CN115337672A - Matrix chromatography device and application method thereof in preparation of flavonoid compound - Google Patents
Matrix chromatography device and application method thereof in preparation of flavonoid compound Download PDFInfo
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- B—PERFORMING OPERATIONS; TRANSPORTING
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- B01D—SEPARATION
- B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
- B01D15/08—Selective adsorption, e.g. chromatography
- B01D15/10—Selective adsorption, e.g. chromatography characterised by constructional or operational features
- B01D15/18—Selective adsorption, e.g. chromatography characterised by constructional or operational features relating to flow patterns
- B01D15/1864—Selective adsorption, e.g. chromatography characterised by constructional or operational features relating to flow patterns using two or more columns
- B01D15/1871—Selective adsorption, e.g. chromatography characterised by constructional or operational features relating to flow patterns using two or more columns placed in series
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
- B01D15/08—Selective adsorption, e.g. chromatography
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
- B01D15/08—Selective adsorption, e.g. chromatography
- B01D15/10—Selective adsorption, e.g. chromatography characterised by constructional or operational features
- B01D15/18—Selective adsorption, e.g. chromatography characterised by constructional or operational features relating to flow patterns
- B01D15/1807—Selective adsorption, e.g. chromatography characterised by constructional or operational features relating to flow patterns using counter-currents, e.g. fluidised beds
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
- B01D15/08—Selective adsorption, e.g. chromatography
- B01D15/26—Selective adsorption, e.g. chromatography characterised by the separation mechanism
- B01D15/38—Selective adsorption, e.g. chromatography characterised by the separation mechanism involving specific interaction not covered by one or more of groups B01D15/265 - B01D15/36
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- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Treatment Of Liquids With Adsorbents In General (AREA)
Abstract
The invention discloses a matrix chromatography device and an application method thereof in flavonoid preparation, and the matrix chromatography device comprises a case, a plurality of chromatography columns arranged in a rectangular array and a bracket positioned at the bottom of the case, wherein the case is positioned at the top of the bracket, the chromatography columns are fixedly arranged in the case, and four corners at the bottom of the case are movably connected with four corners at the top of the bracket; the whole case is rectangular, and a plurality of chromatographic columns are arranged in the case in a rectangular array; the main improvement point of the chromatographic column is that the scheme adopts relatively independent inlets to respectively feed resin and feed materials, and simultaneously adopts relatively independent outlets to respectively discharge the resin and the feed materials; according to the invention, by-products of sweet potatoes are used as raw materials, and through the steps of pulsed electric field and ultrasonic synergistic pretreatment, filtration and clarification and ionic resin separation, flavonoid compounds can be simultaneously prepared from the by-products, so that the fine utilization of the sweet potato by-products is realized, and the method has industrial practicability.
Description
Technical Field
The invention relates to the field of extraction of flavonoid compounds, in particular to a matrix type chromatography device and an application method thereof in preparation of the flavonoid compounds.
Background
Sweet potato is a kind of natural nourishing food with rich nutrients, and contains protein, fat, polysaccharide, P, ca, K, carotene, vitamin A, vitamin C, vitamin E, vitamin B1, vitamin B2 and 8 kinds of amino acids. According to the analysis of scientists, the protein content of the rice is more than 7 times that of the rice; the content of carotene is 3.5 times that of carrot; the content of the vitamin A is 100 times of that of the potato; the contents of sugar, calcium, vitamin B1 and vitamin B2 are all higher than those of rice and flour. Each 100g of fresh potato pieces contains 29.5g of carbohydrate, 0.2g of fat, 20mg of phosphorus, 18mg of calcium and 0.4g of iron. These substances play important roles in promoting the activity of human brain cells and secretory hormones, enhancing the disease resistance of human bodies, improving the immune function, and delaying the decline of intelligence and the aging of organisms.
