KR101233114B1 - Composition for the protection and treatment of Macular degeneration comprising Green tea Acetone extract - Google Patents

Abstract

The present invention relates to a composition for preventing and treating macular degeneration containing green tea acetone extract, and more specifically, it is confirmed that lutein is known to have an inhibitory effect on macular degeneration in green tea acetone extract and is present in an active state in the body. It is intended to be applied for the prevention and treatment of macular degeneration.
According to the present invention, green tea acetone extract can be provided as a source of new lutein applicable for the prevention and treatment of macular degeneration.

Description

Composition for the protection and treatment of Macular degeneration comprising Green tea Acetone extract}

The present invention relates to a composition for the prevention and treatment of macular degeneration containing green tea acetone extract containing lutein as an active ingredient.

The macula lutea is the part that receives the most clear and accurate light due to the densely packed cells in the retina of the eye, and the disease that causes degeneration of the macular degeneration due to various causes is called macular degeneration. do.

The macular degeneration is one of the three major causes of blindness together with glaucoma and diabetic retinopathy. Age is the leading cause of macular degeneration, and family history, race, and smoking are also known as causes. Macular degeneration is the most common cause of vision loss among people over 60 years of age, which can affect central vision and can interfere with a variety of activities that require reading, driving, or fine detailed vision. In addition, damage to the macula causes the eye to lose the ability to recognize details such as small prints, facial features, or small objects.

There are two types of macular degeneration, non-exudative (dry) macular degeneration and exudative (wet) macular degeneration, 90% of people with macular degeneration have a dry form of symptoms. In dry macular degeneration, waste products form a yellow precipitate called drusen and can accumulate in the tissue under the macula. The presence of drusen interferes with blood flow to the retina, especially the macula and, when blood flow is reduced, reduces nutrient supply to the macula, stopping or contracting the efficient action of photosensitive cells. In wet macular degeneration, new, weak blood vessels can grow in or below the retina, causing fluid and blood to leak into the space below the macula. Wet macular degeneration is sometimes described as choroidal neovascularization, which is the region of the subretinal vessels and angiogenesis refers to the growth of new blood vessels in the tissue, as can be inferred from the term choroidal neovascularization. Blood vessels are born from the macula and grow.

The most common early signs of dry macular degeneration are blurred vision. In the case of dry macular degeneration, fewer cells are functional in the macula, which reduces the sharpness when looking at the details of the front face, such as a face or a book letter. As the loss of the photosensitive cells expands, a small (but larger) blind spot appears in the middle of the field of view. A typical early sign of wet macular degeneration is a straight line bend. This occurs when fluid collects from the lacrimal vessels and lifts the macula, distorting the field of view. Small blind spots may also appear in wet macular degeneration, resulting in loss of central vision.

On the other hand, macular degeneration is considered to be a disease that occurs well in the elderly, but it is known that the number of patients in their 4's and 50's has recently increased rapidly. The main cause of the lowered age group of macular degeneration is the westernization of diet such as increased fat intake. It is becoming.

On the other hand, lutein is a naturally occurring carotenoid (carotenoid) that does not have vitamin A activity and is present in a natural state. lutea). Lutein, one of the most carotenoids present in food and human blood, is known to play an important role in the body's antioxidant effects and to protect plants from UV rays. It is known to play an important role in health and vision enhancement.

According to US Department of Agriculture data, Americans currently consume about 1.7 mg of lutein per day and Europeans consume about 2.2 mg of lutein per day, which is cataract and age-related macular degeneration (Age-). lutein in the United States and Europe has been reported to increase the use of lutein in dietary supplements and certain food and beverage components. It has been reported that it will help to increase the intake of to more effective levels.

Lutein is known as a major component of green vegetables and fruits such as broccoli, green beans, mallow beans, cabbage, kale, spinach and kiwi, but the green vegetables and fruits coexist with large amounts of chlorophyll, beta carotene and carotenoid epoxy compounds in addition to lutein. Separation and purification of lutein from these is a problem that requires a high cost and complicated procedures, especially lutein ester compound having a relatively lower physiological activity than lutein in the free state in many cases.

