KR100979524B1 - Method for extracting and refining flavonoid from Onju Orange - Google Patents

Abstract

The present invention relates to a method for extracting and purifying flavonoid components from citrus fruits, and more particularly, extracting a crude extract from citrus fruits; Injecting the extracted crude extract into a continuous column filled with filler and simultaneously injecting ethanol to continuously circulate to adsorb and desorb flavonoids; Concentrating the extract to which the flavonoids are adsorbed and desorbed, and then recovering them by permeation through a silica column; And extracting and purifying flavonoid components from citrus fruits comprising the step of concentrating and drying the recovered extract.

Extraction and purification method according to the present invention can obtain a high purity and high recovery rate of flavonoids from citrus fruits, to produce a variety of functional products including citrus to contribute to the national health promotion and promote citrus consumption through industrialization.

Citrus, Extract, Flavonoids

Description

Method for Extracting and Purifying Flavonoids from Citrus Fruits {Method for extracting and refining flavonoid from Onju Orange}

The present invention can obtain flavonoids from citrus fruits with high purity and high recovery rate, and manufacture and manufacture of various functional products including citrus fruits, which contributes to the improvement of national health and the extraction and purification of flavonoid components from citrus fruits that can promote citrus consumption through industrialization. It is about a method.

Citrus peel contains various bioactive substances such as flavonoids, carotenoids, kurmarins, and essential oils, and various functional evaluations have been actively conducted in Japan and Korea. In particular, the attention of related industries has been focused on recent studies on representative flavonoid components such as hesperidin and naringin among the bioactive substances in citrus peels.

Since 2000, Citrus aurantium (Citrus aurantium), also known as crust, contains a large amount of hesperidin, mainly in Europe and Japan, and has been used as a safe material for various health foods.

Jeju Wenzhou Citrus Powder is manufactured by drying whole flesh and skin, so hesperidin and other physiologically active substances are still alive and have a higher taste for taste. Use is considered and throws hope on Jeju citrus revival.

Furthermore, in the medical field, as the new flavonoids are extracted, diseases to be treated are diversified, such as liver disease, diabetes, rheumatoid arthritis, various cancers, analgesic, viral diseases, allergic diseases, arteriosclerosis, cardiovascular diseases, stress, It has been found that there is pharmacological activity in depression.

Recently, flavonoids (Naringin, Hesperidin, etc.) contained in fruit or citrus peel in Korea have been found to be effective in treating hypoglycemia, arteriosclerosis and liver disease.

Attempts have been made to extract various ingredients contained in citrus fruits.

Republic of Korea Patent Publication No. 2003-0038009 is a flavonoid extraction step of extracting the flavonoids for 1, 2, 3 times (6 hours) at 90 ~ 100 ℃ with a ratio of citrus fruit and methanol to 1:10; Pulverizing and drying the residue produced in the flavonoid extraction step using a mill to remove residual methanol in a dryer at about 30 to 40 ° C .; A hydrolysis and washing step of adding 10% hydrochloric acid and water per 5 g of the powder produced in the grinding and drying step at 0.47 g and 47.14 g, respectively, reacting for 40 minutes, and then adding washing water to completely remove foreign substances; The washing residue in the washing step was added 1.8 times hot water of 80 ° C or more and 0.02 to 0.03 times of 10% hydrochloric acid to the weight ratio, and then reacted for 30 to 120 minutes at 80 to 100 ° C, followed by filtration to extract lyophilized pectin. A method for extracting and purifying flavonoids and pectin by a continuous process in a citrus gourd characterized in that it comprises a step.

This method is not only very complicated but also uses methanol and hydrochloric acid which are harmful to human body as extraction solvent, so there is a concern that these substances remain in the extract.

Korean Patent Publication No. 2004-0001551 discloses a method of extracting active ingredients including hesperdine extracted from a citrus peel, a method of extracting mineral components from the extracts of skins, seaweeds, and the extracting method. The present invention proposes a blended composition obtained by combining the extracted mineral extract, the hesperdine-containing rind extract and the mineral component extract in a predetermined ratio, and a method of using the extract or the blended composition as a food additive.