According to the traditional Chinese, after tubers of sweet potatoes are harvested every year, a large amount of leaves and stems are left, except for being used as food or livestock feed, most of the leaves and stems are discarded beside a field, which is a great waste, and in fact, china always explores a method for changing the sweet potato leaves into valuables, such as sweet potato leaf tea, namely a tea making method is adopted to refine the sweet potato leaves, but the large-scale popularization and application are substantially difficult due to the taste problem; the flavone content of the sweet potato leaves is 1.551 per thousand, the flavone content of the sweet potato stems is 0.860 per thousand, and the flavone content of the sweet potato stems is 0.315 per thousand, so that the market for extracting the flavone from the leaves, stems and stems of the sweet potatoes is quite promising.
The extraction of the flavonoid compounds generally uses a chromatographic column, and the operation of column packing and subsequent elution of the existing chromatographic column is very troublesome.
Disclosure of Invention
The invention aims to provide a matrix type chromatography device and an application method thereof in preparation of flavonoid compounds, so as to solve the problems in the background technology.
In order to achieve the purpose, the invention provides the following technical scheme:
a matrix type chromatography device comprises a case, a plurality of chromatography columns arranged in a rectangular array and a bracket positioned at the bottom of the case,
the case is positioned at the top of the bracket, the chromatographic column is fixedly arranged in the case,
the four corners of the bottom of the case are movably connected with the four corners of the top of the bracket, and the bottom of the rack is also fixedly provided with a vibration motor;
the whole case is rectangular, and a plurality of chromatographic columns are arranged in the case in a rectangular array;
the main improvement point of the chromatographic column is that the scheme adopts relatively independent inlets to respectively feed resin and feed materials, and simultaneously adopts relatively independent outlets to respectively discharge the resin and the feed materials;
the chromatography column is integrally cylindrical, the upper end and the lower end of the chromatography column are provided with openings, the upper end and the lower end of the chromatography column are respectively provided with a column top cover and a column bottom component,
the column top cover is fixedly arranged at the top of the chromatographic column, the top of the column top cover is communicated with a feeding branch pipe, and the side part of the column top cover is communicated with a glue inlet branch pipe;
it is worth noting that the feeding branch pipe is connected with an even distributor inside the chromatographic column, the even distributor is composed of a plurality of coaxially arranged conical cover bodies, the conical cover bodies are fixedly connected, and the even distributor is used for uniformly spraying eluent on the top of the resin column and placing the eluent to influence the resin separation effect;
the column bottom component is fixedly connected to the bottom of the chromatographic column; the bottom side part of the chromatographic column is provided with a plurality of through holes, and the through holes are used for replacing quartz sand (in the figure, the diameter effect of the through holes is amplified for the purpose of showing clarity, and the aperture of the through holes is basically similar to or slightly smaller than the diameter of the quartz sand in practice);
a gap is arranged between the column bottom component and the side wall of the bottom of the chromatographic column, and the gap is used for allowing eluent containing flavone to pass through; and the lateral part intercommunication of column bottom subassembly is equipped with ejection of compact branch pipe, and the discharging pipe sets up with this space intercommunication, and the bottom of column bottom subassembly still communicates to be equipped with out gluey branch pipe simultaneously.
Furthermore, the whole support is rectangular, the support comprises stand columns and transverse connecting rods, the stand columns are at least four, the transverse connecting rods are connected between the adjacent stand columns, and reinforcing rib plates are connected between the adjacent transverse connecting rods so as to increase the overall structural strength of the support.
Furthermore, the case is fixedly connected with the top of the support through a spring damper.
Further, the feeding primary pipe, the glue feeding primary pipe, the discharging primary pipe and the glue discharging primary pipe; the feeding primary pipe is communicated with a plurality of feeding secondary pipes, and a plurality of feeding branch pipes are communicated with the feeding secondary pipes; the glue inlet primary pipe is communicated with a plurality of glue inlet diodes, and the glue inlet diodes are communicated with a plurality of glue inlet branch pipes; the discharging primary pipe is communicated with a plurality of discharging secondary pipes, and a plurality of discharging branch pipes are communicated with the discharging secondary pipes; the glue outlet primary pipe is communicated with a plurality of glue outlet diodes, and the glue outlet diodes are communicated with a plurality of glue outlet branch pipes.