Lutein is also abundant in many yellow / orange fruits and vegetables such as mangoes, papayas, peaches, plums, acorns, pumpkins, zucchini and oranges. Lutein contained in these yellow / orange fruits and vegetables exists in esterified form with organic acids or fatty acids such as myristic acid, lauric acid, and palmitic acid. Since lutein ester compounds have a lower physiological activity than lutein, lutein esters must be hydrolyzed and regenerated into free lutein when consumed in food, and then absorbed and metabolized in the body. Therefore, in order to effectively use the lutein contained in these yellow / orange fruits and vegetables, it is necessary to separate and purify the glass after saponification, which induces relatively high cost and time consumption. Carotenoids also contain high concentrations, resulting in low relative purity.

To date, most of the therapeutic agents used to treat and improve macular degeneration are marigold extracts. Marigold petals are considered good sources of lutein because they contain significant amounts of lutein and not much other carotenoids.

However, marigold has relatively high purity of lutein because it contains a small amount of relatively different kinds of carotenoids, but since most of the forms present in the plant are not present as free lutein, but in the form of lutein ester compounds. In addition, marigold extract is also used to convert lutein fatty acid esters to lutein through saponification, and the resulting lutein is still contaminated with some chemical impurities despite the saponification reaction.

In order to solve the above problems, as a result of searching for the effect of inhibiting yellowing degeneration in plant extracts having relatively less toxicity than synthetic chemicals, the present inventors found that acetone extract of green tea contained a large amount of free lutein. In addition, it was found that the yellowing degeneration inhibitory effect of the green tea acetone extract comprising the same to complete the present invention.

It is an object of the present invention to provide a composition for treating and preventing yellowing degeneration with green tea acetone extract as an active ingredient.

As an example for achieving the above object, the present invention is characterized by a composition for the prevention and treatment of macular degeneration comprising the green tea acetone extract as an active ingredient.

As another example for achieving the above object, the present invention is characterized by a macular degeneration prevention and improvement health supplement comprising the green tea acetone extract as an active ingredient.

The green tea acetone extract is characterized by including lutein represented by the following formula (1) as an active ingredient.

Hereinafter, the composition for preventing and treating macular degeneration of the present invention will be described in detail.

First, “macular degeneration” as used in the specification and claims of the present invention should be interpreted to include all forms of macular degeneration disease, regardless of the age of the patient, although some macular degeneration diseases are more common in certain age groups. will be. Macular degeneration may include, but is not limited to, heredity, physical trauma, diabetes, malnutrition, bacterial infections, and the like.

Method for producing acetone extract from green tea of the present invention is as follows.

First, the green tea leaves are dried and pulverized to form a dry powder, which is immersed in acetone, and the filtrate obtained by filtration is allowed to stand at a low temperature to remove insoluble precipitate, and then saturated saline, distilled water and hexane are added to the acetone extract from which the insoluble precipitate is removed. The mixture is shaken to give a hexane fraction, which is then concentrated under reduced pressure to obtain green tea acetone extract.

Lutein has 40 carbons and 56 hydrogens, but it shows fat solubility due to its structural features including only a few hydroxyl groups (OH). Therefore, it is not dissolved at all in a completely polar solvent such as water. This fat-soluble lutein shows relatively good solubility in polar organic solvents having a polarity of 5 to 6 levels, such as alcohol or acetonitrile. However, these solvents have excellent solubility in various polar substances including sugars. When one polar organic solvent is used, lutein is extracted in addition to other polar substances including sugars, flavonoids, glycosides, and proteins. Therefore, the alcohol-based polar organic solvent is not suitable for extracting lutein in terms of purity and efficiency by the incorporation of relative polar materials.

In addition, lutein, which shows fat-soluble properties, can be extracted well using organic solvents such as ethyl acetate, chloroform, butanol, propanol, dichloromethane, ethyl ether, xylene, and hexane, which exhibit relatively moderate or non-polar characteristics. Solubility of substances with lower polarity than lutein, such as chlorophyll compounds with lower polarity, pheoptin derivatives, carotene compounds including alpha and beta-carotene, lipids, etc., is dramatically increased, resulting in greater incorporation of nonpolar substances. In terms of purity and efficiency, it was found to be more suitable for extracting lutein than acetone.

