (2011,4,135-138) report thinks that oxidative fermentation process is the prerequisite that oriental blueberry melanin forms; Resin is the better selection of purifying oriental blueberry melanin; Can not measure with the content of Flavonoid substances the content of oriental blueberry melanin.Based on this thought, Wei state China waits the patent (application number 201110097948.X) of having applied for " the melanic manufacture method of a kind of water miscible plant " for 2011, different with the myrtillin leave melanochrome extraction thinking of previous patent report, the method be take myrtillin leave and is extracted melanin precursor as raw material adopts water-containing organic solvent at a certain temperature, utilize ultra-filtration membrane to remove after macromolecular components, obtain water-soluble small molecules melanin precursor composition, add polyphenol oxidase catalyzed oxidation, after ultrafiltration membrance filter, obtain again melanochrome crude product, through macroporous resin purification, spraying is dried and obtains sterling again.
Although the research of domestic oriental blueberry melanin is lasting for years, but up to this point also do not form more successful industrialization, trace it to its cause is largely because the preparation method of oriental blueberry melanin goes back utmost point imperfection, Many researchers is just the separation of vat liquor evaporate to dryness, the product of gained or be medicinal extract, and color and luster and content are all unstable, or product is that melanin precursor is as flavonoid or other compounds, still be not enough to be directly used in dyeing, or preparation technology's flow process is very long, is difficult to industrialization.
Summary of the invention
The present invention, just in order to overcome above-mentioned deficiency, provides a kind of method of utilizing polyphenoloxidase and tea-polyphenol to improve melanochrome extraction efficiency yield in Fructus Vaccinii Bracteati.
Technical scheme of the present invention: take myrtillin leave as main raw material, first making beating, endogenous polyphenoloxidase and peroxidase etc. are contributed to form melanic enzyme to be discharged, the melanic precursor substances of formation such as phenolic compound are discharged simultaneously, then add external source polyphenoloxidase or further add tea-polyphenol, make enzymatic oxidation process be accelerated and adjust colourity, more melanochrome that obtains sooner, finally adopt the heating of finite concentration ethanolic soln to extract melanochrome, after filtering, concentrate and being dried, obtain melanochrome product, step is:
(1) making beating raw materials: get appropriate fresh or dry myrtillin leave, add water making beating, cross colloidal mill circular treatment, the slurry obtaining is placed in not higher than placing 1-2 hour under 40 ° of C temperature condition, promotes melanochrome to form;
(2) add tea-polyphenol and adjust product colourity: slurry is required to add tea-polyphenol according to the finished product color, higher than 40 ° of C temperature condition, do not carrying out insulated and stirred reaction, the input amount of tea-polyphenol is 0 ~ 100%(W/W of total phenol quality in slurry);
(3) enzyme-added auxiliary catalysis oxidation: by the slurry through making beating or added the mixture of tea-polyphenol, according to Folin-Ciocalteu method, record total phenol content in slurries and drop into polyphenoloxidase, the input amount of polyphenoloxidase is 1 ~ 10%(W/W of total phenol quality in slurries), higher than 50 ° of C temperature condition, do not carrying out insulated and stirred reaction;
(4) extraction using alcohol: according to the volume of the slurry after catalyzed oxidation, add food grade ethanol and extract, filter extracting solution, obtain melanochrome crude product solution;
(5) dry: melanochrome crude product solution to be reclaimed to ethanol under reduced pressure, through spraying, be dried and obtaining oriental blueberry melanin powder.
In step (1), the fresh myrtillin leave of per kilogram adds water 1-5L making beating, if dry myrtillin leave, the dry myrtillin leave of per kilogram adds water 10-20L making beating, after making beating, slurry is through 2-3 circular treatment of colloidal mill, and the processing fineness of colloidal mill is resized to 2-5 micron; Slurry through colloidal mill is placed 1 ~ 2 hour under 25 ~ 40 ° of C conditions, promotes melanochrome to form.
In step (2), partially black-and-blue if pigment requires, the input amount of tea-polyphenol is 0 ~ 10%(W/W of total phenol quality in slurry); If pigment requires inclined to one side black, the input amount of tea-polyphenol is 10 ~ 40%(W/W of total phenol quality in slurry); If pigment requires inclined to one side reddish black, the input amount of tea-polyphenol is 40 ~ 100%(W/W of total phenol quality in slurry).
In step (3), the specific activity of added polyphenoloxidase is more than 10000 units/g; The optimum temperuture of catalytic oxidation is 25 ~ 50 ° of C, and the reaction times is 2 ~ 6 hours.