The application method of the matrix type chromatography device in the preparation of the flavonoid compound specifically comprises the following steps:
s1, deactivating enzyme of leaves, stems and stalks of sweet potatoes, drying until the water content is 6%, and crushing to obtain material powder with the particle size of 50 meshes for later use;
s2, placing the material powder in water, wherein the using amount ratio of the material powder to the water is 1g; soaking at 50 deg.C for 40-60min to obtain mixture, treating the mixture with pulsed electric field, and performing ultrasonic treatment to obtain mixture stock solution;
s3, purifying the stock solution:
the treated mixture stock solution is subjected to filter pressing through a squeezing or screw press to obtain filtrate, the amount of the obtained filtrate can be increased by obtaining the filtrate through the squeezing or screw press method, and excessive fine particles are prevented from remaining in the filtrate;
collecting filtrate, centrifuging the filtrate by using a disc centrifuge, and collecting supernatant after centrifugation;
s4, carrying out column chromatography on the supernatant:
s4.1, soaking HPD722 macroporous resin in ethanol for 12-36h, washing with distilled water until no alcohol smell exists, soaking with HCl with the mass concentration of 3% -7% for 2-6h, washing with distilled water until the solution is neutral, soaking with NaOH with the mass concentration of 3% -7% for 2-6h, and washing with distilled water until the solution is neutral to obtain activated HPD722 macroporous resin;
s4.2, uniformly mixing the activated HPD722 macroporous resin with water, and filling the mixture into a column to obtain an HPD722 macroporous resin column for later use;
s4.3, adding the supernatant obtained in the step S3 into the HPD722 macroporous resin column at the flow rate of 2.0 mL/min; the ratio of the amount of the flavone powder in the material to the filling amount of the HPD722 resin in the chromatographic column is 1;
putting the target eluent into a vacuum rotary evaporator, and completely volatilizing an alcohol solution in the target eluent at the temperature of 30 ℃ at the speed of 20r/s to obtain a concentrated extracting solution;
s5, freeze-drying at low temperature; drying the concentrated extractive solution at-90 deg.C for 60 hr to obtain high purity flavone powder.
Specifically, the electric field intensity of the pulse is 0.5-2.0kv/cm, and the frequency is 10-30.
Specifically, the ultrasonic treatment frequency is 20-40kHz, the power is 300-800W, and the time is 40-60min.
Specifically, the speed of the centrifugal treatment is 4000r/min, and the centrifugal time is 15min.
Specifically, in the step 4.3, distilled water is firstly used for eluting until the eluent is colorless, then 50% ethanol is used for eluting with the elution rate of 1mL/min, the absorbance of the eluent is measured by an ultraviolet spectrophotometer, and the target eluent is collected when the eluent reaches 510nm until the eluent is colorless and is stopped collecting, so that the target eluent is obtained.
Compared with the prior art, the invention has the beneficial effects that: the invention improves the optimum loading amount of the dynamic adsorption of the macroporous resin, has uniform column loading and good elution and purification effects, ensures the accurate separation and enrichment of the total flavone, and improves the content of the flavone in the extract; the preparation method can realize semi-automatic operation, save chromatography time, reduce production cost and is more suitable for mass production;
the method takes the byproducts of the sweet potatoes as raw materials, and can simultaneously prepare flavonoid compounds from the byproducts through the steps of pulsed electric field and ultrasonic synergistic pretreatment, filtration and clarification and ion resin separation, thereby realizing the refined utilization of the byproducts of the sweet potatoes and having industrial practicability.
Drawings
FIG. 1 is a schematic view of a matrix chromatography device.
FIG. 2 is a schematic side view of a matrix chromatography device.
FIG. 3 is a schematic diagram of a chromatography column in a matrix chromatography apparatus.
FIG. 4 is a schematic diagram of a part of the structure of a chromatography column in a matrix chromatography device.
FIG. 5 is a schematic diagram of a position A of a matrix chromatography device.
FIG. 6 is a bottom view of a homogenizer for a matrix chromatography apparatus.
