CN107245483A - A kind of complex enzyme for being used to extract kudzu root flavone and the method extracted using the complex enzyme - Google Patents
A kind of complex enzyme for being used to extract kudzu root flavone and the method extracted using the complex enzyme Download PDFInfo
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Abstract
A kind of method extracted the invention discloses complex enzyme for being used to extract kudzu root flavone and using the complex enzyme, complex enzyme includes pectase and β glucuroides, wherein, total enzyme concentration of pectase and β glucuroides is 150~400IU, and the enzyme activity of β glucuroides accounts for the 20%~80% of total enzyme concentration；Extracting method comprises the following steps：Weigh the root of kudzu vine to crush and sieve, by liquid ratio 10~40:1 adds the disodium hydrogen phosphate citric acid solution of pH4.0~6.0, adds pectase and β glucuroides, after 0.5~2.5h of reaction under the conditions of 40~60 DEG C, suction filtration, you can.Advantage is that complex enzyme can be applied to the extraction of kudzu root flavone, and can effectively improve recovery rate, safety and environmental protection, pollution-free, simple and convenient extraction, it is easy to operate simultaneously.
Description
Technical field
The invention belongs to complex enzyme field, more particularly to a kind of complex enzyme for being used to extract kudzu root flavone and it is combined using this
The method that enzyme is extracted.
Background technology
The root of kudzu vine is a kind of edible herbaceous plant, rich in flavone compound.There is kudzu root flavone antibacterial, promoting blood circulation, anticancer to prevent
Cancer, at the same have hypotensive reduce myocardial oxygen consumption and promote retina blood circulation, suppress platelet aggregation, anti-arrhythmia cordis,
The multiple pharmacological effects such as anti-oxidant and enhancing immunity of organisms, are the main active components of the root of kudzu vine.Extraction process is to kudzu root flavone
Yield it is most important.At present, kudzu root flavone extracting method mainly has solvent to leach method, reflux extraction etc..Due to traditional
It is to be dissolved into the active ingredient in string tissue using concentration gradient to carry that water, organic solvent, which extract Flavonoid substances method,
Take in solvent.The recovery rate of conventional method is relatively low, and extraction time is long, and the utilization of resources is insufficient.Ultrasonic assistant is extracted and micro-
Although it is short that ripple assisted extraction prepares the Pueraria Flavonid time, because equipment investment is big, operation and maintenance cost are very high, it is difficult to use
In large-scale industrialized production.The development of modern biotechnology promotes exogenous biological enzyme and extracts processing skill in active skull cap components
Application in art, is also the overall development trend of current plant extract processing.
The content of the invention
Goal of the invention：The first object of the present invention, which is to provide one kind, can be applied to extract kudzu root flavone and can effectively carry
The complex enzyme of high extraction；The second object of the present invention is to provide the method extracted using the complex enzyme.
Technical scheme：The present invention is used for the complex enzyme for extracting kudzu root flavone, including pectase and β glucuroides, both
Total enzyme concentration be 150~400IU, wherein, the enzyme activity of β glucuroides accounts for the 20%~80% of total enzyme concentration.It is preferred that, institute
The 30%~50% of total enzyme concentration can be accounted for by stating the enzyme activity of β glucuroides.
Furtherly, the complex enzyme may include cellulase, pectase and β glucuroides, wherein, the cellulase
Enzyme activity is 150~350IU, and pectase enzyme activity is 150~350IU, and β glucuroides enzyme activity is 150~350IU.It is preferred that, it is fine
The plain enzyme enzyme activity of dimension can be 300~350IU, and pectase enzyme activity can be 300~350IU, β glucuroides enzyme activity can for 300~
350IU。
The method that the present invention is extracted using complex enzyme, comprises the following steps：(1) weigh the root of kudzu vine to crush and sieve, press
Liquid ratio 10~40:1 adds disodium hydrogen phosphate-citric acid solution of pH4.0~6.0；
(2) pectase and β glucuroides are added by enzyme activity, after 0.5~2.5h of reaction under the conditions of 40~60 DEG C, taken out
Filter, you can.It is preferred that, the reaction time can be 1~2.5h.
Furtherly, the method that the present invention is extracted using complex enzyme, comprises the following steps：(1) kudzu-vine root powder is weighed
It is broken and sieve, by liquid ratio 10~40:1 adds disodium hydrogen phosphate-citric acid solution of pH3.0~5.5；It is preferred that, liquid
It can be 25~35 to expect ratio:1；PH can be 4.5~5.5.