However, although the acetone used for extracting lutein from green tea in the present invention exhibits a similar level to alcohols in terms of polarity, sugar, glycosides and protein components present in a large amount in plants have a very poor solubility in acetone. When using as a solvent, the extraction is not well done, even if a small amount of cold soaking solubility is very low and precipitated in the form of precipitate, which can be removed by filtration, there is an advantage that can be extracted relatively high purity lutein Confirmed.

Acetone may be used in a weight of 5 to 20 times the weight of the green tea powder, the extraction may be applied to a variety of methods, such as low temperature dark conditions extraction, room temperature extraction, ultrasonic extraction.

In the case of low-temperature dark condition extraction is made in the refrigeration conditions, specifically 3 to 5 ℃, extraction for 11 to 12 hours was found to be high extraction efficiency of lutein. In the case of room temperature extraction, the extraction is performed at 20 to 25 ° C., and the extraction efficiency of lutein is high when the extraction is performed for 22 to 24 hours. In the case of ultrasonic extraction, the extraction time was high when the extraction time was made at 37 to 40 ℃ 160 to 180 minutes. In the case of reflux extraction, when the extraction time was made at 75 to 80 ℃ extraction time was 160 to 180 minutes lutein extraction efficiency was high.

When comparing low temperature dark condition extraction, room temperature extraction, ultrasonic extraction and reflux extraction, it was confirmed that low temperature dark condition extraction is more preferable to obtain a large amount of lutein only.

As described above, the filtrate obtained by directly immersing and filtering the dry powder of green tea in acetone is completely removed by polar immersion in acetone extract using low temperature immersion and hexane distribution, and at the same time prevents decomposition of lutein by heat and oxidation, This makes it possible to stably extract a large amount of lutein with high purity.

The filtrate extracted using acetone as described above is allowed to stand for 12 to 24 hours at low temperature (4 to 20 ℃) to filter the insoluble precipitate. The insoluble precipitate contains various polar substances such as sugars, glycosides, phenolic substances, polar proteins, and vitamin complexes which are recrystallized due to extremely low solubility in acetone at low temperature. The advantage is that some of the polar materials that are less soluble in acetone can be removed through non-thermal treatment.

Some polar materials and mesopolar materials still coexist in the filtrate from which insoluble precipitates are removed at a low temperature as described above, and thus mixed polar materials and mesopolar materials using solvent distribution, which is a non-thermal treatment method that can minimize structural breakdown. Remove it completely.

That is, saturated saline and distilled water are added to the acetone filtrate from which the insoluble precipitate is removed, and hexane layer, which is the upper layer in which lutein is dissolved, is induced by shaking vigorously by adding hexane and a low specific gravity to hexane. By recovering, it can separate and recover without mixing polar substance.

On the other hand, acetone is an organic solvent and is well mixed with other organic solvents or distilled water.Acetone and other organic solvents and distilled water are simply added when distilled water or polar solvent is simply added to remove the polar substance from the acetone extract. Due to the good mixing of each other, it is not possible to induce the separation of the layers and the mixing occurs. Therefore, it is impossible to remove the mesopolar and polar substances in the acetone extract through the layer separation using the polarity of the actual solvent.

Thus, in the present invention, saturated saline is used to remove some mesopolar and polar substances mixed in the acetone extract. As such, the method of using saturated brine can solve the problem caused by heat treatment because it is a non-thermal treatment. In addition, when saturated saline and distilled water are added to the acetone extract at the same time, the saturated saline used with distilled water acts as a polar solvent, but the distilled water is almost dissolved in acetone by the added saturated salt. The polar material may melt, but the level of solubility that can dissolve even the nonpolar material is completely lost. At this time, when hexane, a nonpolar solvent, is added, lutein, a nonpolar substance contained in acetone extract, naturally transitions to a hexane layer having a relatively higher polar affinity than a saturated saline solution. Since the transition to the hexane layer, it is possible to exclude the mixing of the polar material.

Using the hexane partitioning, the intermediate and polar substances were removed, and the green tea acetone extract which had been removed from the hexane solution containing high purity lutein free of hexane as a solvent was concentrated and subjected to column chromatography to obtain a fraction of lutein. Each fraction is concentrated to dryness and redissolved and then separated purely using preparative HPLC.