In step (4), add food grade ethanol, make final alcohol concn reach 30 ~ 60%, the volume ratio of myrtillin leave slurries and ethanol extract is not less than 1:0.8 ~ 1.5; Extraction is to carry out under not higher than 60 ° of C temperature condition, and extraction time is 2-6 hour; After extraction finishes, adopt expression separation; It is the ethanol of 30-60% that separated rear residue adds concentration, and the mass volume ratio of the ethanol of residue dry substance quality and second extraction is 1:6 ~ 10; The time of extracting is for the second time 1-2 hour, and temperature is similarly not higher than 60 ° of C; United extraction liquid, filters, and obtains melanochrome crude product solution.
In step (5), reduced pressure is 0.09MPa, 60 ° of C, and the product of sloughing after ethanol is dried and obtains oriental blueberry melanin powder through spraying.Product yield accounts for prophyll dry weight in 26.1% ~ 36%(yield with product powder dry weight and calculates), depending on the difference of extraction conditions and difference.
The present invention is based on myrtillin leave itself has melanochrome substrate and promotes a kind of like this understanding of enzyme that melanochrome forms, endogenous enzyme and substrate-function have been proposed first to utilize, recycling external source polyphenoloxidase is strengthened melanic formation, then utilize external source phenolic compound strengthening matrix effect and product tone is modified a little, effectively promoted and take melanochrome output and the product stability that myrtillin leave is main material production, improved the problem that myrtillin leave medicinal extract color and luster content is unstable or technological process is long in the past, and promoted the using value of myrtillin leave melanochrome product.Up to now, not yet utilize endogenous enzyme strengthening melanochrome to form, the report that utilizes external source phenolic compound strengthening matrix effect and product tone is modified.
Myrtillin leave powder prepared by the present invention, change based on production process tea-polyphenol add-on, can form based on myrtillin leave originally blue-black from blue-black to black to the product line of red-black different tones, according to the difference of application, be adapted to the needs of the foods prods of different colours demand.
Oriental blueberry melanin powder-product prepared by the inventive method, yield is high, yield 26.1% ~ 36%(yield accounts for prophyll dry weight with product powder dry weight and calculates), depending on the difference of extraction using alcohol condition and difference.Owing to also containing a certain amount of polyphenols in myrtillin leave melanochrome product, can play anti-oxidant and remove free radical and antibacterial effect, can be attributed to a kind of functional foodstuff tinting material, the suitable Worth Expecting of its application prospect.
The natural black pigment that utilizes the present invention to obtain, technique is simple, constant product quality security is good, melanochrome extraction yield improves 30-90% compared with water extraction, according to the variation of tea-polyphenol consumption, final finished pigmentary colours can cover blue-black, black and red-black three tone ranges, have widened well the range of application of natural oriental blueberry melanin.
Embodiment
Embodiment 1
Lyophilize myrtillin leave, according to Folin-Ciocalteu method, measuring total phenol content is that 3.72%(moisture content is 3.55%), accurately take and get the cryodesiccated myrtillin leave of 50g and measure and add 500mL water (i.e. 10 times of water), with tissue mashing machine, smash to pieces, adopt again colloidal mill (gap is adjusted to 2 microns), carry out further fragmentation, carry out 2 circulations, whole shattering process notices that temperature does not surpass 40 ° of C; Place at ambient temperature 2 hours.
Toward adding specific activity in slurry, be more than 10000 units/g polyphenoloxidase 0.093g(5%, w/w), mix, under 35 ° of C conditions, stirring reaction is 4 hours.
After reaction finishes, in reacting slurry, add 95%(v/v) ethanol 750mL(Fructus Vaccinii Bracteati slurries and the volume ratio of ethanol extract be 1:1.5, alcohol concn is finally 57%, v/v), under 35 ° of C conditions, extract 2 hours, expression separation, it is 1:7.5 that filter residue adds the mass volume ratio of the ethanol of 50% ethanolic soln 300mL(residue dry substance quality and second extraction, final alcohol concn is 53%), under 55 ° of C conditions, extract again 1 hour, after expression separation, united extraction liquid, filter, obtain melanochrome crude product solution 1325mL.
Melanochrome crude product solution is reclaimed to ethanol under reduced pressure, and reduced pressure is 0.09MPa, 60 ° of C, and the product of sloughing after ethanol is dried and obtains oriental blueberry melanin powder 13.1g through spraying, and product yield is 26.1%.This product colour tone is black-and-blue.