(2) cellulase, pectase and β glucuroides are added by enzyme activity, react 0.5 under the conditions of 40~60 DEG C~
After 2.5h, suction filtration, you can.It is preferred that, reaction temperature can be 40~50 DEG C.
Beneficial effect：Compared with prior art, remarkable advantage of the invention is：The complex enzyme can be applied to kudzu root flavone
Extraction, and can effectively improve recovery rate simultaneously, it is safety and environmental protection, pollution-free；The simple and convenient extraction, it is easy to operate.
Brief description of the drawings
Fig. 1 is the canonical plotting that kudzu root flavone is determined；
When Fig. 2 is using β glucuroides-pectase complex enzyme, influence figures of the pH to kudzu root flavone recovery rate；
When Fig. 3 is using β glucuroides-pectase complex enzyme, influence figure of the temperature to Pueraria Flavonid recovery rate；
When Fig. 4 is using β glucuroides-pectase complex enzyme, influence of the extraction time to Pueraria Flavonid recovery rate
Figure；
When Fig. 5 is using β glucuroides-pectase complex enzyme, influence figure of the enzyme addition to kudzu root flavone recovery rate；
When Fig. 6 is using β glucuroides-pectase complex enzyme, β glucuroides proportion is extracted to kudzu root flavone
The influence figure of rate；
When Fig. 7 uses β glucuroides-pectase complex enzyme, influence figure of the solid-liquid ratio to kudzu root flavone recovery rate；
When Fig. 8 is using β glucuroide-pectase-cellulase complex enzymes, cellulase addition is to extracting flavonoids
The influence figure of rate；
When Fig. 9 is using β glucuroide-pectase-cellulase complex enzymes, pectase addition is to root of kudzu vine recovery rate
Influence figure；
When Figure 10 is using β glucuroide-pectase-cellulase complex enzymes, β glucuroides are carried to kudzu root flavone
Take the influence figure of rate；
When Figure 11 is using β glucuroide-pectase-cellulase complex enzymes, liquid ratio is to kudzu root flavone recovery rate
Influence figure；
When Figure 12 is using β glucuroide-pectase-cellulase complex enzymes, shadows of the pH to kudzu root flavone recovery rate
Ring figure；
When Figure 13 is using β glucuroide-pectase-cellulase complex enzymes, temperature is to kudzu root flavone recovery rate
Influence figure.
Embodiment
Below in conjunction with the accompanying drawings and embodiment is described in further details to technical scheme.
Embodiment 1
The root of kudzu vine that 3.0g is crushed and crossed 40 mesh sieves is weighed, according to liquid ratio 30:1 adds pH5.0 disodium hydrogen phosphate-lemon
Acid buffering solution, adds β glucuroides (100IU/g)-pectase (20000IU/g) complex enzyme, and complex enzyme addition is
The ratio that 350IU, β glucuroide enzyme activity account for total enzyme activity is 40%, the suction filtration after 45 DEG C of water-bath 1h.Filter cake presses liquid ratio 20:1
Add 70% ethanol, 80 DEG C of refluxing extraction 1h, suction filtration, filtrate, measure Pueraria Flavonid content after mixing twice before and after merging.
Embodiment 2
The root of kudzu vine that 3.0g is crushed and crossed 40 mesh sieves is weighed, β glucuroides (100IU/g) 300IU, pectase is added
(20000IU/g) 300IU, adds cellulase (30000IU/g) 300IU.According to liquid ratio 30:1 addition pH5 citric acid-
Suction filtration after disodium hydrogen phosphate buffer solution, 50 DEG C of water-bath 1h.Filter cake presses liquid ratio 20 during suction filtration:1 adds 70% ethanol, and 80 DEG C are returned
Stream extracts 1h, suction filtration.Filtrate twice before and after merging, measure Pueraria Flavonid content after mixing.
The measure of kudzu root flavone
(1) drafting of standard curve
0.0010g rutins accurately are weighed, are dissolved with 30% ethanol, constant volume in 250mL volumetric flasks is moved completely.Take this rutin mark
Quasi- liquid 0,2.5,5.0,7.5,10.0,12.5mL are respectively put into 6 25mL volumetric flasks, are mended with 30% ethanol to 12.5mL, plus
Enter 0.7mL 5% NaNO2Solution shakes up placement 5min；Add 0.7mL10% Al (NO3)3Solution shakes up, and places 5min；Plus
The NaOH solution for entering 5mL10% is shaken up, and 10min is placed with 30% ethanol constant volume, and absorbance is surveyed at wavelength 510nm.With extinction
Spend for ordinate, rutin concentration is abscissa, draws standard curve, as shown in figure 1, regression equation is y=6.2514x+
0.0024R2=0.9995.