The structure of the obtained lutein was identified by various spectroscopic methods such as NMR, MS, etc. As a result, a lutein having the structure shown in Chemical Formula 1 was obtained.

The lutein plays an important role in protecting the plant from ultraviolet rays and antioxidant effects in the plant body, and is widely known to humans as a major component of the macular and lens of the eye to enhance eye health and vision. In addition, the macular pigment density is affected by the concentration of lutein in the serum according to the intake of lutein, and if the macular pigment density is increased by the intake of lutein, the visual acuity is improved, especially in the state of macular degeneration. Increasing the content of lutein as a diet has been reported to stimulate the production of macular pigment may inhibit the progression of macular degeneration.

On the other hand, in the present invention it was newly confirmed that the green tea acetone extract contains a large amount of the lutein. In particular, the green tea acetone extract of the present invention was confirmed that the lutein is present in the active form of the free form in the body. This is simply obtained by extracting the active lutein in the form of free lutein, which immediately shows lutein activity in the body, by using a solvent, in order to extract lutein existing in the ester salt state from the existing marigold extract. The advantage is that you can do it.

Therefore, green tea acetone extract may be used as a composition for treating, preventing or improving macular degeneration.

Macular degeneration prevention and treatment composition comprising the green tea acetone extract or lutein of Formula 1 as an active ingredient is administered orally or parenterally during clinical administration, for example, intravenous and arterial, intramuscular, subcutaneous, intraperitoneal, mucosal Or topically (eg, eye drops), eye, transdermal, and the like.

The compositions may be formulated for oral administration such as tablets, troches, lozenges, aqueous or oily suspensions, prepared powders or granules, emulsions, hard or soft capsules, syrups or elixirs. And the like. Binders such as lactose, saccharose, sorbitol, mannitol, starch, amylopectin, cellulose or gelatin for formulation into tablets and capsules; Excipients such as dicalcium phosphate; Disintegrants such as corn starch or sweet potato starch; Lubricants such as magnesium stearate, calcium stearate, sodium stearyl fumarate or polyethylene glycol wax. In the case of a capsule formulation, in addition to the above-mentioned substances, a liquid carrier such as fatty oil is contained.

In addition, the composition of the present invention can be administered parenterally, and parenteral administration is applied to subcutaneous injection, intravenous injection, intramuscular injection, intrathoracic injection and mucosal or topical, dispersant, suppository, powder, aerosol ( Nasal sprays or inhalants), eye drops, gels, suspensions (aqueous or non-aqueous liquid suspensions, oil-in-water emulsions or water-in-oil emulsions), liquid dosage forms suitable for parenteral administration, such as solutions. To formulate into a parenteral formulation, the composition may be mixed in water with stabilizers or buffers to prepare a solution and formulated in unit dosage forms of ampoules or vials.

The effective dosage of the green tea acetone extract of the present invention or the lutein of Formula 1 may vary according to the age, physical condition, weight, etc. of the patient, but is generally 1 to 20 mg / day per kg of adult patient, preferably Is 5 to 10 mg / day, and may be divided into several times a day, preferably two to three times a day at regular time intervals according to the judgment of a doctor or a pharmacist.

In addition, the present invention, green tea acetone extract or lutein represented by the formula (1) has been identified and separated from edible green tea, so it is safe for human body and suitable for use in food, developed as a health food having a macular degeneration prevention and improvement effect Can be.

When the green tea acetone extract of the present invention or the lutein represented by Formula 1 is included in a health food, it may be used in a range of 0.01 to 50% (w / w), preferably 1 to 30% (w / w) of the total weight. There will be.

That is, various foods can be prepared by conventionally known methods by adding them to conventional palates such as noodles, tofu, cereals, breads, chewing gum, candy, confectionary, etc., such as ramen and raw noodles, and applying them as edible pigments. It may also be formulated into a general formulation such as tablets, granules, pills, hard capsules, soft capsules or liquid formulations, and may be prepared in a juice, pouch, beverage, or a variety of ingredients, in addition to the above ingredients Silver may be appropriately selected and blended by those skilled in the art according to the formulation.