Embodiment 2
Lyophilize myrtillin leave, according to Folin-Ciocalteu method, measuring total phenol content is that 3.72%(moisture content is 3.55%), accurately take and get the cryodesiccated myrtillin leave of 50g and measure and add 600mL water (i.e. 12 times of water), with tissue mashing machine, smash to pieces, adopt again colloidal mill (gap is adjusted to 3 microns), carry out further fragmentation, carry out 2 circulations, whole shattering process notices that temperature is 25 ° of C; Place at ambient temperature 2 hours.
Toward add in slurry tea-polyphenol powder (polyphenol content 80%) 0.23g(be equivalent to total phenol content in slurry 10%), under room temperature, stir.
Toward adding specific activity in the above-mentioned slurry that is mixed with tea-polyphenol, be more than 10000 units/g polyphenoloxidase 0.02g(1%, w/w), mix, under 50 ° of C conditions, stirring reaction is 2 hours.
After reaction finishes, in reacting slurry, add 75%(v/v) ethanol 500mL(Fructus Vaccinii Bracteati slurries and the volume ratio of ethanol extract be 1:0.8, alcohol concn is finally 34%, v/v), under 50 ° of C conditions, extract 6 hours, expression separation, filter residue adds 80% ethanolic soln 250mL, and (mass volume ratio of the ethanol of residue dry substance quality and second extraction is 1:6, final alcohol concn is 60%), under 50 ° of C conditions, extract again 2 hours, after expression separation, united extraction liquid, filter, obtain melanochrome crude product solution 1200mL.
Melanochrome crude product solution is reclaimed to ethanol under reduced pressure, and reduced pressure is 0.09MPa, 60 ° of C, and the product of sloughing after ethanol is dried and obtains oriental blueberry melanin powder 14.3g through spraying, and product yield is 28.6%.This product colour tone is black-and-blue.
Embodiment 3
Lyophilize myrtillin leave, according to Folin-Ciocalteu method, measuring total phenol content is that 3.72%(moisture content is 3.55%), accurately take and get the cryodesiccated myrtillin leave of 50g and measure and add 750mL water (i.e. 15 times of water), with tissue mashing machine, smash to pieces, adopt again colloidal mill (gap is adjusted to 4 microns), carry out further fragmentation, carry out 3 circulations, whole shattering process temperature is 35 ° of C; Place at ambient temperature 1 hour.
Toward add in slurry tea-polyphenol powder (polyphenol content 80%) 0.58g(be equivalent to total phenol content in slurry 25%), under room temperature, stir.
Toward adding specific activity in the above-mentioned slurry that is mixed with tea-polyphenol, be more than 10000 units/g polyphenoloxidase 0.116g(5%, w/w), mix, under 35 ° of C conditions, stirring reaction is 6 hours.
After reaction finishes, in reacting slurry, add 95%(v/v) ethanol 650mL(Fructus Vaccinii Bracteati slurries and the volume ratio of ethanol extract be 1:0.9, alcohol concn is finally 44%, v/v), under 35 ° of C conditions, extract 2 hours, expression separation, it is 1:7.5 that filter residue adds the mass volume ratio of the ethanol of 50% ethanolic soln 300mL(residue dry substance quality and second extraction, final alcohol concn is 46%), under 60 ° of C conditions, extract again 1 hour, after expression separation, united extraction liquid, filter, obtain melanochrome crude product solution 1480mL.
Melanochrome crude product solution is reclaimed to ethanol under reduced pressure, and reduced pressure is 0.09MPa, 60 ° of C, and the product of sloughing after ethanol is dried and obtains oriental blueberry melanin powder 14.4g through spraying, and product yield is 28.8%.This product colour tone is black.
Embodiment 4
Lyophilize myrtillin leave, according to Folin-Ciocalteu method, measuring total phenol content is that 3.72%(moisture content is 3.55%), accurately take and get the cryodesiccated myrtillin leave of 50g and measure and add 900mL water (i.e. 18 times of water), with tissue mashing machine, smash to pieces, adopt again colloidal mill (gap is adjusted to 5 microns), carry out further fragmentation, carry out 3 circulations, whole shattering process notices that temperature does not surpass 40 ° of C; Place at ambient temperature 1.5 hours.
Toward add in slurry tea-polyphenol powder (polyphenol content 80%) 0.88g(be equivalent to total phenol content in slurry 38%), under room temperature, stir.