Detailed Description
In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention. It is to be understood that the embodiments described are only a few embodiments of the present invention, and not all embodiments. The components of embodiments of the present invention generally described and illustrated in the figures herein may be arranged and designed in a wide variety of different configurations. Thus, the following detailed description of the embodiments of the present invention, presented in the figures, is not intended to limit the scope of the invention, as claimed, but is merely representative of selected embodiments of the invention. All other embodiments, which can be obtained by a person skilled in the art without inventive step based on the embodiments of the present invention, are within the scope of protection of the present invention.
Referring to fig. 1-6, a matrix chromatography device comprises a casing 1, a plurality of chromatography columns 2 arranged in a rectangular array, and a bracket 3 located at the bottom of the casing 1,
the case 1 is positioned at the top of the bracket 3, the chromatographic column 2 is fixedly arranged in the case 1,
the support 3 is rectangular integrally, the support 3 comprises upright posts 301 and transverse connecting rods 302, at least four upright posts 301 are arranged, the transverse connecting rods 302 are connected between the adjacent upright posts 301, and reinforcing rib plates 302 are connected between the adjacent transverse connecting rods 302 to increase the integral structural strength of the support 3;
four corners of the bottom of the case 1 are movably connected with four corners of the top of the bracket 3, specifically, the case 1 is fixedly connected with the top of the bracket 3 through a spring damper 303, and a vibration motor 304 is also fixedly mounted at the bottom of the rack 1;
the whole case 1 is rectangular, and a plurality of chromatographic columns 2 are arranged in the case 1 in a rectangular array;
the main improvement point of the chromatographic column 2 is that the scheme adopts relatively independent inlets to respectively feed resin and materials, and simultaneously adopts relatively independent outlets to respectively discharge resin and materials;
the chromatography column 2 is cylindrical as a whole, the upper end and the lower end of the chromatography column are opened, the upper end and the lower end of the chromatography column 2 are respectively provided with a column top cover 210 and a column bottom component 220,
the column top cover 210 is fixedly arranged at the top of the chromatographic column 2, the top of the column top cover 210 is communicated with a feeding branch pipe 211, and the side part of the column top cover 210 is communicated with a glue inlet branch pipe 212;
it should be noted that the feeding branch pipe 211 is connected with an equilizer 230 inside the chromatography column 2, the equilizer 230 is composed of a plurality of coaxially arranged conical cover 231, the conical cover 231 is fixedly connected, the equilizer 230 is used for uniformly spraying eluent on the top of the resin column, and the resin separation effect is affected by the placement of the eluent;
the column bottom component 220 is fixedly connected to the bottom of the chromatographic column 2; the bottom side of the chromatographic column 2 is provided with a plurality of through holes, and the through holes are used for replacing quartz sand (in the figure, the diameter effect of the through holes is enlarged for the sake of clarity, and the aperture of the through holes is substantially similar to or slightly smaller than the diameter of the quartz sand in practice)
A gap is arranged between the column bottom component 220 and the side wall of the bottom of the chromatographic column 2, and the gap is used for passing through eluent containing flavone; the side part of the column bottom assembly 220 is communicated with a discharge branch pipe 221, the discharge pipe 221 is communicated with the gap, and the bottom of the column bottom assembly 220 is also communicated with a glue discharge branch pipe 222;
a feeding primary pipe 241, a glue feeding primary pipe 243, a discharging primary pipe 245 and a glue discharging primary pipe 247; the feeding primary pipe 241 is communicated with a plurality of feeding secondary pipes 242, and a plurality of feeding branch pipes 211 are communicated with the feeding secondary pipes 242; the glue inlet primary pipe 243 is communicated with a plurality of glue inlet diodes 244, and the glue inlet diodes 244 are communicated with a plurality of glue inlet branch pipes 212; the discharging primary pipe 245 is communicated with a plurality of discharging secondary pipes 246, and a plurality of discharging branch pipes 221 are communicated with the discharging secondary pipes 246; the glue outlet primary tube 247 is communicated with a plurality of glue outlet diodes 248, and the glue outlet diodes 246 are communicated with a plurality of glue outlet branch tubes 222.