According to the present invention described above, the effect that the green tea acetone extract containing a large amount of lutein can be used as a composition for the treatment, prevention and improvement of macular degeneration.

According to the present invention described above, since the green tea acetone extract containing a large amount of lutein in the free lutein state of lutein active in the body can be provided, the effect that can provide a new source of lutein can be expected.

Figure 1 shows the HPLC chromatogram of green tea acetone extract.
Figure 2 shows the HPLC chromatogram of acetone extract to improve the purity of lutein by removing the mesopolar and polar substances by performing a low temperature immersion and hexane fraction of green tea acetone extract.
3 is an NMR spectrum of lutein identified in green tea acetone extract.
4 is a mass spectrometry spectrum of lutein identified from green tea acetone extract.

Hereinafter, the present invention will be described in detail with reference to examples, but the present invention is not limited by the following examples.

Example  1. Preparation of Green Tea Acetone Extract and Isolation of Lutein

The green tea leaves were pulverized and passed through a 60 mesh sieve. Acetone 2L was added to 20 g of the pulverized sample and extracted for 3 hours in an ultrasonic extractor at 40 ° C., and extracted three times in the same manner. All the extracted solutions were combined in Whatman No. Filtration was carried out using 2 spots, and the insoluble precipitate was refiltered while standing at -20 ° C low temperature for 24 hours.

600 mL of the filtered acetone extract was transferred to the fractional filter, 200 mL of saturated saline solution and 1,000 mL of distilled water were added thereto, and 300 mL of hexane was added thereto, followed by vigorous shaking to induce layer separation. The saline layer was discarded and the hexane layer of the upper layer was completely recovered.

The filtered hexane layer was concentrated in a vacuum concentrator at 40 ° C. to prepare a green tea acetone extract in a black brown gel (gel) state in which the hexane solvent was removed.

The green tea acetone extract was separated using preparative HPLC (Agillent 1200 series, USA), and the column was separated by preparative HPLC, and the column was HiQ sil C18-10 (21.0 × 250 mm, KYA TECH, Japan). Analytical wavelength 430 nm, flow rate 10 mL / min, solvent A for solvent 75% methanol, solvent B for ethylacetate gradient (0 min: 70% A, 25 min: 15% A, 26 min: 70%) A, 35 minutes: 70% A), and the sample injection amount was adjusted to 2mL to perform high purity separation. As a result of the separation, several compounds showing yellow, green and brown color were isolated, and each compound was identified by performing ultraviolet visible spectral characteristics and mass spectrometry of these isolates. Particularly, the lutein corresponding to Peak 1 of the isolated compound was identified. NMR and additional mass spectrometry were performed to analyze the complete chemical structure.

Peak Compound lambda max (nm) MS (m / z, M + H) One ( all - trans ) -lutein  422, 446, 474 569 2  chlorophyll b  462, 598, 648 907 3  chlorophyll a  414, 432, 582, 616, 664 894 4  pheophytin b  414, 436, 598, 654 886 5  pheophytin a  410, 505, 535, 608, 665 872 6  β-carotene  452, 476 538

Lutein peak (peak) ¹ H-NMR of 1 for the (300MHz, CDCl ₃₎ The results of measuring a spectrum ¹ H- ¹ H correlation (correlation) of Me-18 '(1.63ppm) and when viewed H Assuming that -3 '(4.25 ppm) correlates with the front H-2' (eq, 1.86 ppm and ax, 1.38 ppm) protons of the alicyclic compound region, the peak at 5.55 ppm is H-4 ' could determine, H-7 '(5.44ppm) is H-8' could be identified by (6.15ppm) and ¹ H- ¹ H correlation of the signal H-6 Pigment '(2.42ppm). In addition, protons of two secondary hydroxy groups corresponding to H-3 and H-3 'were found in the 4.00 ppm and 4.25 ppm regions, respectively, and H-16 / H-2ax by 1D selective NOESY. viewed as a corresponding to the H-16 / H-17, H-18 / H-17 peak indicating the correlation _{CH 3 -16, CH 3 -18,} H-2ax, H-3 are both present in the same ring It can be seen. In addition, the CH ₃ bound to the hydrocarbon group can be identified by the NOESY correlation of the H-19 / H-7, H-19 / H-10 and H-20 / H-14 peaks. 19 'and H-20' were identified. The proton peak of the olefin group was found to be between 5 and 7 ppm. In particular, the multiplet signal up to 6.65 ppm was H-15 / H-15 '(6.65 ppm) and H-11 / H-11. '(6.63ppm), H-7 / H-7' (6.44ppm), H-12 / H-12 '(6.36ppm), H-14 / H-14' (6.25ppm), H-8 / H Peaks were identified by the −8 ′ (6.12 ppm) proton pair. On the other hand, protons of H-2 (1.48ppm and 1.79ppm) and H-2 '(1.38ppm and 1.86ppm) were identified in different regions from protons of other olefin groups. It was found around 1.3 to 1.8 ppm because it was paired with the surrounding methyl group instead of pairing with the protons of the group. Other methyl groups (CH ₃ , H-16-H-20, H-16'-H-20 ') were identified in the vicinity of 1-2 ppm, a region in which typical methyl peaks appear.