Toward adding specific activity in the above-mentioned slurry that is mixed with tea-polyphenol, be more than 10000 units/g polyphenoloxidase 0.128g(5%, w/w), mix, under 25 ° of C conditions, stirring reaction is 2 hours.
After reaction finishes, the volume ratio that adds dehydrated alcohol 1350mL(Fructus Vaccinii Bracteati slurries and ethanol extract in reacting slurry is 1:1.5, alcohol concn is finally 60%, v/v), under 35 ° of C conditions, extract 2 hours, expression separation, it is 1:8.5 that filter residue adds the mass volume ratio of the ethanol of 15% ethanolic soln 350mL(residue dry substance quality and second extraction, final alcohol concn 30%), under 50 ° of C conditions, extract again 2 hours, after expression separation, united extraction liquid, filter, obtain melanochrome crude product solution 2430mL.
Melanochrome crude product solution is reclaimed to ethanol under reduced pressure, and reduced pressure is 0.09MPa, 60 ° of C, and the product of sloughing after ethanol is dried and obtains oriental blueberry melanin powder 15.8g through spraying, and product yield is 31.6%.This product colour tone is black.
Embodiment 5
Lyophilize myrtillin leave, according to Folin-Ciocalteu method, measuring total phenol content is that 3.72%(moisture content is 3.55%), accurately take and get the cryodesiccated myrtillin leave of 50g and measure and add 1000mL water (i.e. 20 times of water), with tissue mashing machine, smash to pieces, adopt again colloidal mill (gap is adjusted to 2 microns), carry out further fragmentation, carry out 2 circulations, whole shattering process notices that temperature does not surpass 40 ° of C; Place at ambient temperature 2 hours.
Toward add in slurry tea-polyphenol powder (polyphenol content 80%) 1.74g(be equivalent to total phenol content in slurry 75%), under room temperature, stir.
Toward adding specific activity in the above-mentioned slurry that is mixed with tea-polyphenol, be more than 10000 units/g polyphenoloxidase 0.244g(7.5%, w/w), mix, under 50 ° of C conditions, stirring reaction is 6 hours.
After reaction finishes, in reacting slurry, add 95%(v/v) ethanol 1300mL(Fructus Vaccinii Bracteati slurries and the volume ratio of ethanol extract be 1:1.3, alcohol concn is finally 53%, v/v), under 50 ° of C conditions, extract 6 hours, expression separation, filter residue adds 30% ethanolic soln 400mL, (mass volume ratio of the ethanol of residue dry substance quality and second extraction is 1:10, final alcohol concn is 38%) under 60 ° of C conditions, extract again 2 hours, after expression separation, united extraction liquid, filter, obtain melanochrome crude product solution 2520mL.
Melanochrome crude product solution is reclaimed to ethanol under reduced pressure, and reduced pressure is 0.09MPa, 60 ° of C, and the product of sloughing after ethanol is dried and obtains oriental blueberry melanin powder 17.3g through spraying, and product yield is 34.6%.This product colour tone is reddish black.
Embodiment 6
Fresh myrtillin leave, according to Folin-Ciocalteu method, measuring total phenol content is that 0.58%(moisture content is 83.72%), accurately take and get the fresh myrtillin leave of 500g and measure and add 1000mL water (being equivalent to per kilogram raw material 2L), with tissue mashing machine, smash to pieces, adopt again colloidal mill (gap is adjusted to 2 microns), carry out further fragmentation, carry out 2 circulations, whole shattering process notices that temperature does not surpass 40 ° of C; Place at ambient temperature 2 hours.
Toward add in slurry tea-polyphenol powder (polyphenol content 80%) 3.6g(be equivalent to total phenol content in slurry 100%), under room temperature, stir.
Toward adding specific activity in the above-mentioned slurry that is mixed with tea-polyphenol, be more than 10000 units/g polyphenoloxidase 0.58g(10%, w/w), mix, under 50 ° of C conditions, stirring reaction is 4 hours.
After reaction finishes, in reacting slurry, add 95%(v/v) ethanol 1500mL(Fructus Vaccinii Bracteati slurries and the volume ratio of ethanol extract be 1:1, alcohol concn is finally 47%, v/v), under 50 ° of C conditions, extract 4 hours, expression separation, it is 1:9 that filter residue adds the mass volume ratio of the ethanol of 30% ethanol 150mL(residue dry substance quality and second extraction, final alcohol concn is 37%), under 60 ° of C conditions, extract again 2 hours, after expression separation, united extraction liquid, filter, obtain melanochrome crude product solution 2620mL.