In the scheme, the chromatography column is filled by a wet method, namely, uniform slurry formed by resin and a solvent is continuously added into the chromatography column, an eluent is used for 'column walking' under pressure, the eluent flows out from the lower end, simultaneously, the resin is uniformly settled in the chromatography column until the whole chromatography column is filled, and only the uniformly settled resin can ensure good separation effect;
the scheme assists resin sedimentation in all chromatographic columns through the vibration motor 304,
because the soft gel column is deformed and blocked by too high extrusion and is generally not more than 1 meter, the height of the chromatographic column 2 in the scheme is 80cm,
the invention improves the optimal sample loading amount of the dynamic adsorption of the macroporous resin, has uniform column loading and good elution and purification effects, ensures the accurate separation and enrichment of the total flavonoids and improves the content of the flavonoids in the extract. The preparation method can realize semi-automatic operation, save chromatography time, reduce production cost and is more suitable for mass production.
The application method of the matrix type chromatography device in the preparation of the flavonoid compound specifically comprises the following steps:
s1, deactivating enzymes of leaves, stems and stalks of sweet potatoes, drying until the water content is 6%, and crushing to obtain material powder with the particle size of 50 meshes for later use.
S2, placing the material powder in water, wherein the using amount ratio of the material powder to the water is 1g; soaking at 50 deg.C for 40-60min to obtain mixture, treating the mixture with pulsed electric field, and performing ultrasonic treatment to obtain mixture stock solution;
specifically, the intensity of the pulse electric field is 0.5-2.0kv/cm, and the frequency is 10-30;
specifically, the ultrasonic treatment frequency is 20-40kHz, the power is 300-800W, and the time is 40-60min;
the pulse electric field increases the permeability of the cell membrane, aggravates the oscillation, weakens the strength of the membrane, so the membrane is damaged, substances in the membrane easily flow out, substances outside the membrane easily permeate, and the protective effect of the cell membrane is weakened or even disappears; have good characteristics that are generally non-thermal processes; the temperature is not obviously increased in the process of pulse electric field treatment; the flavonoid substances can not be reduced;
the ultrasonic wave instantly finishes the rupture of plant cell walls and the whole organism, causes the temperature of the system to rise, and simultaneously enhances the release, diffusion and dissolution of intracellular substances by the vibration effect generated by the ultrasonic wave, thereby obviously improving the extraction efficiency; the two components have synergistic effect, and can reduce the precipitation of tannin components in sweet potato leaves, which have strong viscosity and make the extractive solution extremely difficult to filter.
S3, purifying the stock solution:
the treated mixture stock solution is subjected to filter pressing through a squeezing or screw press to obtain filtrate, the amount of the obtained filtrate can be increased by obtaining the filtrate through the squeezing or screw press method, and excessive fine particles are prevented from remaining in the filtrate;
collecting filtrate, centrifuging by a disc centrifuge, and collecting supernatant after centrifugation, wherein the speed of centrifugation is 4000r/min, and the centrifugation time is 15min;
s4, carrying out column chromatography on the supernatant:
first, regarding column chromatography separation, negative pressure, normal pressure and pressurization are basically performed, respectively;
the negative pressure column can reduce the consumption of the silica gel resin by 50% or more according to experience, a large amount of air can pass through the silica gel under the condition of negative pressure so as to volatilize the solvent, and easily decomposed substances like flavonoid compounds can be reduced along with the volatilization of the solvent, so that a means of using the negative pressure column chromatography is eliminated at first;
the pressure column can make the eluent faster by external pressure source, and the pressure increase can make compressed air and small air pump, the pressure source is simpler, and it is suitable for separating flavone;
the efficiency at atmospheric pressure is best, but the separation time can be very long, and a column can be several months;
therefore, the scheme selects a pressurizing column;
s4.1, soaking HPD722 macroporous resin in ethanol for 12-36h, washing with distilled water until no alcohol smell exists, soaking with HCl with the mass concentration of 3% -7% for 2-6h, washing with distilled water until the solution is neutral, soaking with NaOH with the mass concentration of 3% -7% for 2-6h, and washing with distilled water until the solution is neutral to obtain activated HPD722 macroporous resin;
s4.2, uniformly mixing the activated HPD722 macroporous resin with water, and filling the mixture into a column to obtain an HPD722 macroporous resin column for later use;
s4.3, adding the supernatant obtained in the step S3 into the HPD722 macroporous resin column at the flow rate of 2.0 mL/min; the ratio of the amount of the flavone powder in the material to the filling amount of the HPD722 resin in the chromatographic column is 1;
eluting with distilled water until the eluate is colorless, eluting with 50% ethanol at an elution rate of 1mL/min, measuring absorbance of the eluate with an ultraviolet spectrophotometer, collecting the target eluate when the eluate reaches 510nm, stopping collecting until the eluate is colorless, and obtaining the target eluate;
putting the target eluent into a vacuum rotary evaporator, and completely volatilizing the alcohol solution in the target eluent at the temperature of 30 ℃ at the speed of 20r/s to obtain a concentrated extracting solution.