In the patent, hesperdine is extracted from citrus fruits, but the method is different from the method proposed in the present invention, and the extraction efficiency is also not very satisfactory.

Republic of Korea Patent Publication No. 2007-0078167 step of preparing a dry powder of citrus; Extracting the citrus dried powder with a polar solvent; And through the step of removing impurities from the extract obtained in the step has been proposed a method for separating narirutin from citrus.

In addition to these methods, a variety of extraction methods have been proposed. However, in order to be applied to medicines or foods, they must be extracted with high purity, and considering the industrial aspect, a high recovery rate still remains.

It is an object of the present invention to provide an extraction and purification method that can recover flavonoids, which are active ingredients from tangerines, with high purity and high recovery.

In order to achieve the above object, the extraction and purification method of the flavonoid component from the tangerine according to the present invention comprises the steps of extracting a crude extract from the tangerine; Injecting the extracted crude extract into a continuous column filled with filler and simultaneously injecting ethanol to continuously circulate to adsorb and desorb flavonoids; Concentrating the extract to which the flavonoids are adsorbed and desorbed, and then recovering them by permeation through a silica column; And concentrating and drying the recovered extract.
In addition, the crude extract after washing the citrus with water, finely chopped and rapidly dried at a temperature of 200 ~ 250 ℃, hot water for 10 to 30 minutes in 80-100 ℃ hot water, characterized in that the obtained by filtration.
In addition, the adsorption and desorption step is characterized in that it is continuously repeated by continuously injecting the extraction solvent 80% ethanol to the column filled with SDVB (Styrene divinylbenzene, styrene divinylbenzene) filler through the circulation pump.
In addition, concentration of the extract from which the flavonoids are adsorbed and desorbed is concentrated 3 to 5 times by using an evaporator and passed through a silica column connected with a 256 nm spectrometer, ending at the beginning of the initial peak. It is characterized in that all by the time point is recovered.
In addition, drying after concentration of the recovered extract is carried out using an evaporator, and further characterized in that the purity is confirmed by using HPLC after drying.

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Extraction and purification method according to the present invention can obtain a high purity and high recovery rate of flavonoids from citrus fruits, to produce a variety of functional products including citrus to contribute to the national health promotion and promote citrus consumption through industrialization.

The application of flavonoids, a functional substance contained in conventional citrus fruits, is mostly small scale or laboratory scale and is not popularized.

The present inventors have established the optimum conditions for the mass production of flavonoids, a functional material of citrus fruits, to allow industrialization.

The present invention is described in more detail below.

1 is a schematic diagram showing an extraction and purification method according to the present invention.

1, the extraction and purification method comprises the steps of extracting a crude extract (crude extract) from citrus fruits (S1); Injecting the extracted crude extract into a continuous column filled with filler and simultaneously injecting ethanol to continuously circulate to adsorb and desorb flavonoids (S2); Concentrating the extract from which the flavonoids are adsorbed and desorbed, and then recovering by permeation through a silica column (S3); And concentrating and drying the recovered extract (S4).

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First, in step S1, the crude extract is extracted from citrus fruits.

Specifically, the crude extract effectively removes essential oil components, such as terpene oil, remaining in citrus fruits, such extracts are finely rinsed after washing the citrus and rapidly dried at a temperature of 200-250 ° C., followed by 80-100 ° C. It is heated by hot water for 10 to 30 minutes and then filtered.

The citrus fruit was a high content of the skin peel as a result of performing a separate experiment of the flavonoid content extracted in terms of utilization, the whole fruit may be used in terms of recovery efficiency.

In this case, the filtration is performed twice using a pore size 212 μm molecular sieve and a paper filter (Watman filter paper No. 2).