(2) flavones content is determined in extract solution
Precision pipettes 3ml kudzu root flavones extract solution in 25ml volumetric flasks, enters with reference to the coloration method for preparing standard curve
Row colour developing, calculates flavones concentration in extract solution according to calibration curve equation, is scaled the quality of the flavones extracted, calculates Huang
Ketone recovery rate.Recovery rate formula is：
A is absorbance in formula.
Comparative example 1
The root of kudzu vine that 3.0g is crushed and crossed 40 mesh sieves is weighed, by liquid ratio 20:1 addition disodium hydrogen phosphate-lemon acid buffering is molten
Liquid.It is separately added into 150IU six kinds of enzymes of hemicellulase, cellulase, lipase, β glucuroides, protease, pectase
(optimum temperature and optimal pH are shown in Table 1) processing root of kudzu vine 1h extracts kudzu root flavone under respective optimum condition.Filtering, filter cake presses liquid material
Than 20:1, with suction filtration after 70% ethanol, 80 DEG C of refluxing extraction 1h.Filtrate twice before and after merging, determines Pueraria Flavonid content, counts
Calculate recovery rate.
Enzyme preparation optimum condition used in table 1
| Title | Optimum condition | Recovery rate/% |
| β glucuroides | 45℃、pH4.5 | 5.07 |
| Cellulase | 50℃、pH4.8 | 4.80 |
| Pectase | 50℃、pH5.5 | 5.03 |
| Hemicellulase | 50℃、pH5.0 | 3.73 |
| Lipase | 40℃、pH6.0 | 3.52 |
| Neutral proteinase | 45℃、pH4.8 | 4.41 |
Comparative example 2
The root of kudzu vine that 3.0g is crushed and crossed 40 mesh sieves is weighed, by liquid ratio 20:1 adds pH4.5 disodium hydrogen phosphate-citric acid
Suction filtration after cushioning liquid, 50 DEG C of water-bath 1h.Filter cake presses liquid ratio 20:1, with suction filtration after 70% ethanol, 80 DEG C of refluxing extraction 1h.Close
And front and rear filtrate twice, Pueraria Flavonid content is determined after mixing, recovery rate is calculated.
Comparative example 3
The root of kudzu vine that 3.0g is crushed and crossed 40 mesh sieves is weighed, by liquid ratio 20:1, after 70% ethanol, 80 DEG C of refluxing extraction 2h
Suction filtration.Pueraria Flavonid content in filtrate is determined, recovery rate is calculated.
Embodiment 1-2 and comparative example 1-3 recovery rate are measured and understood, it is compound using β glucuroides-pectase
Enzyme is first handled the root of kudzu vine, can effectively improve the recovery rate of kudzu root flavone.Gained kudzu root flavone recovery rate is 815%, with
It is not enzyme treated to be 192 times of two hours recovery rates of alcohol extracting, be 1.60 times of β glucuroides compared to improving 222 times, it is fruit
1.62 times of glue enzyme, are 1.70 times of cellulase extraction effect.
The root of kudzu vine is handled using β glucuroide-pectase-cellulase complex enzymes, its root of kudzu vine recovery rate is
9.44%, 2.57 times are improved compared with not enzyme treated, is 2.23 times of two hours recovery rates of alcohol extracting, is β glucuroides
1.86 times, be 1.88 times of pectase, is 1.97 times of cellulase.
The β glucuroides of embodiment 3-pectase is combined ferment treatment and extracts kudzu root flavone
Influences of the 3-1 pH to kudzu root flavone recovery rate
(1) root of kudzu vine that 3.0g is crushed and crossed 40 mesh sieves is weighed, according to liquid ratio 20:1 be separately added into pH 4.0,4.5,5.0,
5.5 and 6.0 disodium hydrogen phosphate-citric acid solution, complex enzyme addition is 300IU, and β glucuroide proportions are
50%, the suction filtration after 50 DEG C of water-bath 1h.Filter cake presses liquid ratio 20:1 adds 70% ethanol, 80 DEG C of refluxing extraction 1h, suction filtration.Merge
Front and rear filtrate twice.Kudzu root flavone recovery rate is as shown in Fig. 2 with pH increase, the extraction of kudzu root flavone takes the lead in dropping after rise
It is low, the kudzu root flavone recovery rate highest when pH is 5.0.