Melanochrome crude product solution is reclaimed to ethanol under reduced pressure, and reduced pressure is 0.09MPa, 60 ° of C, and the product of sloughing after ethanol is dried and obtains oriental blueberry melanin powder 7.5g through spraying, and product yield is 35.1%.This product colour tone is reddish black.
Embodiment 7
Fresh myrtillin leave, according to Folin-Ciocalteu method, measuring total phenol content is that 0.58%(moisture content is 83.72%), accurately take and get the fresh myrtillin leave of 500g and measure and add 500mL water (being equivalent to per kilogram 1L), with tissue mashing machine, smash to pieces, adopt again colloidal mill (gap is adjusted to 2 microns), carry out further fragmentation, carry out 2 circulations, whole shattering process notices that temperature does not surpass 40 ° of C; Place at ambient temperature 2 hours.
Toward add in slurry tea-polyphenol powder (polyphenol content 80%) 0.72g(be equivalent to total phenol content in slurry 20%), under room temperature, stir.
Toward adding specific activity in the above-mentioned slurry that is mixed with tea-polyphenol, be more than 10000 units/g polyphenoloxidase 0.18g(5%, w/w), mix, under 35 ° of C conditions, stirring reaction is 2 hours.
After reaction finishes, in reacting slurry, add 95%(v/v) ethanol 1500mL(Fructus Vaccinii Bracteati slurries and the volume ratio of ethanol extract be 1:1.5, alcohol concn is finally 57%, v/v), under 40 ° of C conditions, extract 6 hours, expression separation, it is 1:10 that filter residue adds the mass volume ratio of the ethanol of 25% ethanol 200mL(residue dry substance quality and second extraction, final alcohol concn is 35%), under 50 ° of C conditions, extract again 2 hours, after expression separation, united extraction liquid, filter, obtain melanochrome crude product solution 2570mL.
Melanochrome crude product solution is reclaimed to ethanol under reduced pressure, and reduced pressure is 0.09MPa, 60 ° of C, and the product of sloughing after ethanol is dried and obtains oriental blueberry melanin powder 5.1g through spraying, and product yield is 29.9%.This product colour tone is black.
Embodiment 8
Fresh myrtillin leave, according to Folin-Ciocalteu method, measuring total phenol content is that 0.58%(moisture content is 83.72%), accurately take and get the fresh myrtillin leave of 500g and measure and add 2500mL water (being equivalent to per kilogram 5L), with tissue mashing machine, smash to pieces, adopt again colloidal mill (gap is adjusted to 2 microns), carry out further fragmentation, carry out 2 circulations, whole shattering process notices that temperature does not surpass 40 ° of C; Place at ambient temperature 2 hours.
Toward adding specific activity in slurry, be more than 10000 units/g polyphenoloxidase 0.03g(1%, w/w), mix, under 40 ° of C conditions, stirring reaction is 6 hours.
After reaction finishes, in reacting slurry, add 70%(v/v) ethanol 2400mL(Fructus Vaccinii Bracteati slurries and the volume ratio of ethanol extract be 1:0.8, alcohol concn is finally 30%, v/v), under 60 ° of C conditions, extract 2 hours, expression separation, it is 1:6 that filter residue adds the mass volume ratio of the ethanol of dehydrated alcohol 100mL(residue dry substance quality and second extraction, final alcohol concn is 60%), under 60 ° of C conditions, extract again 1 hour, after expression separation, united extraction liquid, filter, obtain melanochrome crude product solution 5410mL.
Melanochrome crude product solution is reclaimed to ethanol under reduced pressure, and reduced pressure is 0.09MPa, 60 ° of C, and the product of sloughing after ethanol is dried and obtains oriental blueberry melanin powder 4.7g through spraying, and product yield is 26.7%.This product colour tone is black-and-blue.