Next, in step S2, the crude extract obtained in step S1 is injected into a continuous column filled with filler and ethanol is injected at the same time to continuously circulate.

Specifically, the obtained crude extract is injected into the packed column to adsorb the flavonoid component contained in the crude extract to the filler, and continuously injected with ethanol to desorb the adsorbed flavonoid component from the filler to dissolve in ethanol, and Desorption process takes place.

This adsorption and desorption step is continuously repeated by continuously injecting ethanol, an extractant, continuously into the column filled with filler through a circulation pump.

The filler may be an adsorbent conventionally used in this field, preferably SDVB (Styrene divinylbenzene, styrene divinylbenzene), Amberile Xad, LiChrosorb (Si60, 100/10 ㎛), LiChrolut (RP-18 , EN) and the like, more preferably SDVB.

The ethanol is preferably 60 to 100% ethanol, more preferably 80% ethanol.

Next, in step S3, the extract obtained in step S2 is concentrated and then recovered by permeation through a silica column.

Specifically, the extract (containing flavonoids) obtained in the previous step (S2) was concentrated 3 to 5 times with an evaporator, and then permeated through a silica column (2000 g-150 ml) connected with a 256 nm spectrometer, Recover everything from the beginning of the initial peak to the end.

The transmission is capable of recovering only flavonoid components using a 256 nm spectrometer.

Next, in step S4, the recovered extract is concentrated and dried to obtain flavonoid components from citrus fruits in high purity and high yield.

The concentration may be an evaporator concentrator, and after drying through concentration, check the purity using HPLC.

This extraction and purification method is to have a large amount of functional material, ie flavonoids during extraction, and in particular to minimize the loss rate to meet the economics.

Flavonoid components obtained through the above steps include limonene, narigin, rutin, hesperidin. Such ingredients may be applied to various beverages and may be applied to the pharmaceutical field.

Extraction method according to the invention can be carried out using a variety of extraction apparatus, it is not particularly limited in the present invention.

Figure 2 is a schematic diagram showing the sequence of the extraction and purification apparatus according to the present invention.

Referring to FIG. 2, the apparatus is largely divided into an extraction apparatus 100 and a concentration and identification apparatus 200.

The extraction device 100 has an inlet and an outlet, and is filled with a filler therein, so that the crude extract is injected and desorbed by continuous circulation of the filler and flavonoids when the crude extract is injected to continuously generate a flash module column ( A large silica column 110 with a flash module column.

The crude extract continuous injector 120 is connected to the inlet side of the large silica column 110 and continuously injects crude extract into the column (ie, flash module column).

The outlet side of the large silica column 110 is connected to the collecting unit 130 for collecting the extracted components.

On the other hand, the eluent reservoir 140 for injecting eluent into the large silica column 110 is connected to the inlet side of the large silica column 110, and the flavonoid component in the crude extract is extracted by the eluent. .

The other side of the eluent storage tank 140 is provided with a pressure type forced transfer pump 150 for transferring the eluent from the large silica column 110 therefrom.

One side of the outlet of the large silica column 110 is provided with a solvent conveying line 160 for conveying the unfractionated material to the eluent storage tank 140.

At this time, if necessary to maintain the clarity of the eluent filtration between the eluent reservoir 140 and the large silica column 110 (170), for example, a silica-filled coatec filter (filter, silica cotec) Go through.

In order to separate the large silica column 110, it is continuously passed through a spectrometer (230-290 nm / spectometer), and all the fractions are collected from the time point at which the initial peak starts and ends, and then sent to the collecting unit 120. The solvent is sent back through the solvent return line 160 to the eluent reservoir 140.

The collecting unit 130 collects fractions through a large silica column 110 (flash module column) connected thereto using a collector equipped with a stopcock-valve.

The extract fractionated from the extraction device 100 is transferred to the concentration and confirmation device 200, which is first concentrated in the evaporator 210, and then separated by a spectrometer (230-290 nm / spectometer) connected again for purity inspection. It penetrates and examines the small silica column 220 for liquid confirmation.