Influence of the 3-2 temperature to Pueraria Flavonid recovery rate
The root of kudzu vine that 3.0g is crushed and crossed 40 mesh sieves is weighed, according to liquid ratio 20:1 adds pH5.0 disodium hydrogen phosphate-lemon
Acid buffering solution, complex enzyme addition is 300IU, and β glucuroides proportion is 50%, respectively in 40,45,50,55 and
Suction filtration after 60 DEG C of water-bath 1h.Filter cake presses liquid ratio 20:1 adds 70% ethanol, 80 DEG C of refluxing extraction 1h, suction filtration.Two before and after merging
Secondary filtrate.Kudzu root flavone recovery rate is as shown in figure 3, as temperature is raised, extraction takes the lead in reducing after increase, when temperature is 45 DEG C
Recovery rate highest.
Influence of the 3-3 extraction times to Pueraria Flavonid recovery rate
The root of kudzu vine that 3.0g is crushed and crossed 40 mesh sieves is weighed, according to liquid ratio 20:1 adds pH5.0 disodium hydrogen phosphate-lemon
Acid buffering solution, complex enzyme addition is 300IU, and β glucuroides proportion is 50%, respectively in 45 DEG C of water enzyme digestions
0.5th, suction filtration after 1,1.5,2 and 2.5h.Filter cake presses liquid ratio 20:1 adds 70% ethanol, 80 DEG C of refluxing extraction 1h, suction filtration.Close
And front and rear filtrate twice.Kudzu root flavone recovery rate is as shown in figure 4, extraction time 1h is best, and this is due to that enzymolysis time is too short, enzyme
Solution is insufficient, and recovery rate is not high, and enzymolysis time is oversize that the production cycle can be caused to increase, while enzyme activity also can be reduced gradually, therefore
Enzymolysis time is limited in the range of 0.5-2.5h.With the extension of time, kudzu root flavone is reduced after extracting the rise that takes the lead in.When carrying
When taking the time more than 1h, recovery rate tends towards stability.
Influence of the 3-4 enzymes addition to kudzu root flavone recovery rate
The root of kudzu vine that 3.0g is crushed and crossed 40 mesh sieves is weighed, according to liquid ratio 20:1 adds pH5.0 disodium hydrogen phosphate-lemon
Acid buffering solution, complex enzyme addition is respectively 150IU, 200IU, 250IU, 300IU, 350IU and 400IU, β glucuroides
Proportion is 50%, respectively the suction filtration after 1h is extracted in 45 DEG C of water-baths.Filter cake presses liquid ratio 20:1 adds 70% ethanol, and 80 DEG C are returned
Stream extracts 1h, suction filtration.Filtrate twice before and after merging.Kudzu root flavone recovery rate is as shown in figure 5, complex enzyme total addition level is 350IU
Preferably, this be due to enzyme concentration very little, digest it is insufficient, recovery rate is not high.And enzyme concentration then can greatly increase cost very much greatly,
Cause to waste, therefore enzyme concentration is limited in the range of 150-400IU.With the increase of enzyme addition, kudzu root flavone recovery rate by
Edge up height, and when enzyme addition is 350IU, recovery rate is maximum, tends towards stability afterwards.
Influence of the 3-5 β glucuroides proportions to kudzu root flavone recovery rate
The root of kudzu vine that 3.0g is crushed and crossed 40 mesh sieves is weighed, according to liquid ratio 20:1 adds pH5.0 disodium hydrogen phosphate-lemon
Acid buffering solution, complex enzyme addition is 300IU, β glucuroide enzyme activity account for total enzyme activity proportion be respectively 20%,
30%th, 40%, 50%, 60% and 80%, the suction filtration after 1h is extracted in 45 DEG C of water-baths respectively.Filter cake presses liquid ratio 20:1 adds 70%
Ethanol, 80 DEG C of refluxing extraction 1h, suction filtration.Filtrate twice before and after merging.Kudzu root flavone recovery rate is as shown in fig. 6, β Portugals in complex enzyme
When polyglycoside enzyme proportion is 40% preferably, this is due to that pectin is primarily present in iuntercellular, and pectin is to constitute plant
The main component of cell membrane outermost layer secondary wall, so raising of a certain amount of pectase to kudzu root flavone recovery rate has very big side
Help.And a certain amount of β glucuroides can make the precursor of Flavonoid substances degrade, so as to effectively improve carrying for Pueraria Flavonid
Take rate.Therefore β glucuroides institute accounting in complex enzyme is limited in the range of 20-80%.With β glucuroide proportions
Increase, kudzu root flavone extract take the lead in rise after reduce.When β glucuroides proportion is 40%, kudzu root flavone is extracted
Rate highest.