Position 隆_H Position 隆_H One One' 2 ax  1.48 (m)  2' ax  1.38 (dd, 13, 7) eq  1.79 (m) eq  1.86 (m) 3  4.00 (m) 3 '  4.25 (m, br) 4 ax  2.05 (m) 4'  5.55 (m, br) eq  2.38 (m) 5 5 ' 6 6 '  2.42 (d, 9.5) 7 6.12 ov ^a 7 '  5.44 (dd, 15.5, 10.1) 8 6.12 ov ^a 8' 6.15 ov ^a 9 9 ' 10 6.17 ov ^a 10 ' 6.14 ov ^a 11  6.63 (dd, 14.3, 11.4) 11 '  6.61 (dd, 15.0, 11.4) 12  6.36 (d, 14.7) 12 '  6.36 (d, 14.7) 13 13 ' 14  6.25 (m) 14 '  6.25 (m) 15  6.65 (m) 15 '  6.65 (m) 16  1.07 (s) 16 '  1.00 (s) 17  1.07 (s) 17 '  0.85 (s) 18  1.74 (s) 18 '  1.63 (s) 19  1.97 (s) 19 '  1.91 (s) 20  1.97 (s) 20 '  1.97 (s)

Example  2. Comparison of Lutein Content in Various Green Tea Resources

The acetone extracts were obtained by the method of Example 1 from 497 strains of domestic and international tea tree collection genes grown at the test packaging of Pacific Sulloccha Research Institute in Jeju Island, and then quantitatively evaluated for lutein content using analytical HPLC. 3 is shown. That is, it was confirmed that the lutein is present in a variety of content range of 99.4 to 555.8 ㎍ / g, an average of about 290.0 ㎍ / g [SD: 78.4] was confirmed, the most contained in terms of lutein content In the case of genetic resources, it shows 5.5 times higher than the resource representing the smallest content, and it can be seen that lutein contained in green tea has variety in terms of content. In addition, the average lutein content is about 5.6% of the total amount of the multi-pyrrole compounds (chlorophyll a, b, pheophytin a and b) and the carotenoid compounds present in the green tea acetone extract. Predictable applicability as a source of new lutein.

Content (µg / g) Lutein Chl b Chl a Phe b Phe a B-carotene Total at least 99.4 187.9 18.8 25.5 317.6 126.4 775.9 maximum 555.8 1,111.0 4,096.1 407.2 3,745.9 1,593.8 11,510.0 Average 290.0 569.7 1,676.4 165.0 2,029.8 459.8 5,190.9 SD 78.4 136.7 622.4 61.6 512.0 165.4 1,576.7 ratio(%) 5.6 11.0 32.3 3.2 39.1 8.8 100

Example  3. Lutein Content in Green Tea Acetone Extracts by Extraction Conditions

In order to examine the optimal extraction method of lutein from green tea, acetone extract was obtained by the method of Example 1 using acetone as an extraction solvent using domestic green tea native resources, but the extraction method was a low temperature dark condition (4 ℃), room temperature conditions ( The extraction efficiency of lutein for each extraction method was examined when the extraction method was changed by 25 ° C.), ultrasonic wave (40 ° C.) and reflux extraction (80 ° C.). In the following Tables 4 to 8, the capitalization of English letters is the result of statistical processing by Duncan's multiple test at 95% confidence level, indicating that A is the highest result.