Then, the separation complete solution is collected again, and then concentrated in the final concentration tank 230 for final concentration to obtain the final flavonoid component from the tangerine.

Hereinafter, preferred embodiments of the present invention will be described in order to facilitate understanding of the present invention. However, the following examples are merely provided to more easily understand the present invention, and the contents of the present invention are not limited by the examples.

Experimental Example 1

Flavonoids were extracted from citrus fruits using the apparatus shown in FIG. 2. In this case, various kinds of adsorbents and ethanol concentrations were varied, and the obtained results are shown in Tables 1 and 2 below.

The analysis of limonene and narizine was carried out under the following conditions.

Limonene Narizin Instrument: Shimazu GC / MS QP5050A
Column: HP Innowax (0.25mm ID × 0m L)
Oven Temp. : 100 ℃
Interface Temp. 230 ℃
Injection Temp: 230 ℃
Injection volume: 1µl Instrument: Waters MicroMass ZQ
Pump: Waters 600
Column: Versapack C-18 (4.1mm ID × 300mmL)
Add Decteor: Shimazu SPD-10 vp
Det wavelength: 254nm
Flowrate: 1.0ml / min, Injection volume: 20µl

Limonene Recovery and Purity Reclaimer absorbent Amberile Xad LiChrosorb
(Si60, 100 / 10㎛) LiChrolut
(RP-18, EN) SDVB
I SDVB
II SDVB
III Ethanol (60%) tr. tr. tr. tr. tr. tr. 70% tr. tr. tr. tr. tr. tr. 80% 17.61 ^* 0.13 tr. 14.69 0.30 tr. 90% 0.14 0.05 tr. 0.22 0.08 tr. 100% tr. tr. tr. tr. tr. tr.

* Calibration purity by initial MS: 100%, ^* Total value of detection area%

Narizine Recovery and Purity Reclaimer absorbent Amberile Xad LiChrosorb
(Si60, 100 / 10㎛) LiChrolut
(RP-18, EN) SDVB
I SDVB
II SDVB
III Ethanol (60%) tr. tr. tr. tr. tr. tr. 70% tr. tr. tr. tr. tr. tr. 80% 71.31 ^* tr. 91.42 93.22 85.78 70.94 90% 66.78 tr. 22.55 85.07 84.48 80.66 100% tr. tr. tr. tr. tr. tr.
* Calibration purity by initial MS: 100%, ^* Total value of detection area%

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Referring to Table 1 and Table 2, the degree of recovery of limonene was examined using GC / MS. As a result, the recovery rate was higher in the order of Amberile Xad> SDVB I in 80% ethanol, and SDVB in 80% ethanol in the case of narizine. I> LiChrolut (RP-18, EN)> Amberile Xad showed high recovery rate.

Experimental Example 2

The experiment was carried out in the same manner as in Experiment 1, the tangerines harvested in October and November were adsorbed the main flavonoid components using SDVB to determine the recovery rate.

Detection ingredient  Main extract flavonoid content (% during drying) October harvest November harvest Ethanol (80%) Ethanol (90%) Ethanol (80%) Ethanol (90%) Rutin pulp 0.34 0.49 0.58 0.43 pericarp 0.45 0.81 2.38 2.02 Hesperidin
(Hesperidin) pulp 0.69 1.00 0.53 0.77 pericarp 1.51 1.54 2.73 1.52 Narizin
(Naringin) pulp 6.94 18.42 10.70 10.81 pericarp 14.0 18.10 19.67 19.22

Referring to Table 3 above, 80% ethanol was found to be highly recovered in rutin, hesperidin, narizine, etc. in the efficiency review of the concentration of recovery agent after adsorption of main flavonoids using citrus SDVB. On the other hand, there was no difference in treatment between the flesh.

Experimental Example 3

By using the following extraction conditions, the column size and filler were changed to
Lavonoids were extracted and the recovery rates obtained are shown in Tables 4 and 5.