S5, freeze-drying at low temperature; drying the concentrated extractive solution at-90 deg.C for 60 hr to obtain high purity flavone powder
The application method of the matrix type chromatography device in the preparation of the flavonoid compound specifically comprises the following steps:
s1, removing green of leaves, stems and stalks of sweet potatoes, drying until the water content is 6%, and crushing to obtain material powder with the particle size of 50 meshes for later use.
S2, placing 1000kg of material powder in water, wherein the using amount ratio of the material powder to the water is 1g; soaking at 50 deg.C for 40-60min to obtain mixture, treating the mixture with pulsed electric field, and performing ultrasonic treatment to obtain mixture stock solution; the intensity of the pulse electric field is 0.5-2.0kv/cm, and the frequency is 10-30; the ultrasonic treatment frequency is 20-40kHz, the power is 300-800W, and the time is 40-60min;
s3, purifying the stock solution:
the treated mixture stock solution is subjected to filter pressing through a squeezing or screw press to obtain filtrate, the amount of the obtained filtrate can be increased by obtaining the filtrate through the squeezing or screw press method, and excessive fine particles are prevented from remaining in the filtrate;
collecting filtrate, centrifuging by a disc centrifuge, and collecting supernatant after centrifugation, wherein the speed of centrifugation is 4000r/min, and the centrifugation time is 15min;
s4, carrying out column chromatography on the supernatant:
s4.1, carrying out activation treatment on HPD722 macroporous resin;
s4.2, uniformly mixing the activated HPD722 macroporous resin with water, and filling the mixture into a column to obtain an HPD722 macroporous resin column for later use;
s4.3, adding the supernatant obtained in the step S3 into the HPD722 macroporous resin column at the flow rate of 2.0 mL/min; the ratio of the amount of the flavone powder in the material to the filling amount of the HPD722 resin in the chromatographic column is 1;
eluting with distilled water until the eluate is colorless, eluting with 50% ethanol at an elution rate of 1mL/min for 36h, measuring absorbance of the eluate with an ultraviolet spectrophotometer, collecting the target eluate when the eluate reaches 510nm, stopping collecting the target eluate when the eluate is colorless, and obtaining the target eluate;
putting the target eluent into a vacuum rotary evaporator, and completely volatilizing the alcohol solution in the target eluent at the temperature of 30 ℃ at the speed of 20r/s to obtain a concentrated extracting solution.
S5, freeze-drying at low temperature; drying the concentrated extractive solution at-90 deg.C for 60 hr to obtain high purity flavone powder of about 963g, which is lower than theoretical measurement value, but has yield meeting expected standard considering loss in production.
The invention provides a method for extracting flavone from sweet potato byproducts, which takes the sweet potato byproducts containing the flavone as raw materials, and firstly increases the permeability of cell membranes, aggravates oscillation, weakens the membrane strength, easily flows out substances in the membranes, easily permeates substances outside the membranes, weakens the protective action of the cell membranes and even disappears by a pulse electric field.
Then the cell wall of the leaf is broken by ultrasound, and the temperature of the system is raised, so that the release, diffusion and dissolution of the intracellular substances are enhanced. The two have synergistic effect, and greatly promote the high-efficiency dissolution of flavone. Because the cell walls of the materials are not broken, the extracting solution is not sticky, the contained viscous substances are less, clear and transparent clear liquid can be obtained after the solid-liquid separation of disc centrifugation, and the matrix type chromatography device of the scheme flows out the flavone along with the eluent, so that the further separation of the flavone is realized.