Claims (9)
1. utilize polyphenoloxidase and tea-polyphenol to improve a method for melanochrome extraction yield in Fructus Vaccinii Bracteati, it is characterized in that manufacturing step is:
(1) making beating raw materials: get appropriate fresh or dry myrtillin leave, add water making beating, cross colloidal mill circular treatment, the slurry obtaining is placed in not higher than placing 1-2 hour under 40 ° of C temperature condition, promotes melanochrome to form;
(2) add tea-polyphenol and adjust product colourity: slurry is required to add tea-polyphenol according to the finished product color, higher than 40 ° of C temperature condition, do not carrying out insulated and stirred reaction, the input amount of tea-polyphenol is 0 ~ 100%(W/W of total phenol quality in slurry) and tea-polyphenol input amount be not 0;
(3) enzyme-added auxiliary catalysis oxidation: by the slurry through making beating or added the mixture of tea-polyphenol, according to Folin-Ciocalteu method, record total phenol content in slurries and drop into polyphenoloxidase, the input amount of polyphenoloxidase is 1 ~ 10%(W/W of total phenol quality in slurries), higher than 50 ° of C temperature condition, do not carrying out insulated and stirred reaction;
(4) extraction using alcohol: according to the volume of the slurry after catalyzed oxidation, add food grade ethanol and extract, filter extracting solution, obtain melanochrome crude product solution;
(5) dry: melanochrome crude product solution to be reclaimed to ethanol under reduced pressure, through spraying, be dried and obtaining oriental blueberry melanin powder.
2. polyphenoloxidase and the tea-polyphenol of utilizing according to claim 1 improves the method that melanochrome in Fructus Vaccinii Bracteati extracts yield, it is characterized in that in step (1), the fresh myrtillin leave of per kilogram adds water 1-5L making beating or the dry myrtillin leave of per kilogram adds water 10-20L making beating.
3. polyphenoloxidase and the tea-polyphenol of utilizing according to claim 2 improves the method that melanochrome in Fructus Vaccinii Bracteati extracts yield, it is characterized in that in step (1), after making beating, slurry is through 2-3 circular treatment of colloidal mill, the processing fineness of colloidal mill is resized to 2-5 micron.
4. the method for utilizing polyphenoloxidase and tea-polyphenol to improve melanochrome extraction yield in Fructus Vaccinii Bracteati according to claim 1, is characterized in that in step (1), the slurry through colloidal mill is placed 1 ~ 2 hour under 25 ~ 40 ° of C conditions, promotes melanochrome to form.
5. polyphenoloxidase and the tea-polyphenol of utilizing according to claim 1 improves the method that melanochrome in Fructus Vaccinii Bracteati extracts yield, it is characterized in that in step (2), it is partially black-and-blue that pigment requires, and the input amount of tea-polyphenol is 0 ~ 10%(W/W of total phenol quality in slurry) and tea-polyphenol input amount be not 0; Pigment requires inclined to one side black, and the input amount of tea-polyphenol is 10 ~ 40%(W/W of total phenol quality in slurry); Pigment requires inclined to one side reddish black, and the input amount of tea-polyphenol is 40 ~ 100%(W/W of total phenol quality in slurry).
6. the method for utilizing polyphenoloxidase and tea-polyphenol to improve melanochrome extraction yield in Fructus Vaccinii Bracteati according to claim 1, is characterized in that, in step (3), the specific activity of added polyphenoloxidase is more than 10000 units/g.
7. polyphenoloxidase and the tea-polyphenol of utilizing according to claim 1 improves the method that melanochrome in Fructus Vaccinii Bracteati extracts yield, and the temperature that it is characterized in that catalytic oxidation is 25 ~ 50 ° of C, and the reaction times is 2 ~ 6 hours.
8. polyphenoloxidase and the tea-polyphenol of utilizing according to claim 1 improves the method that melanochrome in Fructus Vaccinii Bracteati extracts yield, it is characterized in that in step (4), add food grade ethanol, make final alcohol concn reach 30 ~ 60%, the volume ratio of myrtillin leave slurries and ethanol extract is not less than 1:0.8 ~ 1.5, extraction is to carry out under not higher than 60 ° of C temperature condition, and extraction time is 2-6 hour; After extraction finishes, adopt expression separation; It is the ethanol of 30-60% that separated rear residue adds concentration, and the mass volume ratio of the ethanol of residue dry substance quality and second extraction is 1:6 ~ 10, and the time of extracting is for the second time 1-2 hour, and temperature is similarly not higher than 60 ° of C; United extraction liquid, filters, and obtains melanochrome crude product solution.
9. the method for utilizing polyphenoloxidase and tea-polyphenol to improve melanochrome extraction yield in Fructus Vaccinii Bracteati according to claim 1, is characterized in that in step (5), reduced pressure is 0.09MPa, 60 ° of C.
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| CN113383892A (en) * | 2021-07-01 | 2021-09-14 | 苏州求是本草健康科技有限公司 | Preparation method and application of dried leaves of oriental blueberry |
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