1.Pump condition: Gradient HPFC Pump
Flow Rate: 1-100ml / min (Pressure Limit: 100Psi (7 Bar))
-Solvent Delivery: Constant Volume (3ml) Electric
2. Wavelength condition
UV Detector (dual wavelenght (200-320nm))
3. Extraction condition
-Fraction modes: Extraction by UV Slope, Volume, and Time
4. Large capacity extraction condition
-2.5g, 10g of Silica filled with Compression Module
Extraction continuously (12mm, 25mm, 40mm)

Column aperture 12 mm 25 mm 40 mm 75 mm % Recovery 35 50 75 89

Referring to Table 4, after extracting with SDVB, after confirming the column efficiency by flavonoid recovery by KILO Prep, the higher the diameter of the column, the higher the recovery. This result means that the method according to the invention can be applied well to large scale for industrialization.

absorbent HP-20M (SDVB) Silica KP-C18-HS % Recovery 65 75 70

Referring to Table 5 above, the optimum flavonoid recovery column filler with KILO Prep after SDVB extraction showed the highest recovery of silica as the adsorbent.

Experimental Example 4

Flavonoids were extracted through the following conditions, and flavonoids were extracted from citrus fruits by column passing using a known general batch method for comparison, and the obtained results are shown in Table 6 below.

1.Pump condition: Pressurized Press pump
-Solvent Delivery: Constant Volume 150ml
2. Wavelength condition
-UV 253nm
3. Extraction condition
Fraction modes: Continuous adsorption extraction
4. Large capacity extraction condition
-Solvent Transportation: Compression Module
-Column condition: 2,000g-150ml Silica filling

system Recovery Invention Continuous pressurized 85% Conventional technology General batch column pass 70%

Referring to Table 6, in the case of the method according to the invention it can be seen that the recovery of the flavonoids is more excellent.

The method according to the present invention can produce at least 30% of the added value when producing 100% reduced juice 50,000 tons in the functional juice when processing 100,000 tons of Jeju citrus fruit, can achieve a very high profit.

Furthermore, with citrus
Since the various effects of flavonoids obtained from can be applied to various fields, it can be applied to various fields such as food and medical fields.

1 is a schematic diagram showing an extraction and purification method according to the present invention.

2 is a schematic diagram showing the sequence of the apparatus according to the present invention.

Claims (5)

Extracting a crude extract from citrus fruits (S1); Injecting the extracted crude extract into a continuous column filled with filler and simultaneously injecting ethanol to continuously circulate to adsorb and desorb flavonoids (S2); Concentrating the extract from which the flavonoids are adsorbed and desorbed, and then recovering by permeation through a silica column (S3); And Concentrating and drying the recovered extract (S4); Extraction and purification method of flavonoid components from citrus fruits, characterized in that it comprises a. The method of claim 1, The crude extract is washed with citrus, finely chopped and rapidly dried at a temperature of 200-250 ° C., then boiled in hot water at 80-100 ° C. for 10-30 minutes, followed by filtration to obtain flavonoids from citrus fruits. Extraction and Purification Methods. The method of claim 1, The adsorption and desorption step is carried out by continuously injecting 80% ethanol, an extraction solvent, into the column filled with SDVB (Styrene divinylbenzene, styrene divinylbenzene) filler through the circulation pump from the citrus citrus flavonoids Extraction and purification methods. The method of claim 1, Concentration of the extract from which the flavonoids were adsorbed and desorbed was concentrated 3 to 5 times by using an evaporator, and permeated through a silica column connected with a 256 nm spectrometer, from the beginning of the initial peak to the end. Extracting and purifying flavonoid components from citrus fruits, all of which are recovered. The method of claim 1, Concentration and drying of the recovered extract is carried out using an evaporator, and further drying and extracting and purifying the flavonoid component from the tangerine, characterized in that to confirm the purity using HPLC.

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