KR101694660B1 - Ultrasonicating extract of Perilla frutescens buds under darkroom condition with anti-inflamatory effect - Google Patents

Abstract

The present invention relates to an extract of perilla seedlings cultivated in a dark condition with anti-inflammatory effects, in which the seeds of perilla seeds are sown and cultivated in a light source of dark (dark), fluorescent, red and blue LEDs The present invention relates to a method for producing an extract of perilla spp., Which comprises preparing an ultrasonic extract containing propanediol of perilla sprouts and then examining the antioxidative activity, antiinflammatory activity and whitening activity of the extract. The ultrasonic extract of perilla sprouts prepared according to the present invention has excellent antiinflammatory It is effective.

Description

{Ultrasonicating extract of Perilla frutescens buds under darkroom condition with anti-inflammatory effect in cancer (dark) condition with anti-inflammatory effect}

The present invention relates to a perennial after-planted perilla sprout extract in dark (dark) conditions with anti-inflammatory effects.

Perilla Frutescens is a perennial herbaceous plant belonging to the genus Laviatae. Perilla seeds are used for perilla oil, tea, porridge, and confectionery. Leaflets are used for fresh picking, pickling, canning, etc. (Choung, MG, and vegetable perilla, Korean J. Crop. Sci., 50, 171-174, 2005). The seeds of perilla seeds are germinated more than ordinary mature vegetables, and the contents of amino acids, fatty acids, carbohydrates and other substances showing antioxidative effects such as vitamins, minerals and polyphenols are higher than those of seeds and maturity. (Cicer arietinum L.), LWT-Food Sci. Technol., 40, 937-945, 1999), and is recognized as a health functional food (Khalil, AW et al., Comparison of sprout quality characteristics of desi and acceptance type chickpea cultivars 2007). There have been many studies on seeds and leafhoppers of perilla seedlings, but studies on perilla sprouts are insufficient.

In skin aging, intrinsic aging is a decline in the structure and physiological function of the skin while aging, and extrinsic aging is the aging phenomenon caused by external sunlight (Gilchrest BA Skin aging and photoaging. Dermatol Nurs 2, 79-82, 1990). In particular, external aging is caused by skin cells producing melanin pigment for the purpose of defending against ultraviolet absorption, resulting in staining, wrinkling, etc. (Wang KH et al., Cosmetic applications of selected traditional Chinese herbal medicines, J Ethnopharmacol, 353-359, 2006).

In addition, factors such as the activity of melanocytes that make melanin pigment, distribution of blood vessels, thickness of skin, carotenoid, presence of pigment inside and outside the body such as bilirubin are important factors in determining human skin color. The most important factor is melanin, a melanin produced by the action of several enzymes such as tyrosinase in melanosomes in the human body, and it is important to block ultraviolet rays emitted from the sun. Many studies have shown that these skin troubles are also associated with active oxygen. Active oxygen breaks collagen, elastin, and hyaluronic acid which are connective tissues of the dermis, causing wrinkles of the skin, oxidizing the lipid part of the cell membrane, causing cell destruction and causing diseases such as dermatitis, acne and skin cancer, It is involved in the spontaneous oxidation reaction during the formation process, and causes the causes such as stain, freckles, and wrinkles (Korean Journal of Cosmetic Science, Vol. 23, No. 1, 75-132). Vitamin C, arbutin, ascorbic acid, kojic acid, hydroquinone, and retinol collagen are used as various materials effective for improving skin aging. However, there are constant problems in product safety.

Korean Patent Laid-Open Publication No. 10-2014-0015668 discloses a skin-improving cosmetic composition containing a sprout extract such as wheat sprout, buckwheat sprout, and perilla sprout, which is a cosmetic composition containing a perilla sprout extract. A method of cultivating sprouting vegetables such as non-seeded, rapeseed, and perilla seedlings using a light source irradiation in a region has been disclosed in Korean Patent No. 10-1135588. In addition, studies on antioxidant and anti-inflammatory effects by extracting perilla sprouts with ethanol are disclosed in Jung Seung Il et al., Journal of Food Science and Technology, Vol. 46, No. 87-93, However, none of the above documents discloses an antioxidant, anti-inflammatory, and whitening functional cosmetic composition containing an extract of perilla sp. By an LED light source as an effective ingredient.

Accordingly, an object of the present invention is to provide an ultrasound extract of perilla sprouts cultivated after childhood in a dark condition with anti-inflammatory effect.

The above object of the present invention is achieved by a method for growing seedlings, comprising seeding perilla seeds and cultivating seedlings at a light source of dark, fluorescent, red, and blue LEDs; Adding propanediol to the perilla sprouts as a solvent and ultrasonically extracting the extract; Antioxidant activity, antiinflammatory activity and whitening activity of the extract were compared and confirmed.

The present invention has an excellent effect of enhancing the anti-inflammatory activity by the combination of the ultrasonic extraction method using propanediol as a solvent in the preparation of the best perilla seed extract under dark conditions.