The invention has the beneficial effects that: the method takes the by-products of the sweet potatoes as raw materials, and can simultaneously prepare flavonoid compounds from the by-products through the steps of pulsed electric field and ultrasonic synergistic pretreatment, filtration and clarification and ion resin separation, thereby realizing the refined utilization of the by-products of the sweet potatoes and having industrial practicability.
In the description of the present invention, it should be noted that the terms "upper", "lower", "inner", "outer", "left", "right", and the like indicate orientations or positional relationships based on orientations or positional relationships shown in the drawings, or orientations or positional relationships that the product of the present invention is conventionally placed in use, or orientations or positional relationships that are conventionally understood by those skilled in the art, which are merely for convenience of description and simplification of the description, and do not indicate or imply that the device or element referred to must have a specific orientation, be constructed in a specific orientation, and operate, and thus, should not be construed as limiting the present invention. Furthermore, the terms "first," "second," and the like are used merely to distinguish one description from another, and are not to be construed as indicating or implying relative importance. In the description of the present invention, it should be further noted that, unless otherwise explicitly stated or limited, the terms "disposed" and "connected" and the like are to be broadly construed, for example, "connected" may be a fixed connection, a detachable connection, or an integral connection; can be mechanically or electrically connected; the connection may be direct or indirect via an intermediate medium, and may be a communication between the two elements. The specific meanings of the above terms in the present invention can be understood according to specific situations by those of ordinary skill in the art.
Claims (9)
1. A matrix type chromatography device comprises a case (1), a plurality of chromatography columns (2) arranged in a rectangular array and a bracket (3) positioned at the bottom of the case (1),
the case (1) is positioned at the top of the bracket (3), the chromatographic column (2) is fixedly arranged in the case (1),
four corners of the bottom of the case (1) are movably connected with four corners of the top of the bracket (3), and a vibration motor (304) is fixedly mounted at the bottom of the rack (1);
the whole case (1) is rectangular, and a plurality of chromatographic columns (2) are arranged in the case (1) in a rectangular array;
the chromatographic column (2) adopts relatively independent inlets to respectively feed resin and feed materials, and simultaneously adopts relatively independent outlets to respectively discharge the resin and the feed materials;
the chromatography column (2) is integrally cylindrical, the upper end and the lower end of the chromatography column (2) are provided with openings, the upper end and the lower end of the chromatography column (2) are respectively provided with a column top cover (210) and a column bottom component (220),
the column top cover (210) is fixedly arranged at the top of the chromatographic column (2), the top of the column top cover (210) is communicated with a feeding branch pipe (211), and the side part of the column top cover (210) is communicated with a glue inlet branch pipe (212);
the feeding branch pipe (211) is connected with an even distributor (230) in the chromatographic column (2), the even distributor (230) consists of a plurality of coaxially arranged conical cover bodies (231), the conical cover bodies (231) are fixedly connected, and the even distributor (230) is used for uniformly spraying eluent on the top of the resin column and placing the eluent to influence the separation effect of the resin;
the column bottom component (220) is fixedly connected to the bottom of the chromatographic column (2); the side part of the bottom of the chromatographic column (2) is provided with a plurality of through holes, and the through holes are used for replacing quartz sand;
a gap is arranged between the column bottom component (220) and the side wall of the bottom of the chromatographic column (2), and the gap is used for allowing eluent containing flavone to pass through; and the lateral part intercommunication of column bottom subassembly (220) is equipped with ejection of compact branch pipe (221), and ejection of compact pipe (221) and this space intercommunication set up, and the bottom of column bottom subassembly (220) still communicates simultaneously and is equipped with out gluey branch pipe (222).
2. Matrix chromatography device according to claim 1, wherein the bracket (3) is rectangular in whole, the bracket (3) comprises at least four columns (301) and transverse connecting rods (302), the transverse connecting rods (302) are connected between adjacent columns (301), and reinforcing ribs (302) are connected between adjacent transverse connecting rods (302) to increase the whole structural strength of the bracket (3).