Extraction amount at low temperature dark condition (4 ℃, ug / g) Lutein Chl b Chl a Phe b Phe a B-carotene 10 minutes 35.6 E 111.2 E 194.7 E 22.2 F 539.7 E 58.4 F 30 minutes 32.4 E 115.2 E 195.9 E 23.0 EF 572.2 DE 53.4 F 60 minutes 37.4 E 119.2 E 208.8 E 23.3 EF 576.8 DE 63.2 F 2 hours 48.1 D 137.0 D 238.8 D 25.1 DE 618.5 D 105.0 E 3 hours 52.2 CD 141.5 CD 246.4 CD 25.8 CD 628.8 D 126.8 D 6 hours 54.0 C 151.6 C 261.2 C 27.8 C 685.4 C 137.2 CD 9 hours 59.1 B 176.7 B 302.9 B 32.3 B 802.9 B 146.9 BC 12 hours 66.2 A 183.5 AB 315.7 AB 33.3 AB 825.8 B 175.1 A 24 hours 61.1 B 191.5 A 326.9 A 35.0 A 881.0 A 157.6 B

As shown in Table 4, the extraction of lutein and beta-carotene was the best for 12 hours in the case of 4 ° C. low temperature dark condition extraction, and other multi-pyrrole compounds such as chlorophyll a and b and pheophytin a and b for 24 hours Extraction showed the highest extraction efficiency.

Extraction amount at room temperature (25 ℃, ug / g) Lutein Chl b Chl a Phe b Phe a B-carotene 10 minutes 18.9 C 111.1 G 173.4 E 22.8 G 558.6 G 32.4 D 30 minutes 21.2 BC 121.0 F 187.0 E 25.2 F 611.4 F 34.9 D 60 minutes 19.6 BC 118.7 FG 178.7 E 26.1 F 625.0 F 32.0 D 2 hours 32.7 AB 147.6 E 227.2 D 30.9 E 755.6 E 67.1 B 3 hours 38.5 A 157.8 D 247.8 C 31.1 E 767.1 E 88.7 A 6 hours 29.7 ABC 162.1 D 247.2 C 35.8 D 865.6 D 49.3 C 9 hours 31.6 ABC 170.6 C 256.6 BC 37.3 C 909.0 C 58.9 BC 12 hours 31.1 ABC 180.2 B 270.0 B 39.0 B 954.4 B 52.8 C 24 hours 40.5 A 224.9 A 347.6 A 47.2 A 1150.7 A 61.6 BC

As shown in Table 5, 24 hours of lutein at 25 ℃ room temperature extraction showed the highest pattern, the extraction pattern was lower than 4 ℃ low temperature dark conditions, beta carotene 3 hours extraction was the best Also, the extraction pattern was lower than that of 4 ° C low temperature dark condition. Other multi-pyrrole compounds, chlorophyll-a and b, pheophytin-a and b, showed the highest extraction efficiency for 24 hours extraction, and showed higher or similar extraction patterns than 4 ° C low temperature dark conditions.

Extraction amount by ultrasonic extraction (40 ℃, ug / g) Lutein Chl b Chl a Phe b Phe a B-carotene 10 minutes 22.7 E 137.0 E 199.9 E 27.9 F 668.2 F 36.3 D 20 minutes 26.0 DE 155.3 D 227.2 D 32.5 E 774.5 E 35.9 D 30 minutes 29.2 CD 157.8 D 231.4 CD 31.7 E 770.5 E 46.3 C 60 minutes 32.9 C 174.6 C 256.2 C 37.1 D 891.6 D 49.9 C 90 minutes 44.0 B 194.7 B 291.0 B 39.9 C 971.4 C 68.7 B 120 minutes 47.3 B 211.8 A 311.3 B 43.1 B 1040.7 B 75.9 B 180 minutes 63.3 A 222.8 A 362.6 A 46.4 A 1138.8 A 90.6 A

As shown in Table 6, in the case of ultrasonic extraction at 40 ℃ condition all components showed the highest pattern in 180 minutes extraction, except for beta carotene extraction efficiency of all components is higher than or similar to 4 ℃ low temperature extraction An aspect was shown.