Influence of the 3-6 solid-liquid ratios to kudzu root flavone recovery rate
The root of kudzu vine that 3.0g is crushed and crossed 40 mesh sieves is weighed, according to liquid ratio 10:1、15:1、20:1、25:1、30:1、35:1
With 40:1 adds pH 5.0 disodium hydrogen phosphate-citric acid solution, and complex enzyme addition is 350IU, β glucuroides institute
Accounting example is 40%, respectively the suction filtration after 1h is extracted in 45 DEG C of water-baths.Filter cake presses liquid ratio 20:1 adds 70% ethanol, 80 DEG C of backflows
Extract 1h, suction filtration.Merge before and after twice filtrate, recovery rate as shown in fig. 7, liquid ratio is 30:1 kudzu root flavone recovery rate highest,
This is due to that liquid ratio is too small, and wetting of the enzyme liquid to the raw material root of kudzu vine is insufficient, causes hydrolysis result to decline, and reduces recovery rate；And
Liquid ratio is excessive, and the enzyme activity of unit volume enzymolysis liquid is relatively low, and hydrolysis effect is bad, can also reduce recovery rate, while wave can be caused
Take.Therefore solid-liquid ratio is limited to 10-40:In the range of 1.With the increase of liquid ratio, kudzu root flavone is reduced after extracting the rise that takes the lead in,
When liquid ratio is 30:When 1, recovery rate highest.
Kudzu root flavone is extracted in the effect of the β glucuroide-pectases-cellulase synergistic of embodiment 4
Influence of the 4-1 cellulases addition to extracting flavonoids rate
3.0g kudzu-vine root powders are weighed, β glucuroides 150IU, pectase enzyme 150IU is added, is separately added into cellulase
150IU, 200IU, 250IU, 300IU and 350IU.According to liquid ratio 20:1 adds the citrate-phosphate disodium hydrogens of pH 4.5 buffering
Suction filtration after solution, 50 DEG C of water-bath 1h.Filter residue presses liquid ratio 20:1 adds 70% ethanol, 80 DEG C of refluxing extraction 1h, suction filtration.Before merging
Filtrate twice, determines Pueraria Flavonid content, recovery rate as shown in figure 8, cellulase addition is 300IU extractions after mixing afterwards
Rate highest.
Influence of the 4-2 pectases addition to root of kudzu vine recovery rate
3.0g kudzu-vine root powders are weighed, β glucuroides 150IU, cellulase 300IU is added, is separately added into pectase
150IU, 200IU, 250IU, 300IU and 350IU.According to liquid ratio 20:1 addition pH4.5 citrate-phosphates disodium hydrogen buffering is molten
Suction filtration after liquid, 50 DEG C of water-bath 1h.Filter residue presses liquid ratio 20:1 adds 70% ethanol, 80 DEG C of refluxing extraction 1h, suction filtration.Merge twice
General flavone content, recovery rate are determined after filtrate, mixing as shown in figure 9, pectase addition be 300IU when recovery rate highest.
Influence of the 4-3 β glucuroides to kudzu root flavone recovery rate
3.0g kudzu-vine root powders are weighed, pectase 300IU, cellulase 300IU is added, is separately added into β glucuroides
150IU, 200IU, 250IU, 300IU and 350IU.According to liquid ratio 20:1 adds the citrate-phosphate disodium hydrogens of pH 4.5 buffering
Suction filtration after solution, 50 DEG C of water-bath 1h.Filter residue presses liquid ratio 20:1 adds 70% ethanol, 80 DEG C of refluxing extraction 1h, suction filtration.Merge two
General flavone content is determined after secondary filtrate, mixing, as shown in Figure 10, β glucuroide enzymes addition is extracted recovery rate when being 300IU
Rate highest.