3. Matrix chromatography device according to claim 1, wherein the chassis (1) is fixedly connected to the top of the frame (3) by means of spring dampers (303).
4. Matrix chromatography device according to claim 1, wherein said feed (241), gel (243), exit (245) and exit (247) primary tubes; the feeding primary pipe (241) is communicated with a plurality of feeding secondary pipes (242), and a plurality of feeding branch pipes (211) are communicated with the feeding secondary pipes (242); the glue inlet primary pipe (243) is communicated with a plurality of glue inlet diodes (244), and a plurality of glue inlet branch pipes (212) are communicated with the glue inlet diodes (244); the discharging primary pipe (245) is communicated with a plurality of discharging secondary pipes (246), and a plurality of discharging branch pipes (221) are communicated with the discharging secondary pipes (246); the glue outlet primary tube (247) is communicated with a plurality of glue outlet diodes (248), and the glue outlet diodes (246) are communicated with a plurality of glue outlet branch tubes (222).
5. The application method of the matrix type chromatography device in the preparation of the flavonoid compound specifically comprises the following steps:
s1, deactivating enzyme of leaves, stems and stalks of sweet potatoes, drying until the water content is 6%, and crushing to obtain material powder with the particle size of 50 meshes for later use;
s2, placing the material powder in water, wherein the using amount ratio of the material powder to the water is 1g; soaking at 50 deg.C for 40-60min to obtain mixture, treating the mixture with pulsed electric field, and performing ultrasonic treatment to obtain mixture stock solution;
s3, purifying the stock solution:
the treated mixture stock solution is subjected to filter pressing through a squeezing or screw press to obtain a filtrate, the amount of the obtained filtrate can be increased by the method of squeezing or screw press, and excessive fine particles are prevented from remaining in the filtrate;
collecting filtrate, centrifuging by a disc centrifuge, and collecting supernatant after centrifugation;
s4, carrying out column chromatography on the supernatant:
s4.1, soaking HPD722 macroporous resin in ethanol for 12-36h, washing with distilled water until no alcohol smell exists, soaking with HCl with the mass concentration of 3% -7% for 2-6h, washing with distilled water until the solution is neutral, soaking with NaOH with the mass concentration of 3% -7% for 2-6h, and washing with distilled water until the solution is neutral to obtain activated HPD722 macroporous resin;
s4.2, uniformly mixing the activated HPD722 macroporous resin with water, and filling the mixture into a column to obtain an HPD722 macroporous resin column for later use;
s4.3, adding the supernatant obtained in the step S3 into the HPD722 macroporous resin column at the flow rate of 2.0 mL/min; the ratio of the amount of the flavone powder in the material to the filling amount of the HPD722 resin in the chromatographic column is 1;
putting the target eluent into a vacuum rotary evaporator, and completely volatilizing an alcohol solution in the target eluent at the temperature of 30 ℃ at the speed of 20r/s to obtain a concentrated extracting solution;
s5, freeze-drying at low temperature; drying the concentrated extractive solution at-90 deg.C for 60 hr to obtain high purity flavone powder.
6. The use of a matrix chromatography device in the preparation of flavonoids according to claim 1, wherein the intensity of the pulsed electric field is 0.5-2.0kv/cm for a number of times of 10-30.
7. The application method of the matrix type chromatography device in the preparation of flavonoids compounds according to claim 1, wherein the ultrasonic treatment frequency is 20-40kHz, the power is 300-800W, and the time is 40-60min.
8. The use of a matrix chromatography device in the preparation of flavonoids according to claim 1, wherein the centrifugation rate is 4000r/min and the centrifugation time is 15min.
9. The application method of the matrix type chromatography device in flavonoid compound preparation according to claim 1, wherein in step 4.3, the elution is carried out by using distilled water until the eluate is colorless, then the elution is carried out by using 50% ethanol at an elution rate of 1mL/min, the absorbance of the eluate is measured by using an ultraviolet spectrophotometer, and the target eluate is collected when the eluate reaches 510nm and is colorless, and then the collection is stopped, so as to obtain the target eluate.
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