Extraction amount by reflux extraction (80 ℃, ug / g) Lutein Chl b Chl a Phe b Phe a B-carotene 10 minutes 48.0 D 166.8 E 267.7 D 30.1 D 745.6 F 112.8 E 20 minutes 50.7 D 171.7 E 276.5 D 32.0 D 792.4 E 117.8 D 30 minutes 59.9 C 194.2 D 313.1 C 36.7 C 915.2 D 111.5 E 60 minutes 58.7 C 207.4 C 335.4 B 41.3 B 1024.2 C 139.5 C 90 minutes 65.1 B 213.2 C 335.0 B 42.6 AB 1057.4 C 144.5 B 120 minutes 65.8 B 224.0 B 362.8 A 44.4 A 1114.0 B 150.0 A 180 minutes 72.7 A 233.3 A 376.1 A 45.1 A 1163.5 A 148.5 AB

As shown in Table 7, all components except beta-carotene (120 minutes extraction is the most efficient) in the reflux extraction at 80 ℃ condition showed the highest pattern in 180 minutes extraction, beta carotene in terms of extraction efficiency Except for 4 ℃ cold extraction showed a higher aspect.

Extraction method Extraction condition Extraction amount (ug / g) Lutein Chl b Chl a Phe b Phe a B-carotene Low temperature 24 hours 61.1 A 191.5 B 326.9 B 35.0 B  881.0 B 157.6 A Room temperature 24 hours 40.5 B 224.9 B 347.6 AB 47.2 A 1150.7 A  61.6 C ultrasonic wave 180 minutes 63.3 A 222.8 A 362.6 A 46.4 A 1138.8 A  90.6 B reflux 180 minutes 72.7 A 233.3 A 376.1 A 45.1 A 1163.5 A 148.5 A

As shown in Table 8 above, when comparing the arithmetic aspects when comparing the optimum extraction time with the best extraction efficiency in each extraction method above, all components except beta-carotene are reflux extraction at 80 ° C. Although evaluated as the best, considering the ease of extraction operation, ultrasonic extraction at 40 ℃ conditions is also considered an excellent method.

The present invention described above is not limited to the above-described embodiment and the accompanying drawings, and various substitutions, modifications, and changes are possible within the scope without departing from the technical spirit of the present invention. It will be evident to those who have knowledge of.

In each figure, Chl means chlorophyll and Phe means pheophytin.

Claims (6)

Macular degeneration prevention and treatment composition comprising the green tea acetone extract as an active ingredient.
The method according to claim 1,
The green tea acetone extract is a composition for preventing and treating macular degeneration comprising a lutein represented by the following formula (1) as an active ingredient.
[Formula 1]

The method according to claim 1,
The green tea acetone extract is obtained by immersing green tea leaf dry powder in acetone and filtering the filtrate to stand for 12 to 24 hours at a low temperature of 4 to -20 ℃ to remove insoluble precipitate, saturated saline in acetone extract from which the insoluble precipitate is removed. After the addition of distilled water and hexane shaken, the hexane fraction is taken, and then the hexane fraction is concentrated under reduced pressure to remove hexane, characterized in that the composition for preventing and treating macular degeneration.
Macular degeneration improvement and prevention of functional foods, comprising the green tea acetone extract as an active ingredient.
The method according to claim 1,
The green tea acetone extract is a functional food for improving and preventing macular degeneration, characterized in that it comprises lutein represented by the following formula (1) as an active ingredient.
[Formula 1]

The method of claim 3,
The green tea acetone extract is obtained by immersing green tea leaf dry powder in acetone and filtering the filtrate to stand for 12 to 24 hours at a low temperature of 4 to -20 ℃ to remove insoluble precipitate, saturated saline in acetone extract from which the insoluble precipitate is removed. After adding a distilled water and hexane and shaking to take the hexane fraction, the hexane fraction is concentrated under reduced pressure to remove the hexane, characterized in that macular degeneration improvement and preventive health functional food.

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