Influence of the 4-4 liquid ratios to kudzu root flavone recovery rate
3.0g kudzu-vine root powders are weighed, pectase 300IU, cellulase 300IU, β glucuroide 300IU is added.By liquid material
Than being respectively 10:1、15:1、20:1、25:1、30:1、35:1 and 40:1 addition pH 4.5 citrate-phosphate disodium hydrogen bufferings are molten
Suction filtration after liquid, 50 DEG C of water-bath 1h.Filter residue presses liquid ratio 20:1 adds 70% ethanol, 80 DEG C of refluxing extraction 1h, suction filtration.Merge twice
General flavone content is determined after filtrate, mixing, recovery rate is as shown in figure 11, when liquid ratio is 30:Recovery rate highest when 1.
Influences of the 4-5 pH to kudzu root flavone recovery rate
3.0g kudzu-vine root powders are weighed, pectase 300IU, cellulase 300IU, β glucuroide 300IU is added.Liquid ratio
30:1 adds suction filtration after pH 3,3.5,4,4.5,5 and 5.5 citrate-phosphate disodium hydrogen cushioning liquid, 50 DEG C of water-bath 1h.Filter residue
By liquid ratio 20:1 adds 70% ethanol, 80 DEG C of refluxing extraction 1h, suction filtration.Merge filtrate twice, general flavone is determined after mixing and is contained
Amount, recovery rate is as shown in figure 12, the recovery rate highest when pH is 5.
Influence of the 4-6 temperature to kudzu root flavone recovery rate
3.0g kudzu-vine root powders are weighed, pectase 300IU, cellulase 300IU, β glucuroide 300IU is added.Liquid ratio
30:1 adds pH5 citrate-phosphate disodium hydrogen cushioning liquid, respectively again 40, suction filtration after 45,50,55 and 60 DEG C of water-bath 1h.Filter residue
By liquid ratio 20:1 adds 70% ethanol, 80 DEG C of refluxing extraction 1h, suction filtration.Merge filtrate twice, general flavone is determined after mixing and is contained
Amount, recovery rate is as shown in figure 13, the recovery rate highest when temperature is 50 DEG C.
Claims (10)
1. a kind of complex enzyme for being used to extract kudzu root flavone, it is characterised in that：The complex enzyme includes pectase and β glucosides
Enzyme, both total enzyme concentrations are 150~400IU, wherein, the enzyme activity of β glucuroides accounts for the 20%~80% of total enzyme concentration.
2. the complex enzyme according to claim 1 for being used to extract kudzu root flavone, it is characterised in that：The complex enzyme includes fiber
Plain enzyme, pectase and β glucuroides, wherein, the cellulose enzyme activity be 150~350IU, pectase enzyme activity be 150~
350IU, β glucuroide enzyme activity are 150~350IU.
3. the complex enzyme according to claim 1 for being used to extract kudzu root flavone, it is characterised in that：The β glucuroides
Enzyme activity account for the 30%~50% of total enzyme concentration.
4. the complex enzyme according to claim 2 for being used to extract kudzu root flavone, it is characterised in that：The cellulose enzyme activity
For 300~350IU, pectase enzyme activity is 300~350IU, and β glucuroides enzyme activity is 300~350IU.
5. the method that the complex enzyme described in a kind of use claim 1 is extracted, it is characterised in that comprise the following steps：(1)
Weigh the root of kudzu vine to crush and sieve, by liquid ratio 10~40:1 adds disodium hydrogen phosphate-citric acid solution of pH4.0~6.0；
(2) pectase and β glucuroides are added by enzyme activity, after 0.5~2.5h of reaction under the conditions of 40~60 DEG C, suction filtration, i.e.,
Can.
6. the method that the complex enzyme described in a kind of use claim 2 is extracted, it is characterised in that comprise the following steps：(1)
Weigh the root of kudzu vine to crush and sieve, by liquid ratio 10~40:1 adds disodium hydrogen phosphate-citric acid solution of pH3.0~5.5；
(2) cellulase, pectase and β glucuroides are added by enzyme activity, 0.5~2.5h is reacted under the conditions of 40~60 DEG C
Afterwards, suction filtration, you can.
7. the method that use complex enzyme according to claim 5 is extracted, it is characterised in that：The reaction time is 1
~2.5h.
8. the method that use complex enzyme according to claim 6 is extracted, it is characterised in that：The liquid ratio be 25~
35:1。
9. the method that use complex enzyme according to claim 6 is extracted, it is characterised in that：The pH be 4.5~
5.5。
10. the method that use complex enzyme according to claim 6 is extracted, it is characterised in that：The reaction temperature is
40~50 DEG C.
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