FIG. 1 is a graph showing the antioxidative effect of an ultrasonic solvent extract of Perilla sp.
FIG. 2 is a graph showing anti-inflammatory effects of 0.5%, 1.0%, and 2.0% of ultrasound solvent extract according to the LED light source of the perilla seedlings of the present invention.
FIG. 3 is a graph showing the whitening effect of 0.5%, 1.0% and 2.0% of ultrasonic solvent extract according to the LED light source of the perilla seedlings of the present invention.

The present invention provides an extract of the perennially grown perilla sprouts in dark conditions.

The extract of the present invention can be obtained only by extraction with water, an organic solvent or a mixed solvent thereof. The organic solvent used herein is ethanol, butylene glycol, propylene glycol, propane diol, and the like. It may also be a dried product obtained by drying the extract obtained by the above extract, a diluted solution of the extract, a concentrate or an extract, or a preparation or a purified product of the extract. However, in the present invention, most preferably, the perilla seed extract is prepared by adding propanediol to a shrunken sample and extracting it using an ultrasonicator, wherein the temperature is fixed at 40 캜 and then subjected to ultrasonic treatment.

According to the present invention, it is necessary that rupture of the cell membrane by ultrasonic treatment is absolutely necessary because the embryonic cell tissue is hard. In addition, in order to whiten skin due to aging, as in the present invention, it is necessary to extract an active ingredient from the cell material but to select a polar solvent without removing any solvent.

Propanediol, which is capable of maximizing the extraction efficiency of the cell material without polar solvent removal, is preferably selected.

According to the present invention, it is most preferable to maximize the extraction of the flavonoid substance or the polyphenol component when the organic solvent is added and to treat the ultrasonic wave, and furthermore, the temperature is 35 to 40 ° C at which there is no thermal denaturation of the protein enzyme.

Hereinafter, the present invention will be described in more detail by way of examples. However, the following examples are intended to illustrate the invention and are not intended to limit the scope of the invention.

< Example  1 > According to the present invention, Perilla sprout  culture

Perilla frutescens ) seeds were purchased in 2013 in Youngwol, Gangwon province and used in the experiment while refrigerated.

After the perilla seed inoculated by 2 g cultivation vessel photosynthesis Gwangyang jaryang (PPFD: Photosynthetic Photon Flux Density) 50 μmol · m -2 · s - each grown in ^one fluorescent lamp, the red LED, blue LED, cancer (暗) Respectively. 25 ± 2 ° C., 12 hours and 18 ± 2 ° C for 12 hours under the above light source conditions, and the perilla seedlings of the present invention were grown for 8 days in total.

<Example 2> of perilla sprout cultivation was choeah to the present invention cancer (暗) conditions Preparation of propanediol ultrasonic wave extract

According to the present invention, the perilla sprouts obtained from each of the cultivated perilla sprouts according to the present invention were crushed, and 40 to 60 mL of 40% propanediol was added to 2 ~ 3 g of the shrubs, and the mixture was filtered through an ultrasolificator (JAC 4020, Jinwoo, Korea), and the temperature was fixed at 35 ~ 40 ° C or lower, respectively. Eight species of perilla sprout extracts of the present invention were obtained. All the procedures were repeated three times for 1 hour. The extracted samples were filtered with a 0.45 μm membrane filter, and then subjected to an efficacy test.

< Experimental Example  1> invention Perilla sprout  Antioxidant activity test of ultrasound extract

In this experimental example, the skin fibroblast (HDFn) was suspended in DMEM (containing 10% serum and 1% antibiotic), cultured in a 5% CO ₂ incubator at 37 ° C, .

Wells were inoculated in 96 wells at a rate of about 1 × 10 ⁵ cells per well. Then, the perilla sprout ultrasonic extract prepared according to the above example was treated with 0.5%, 1% and 2%, and then incubated at 37 ° C in a 5% CO ₂ incubator And cultured for 20 minutes. 1 mM H ₂ O ₂ was added and 20 μM DCFH-DA (stock 20 mM) was added. The reaction was monitored by a spectrofluorophotometer (excitation 485 nm, emission 535 nm) at 37 ° C. for 90 minutes . The inhibition rate of reactive oxygen species was determined by comparing the formation of reactive oxygen species with the absence of the sample as a control group, and compared with the group treated with a sample to remove active oxygen species.

As a result of the experiment, as shown in FIG. 1, The antioxidative activity of the extracts increased with time under the four light conditions. Especially, in the case of blue LED condition, the antioxidative activity of extracts showed 27% active oxygen synthesis inhibition at the maximum concentration of 2% and 24% activity at the maximum concentration of 2% And the oxygen synthesis inhibitory effect was observed.

< Experimental Example  2> invention Perilla sprout  Nitric Oxide of Ultrasonic Extract Low performance  Experiment

In this experiment, LPS (Lipopolysaccharide), an inflammation inducing substance, was artificially treated with mouse macrophage cell RAW 264.7 cell, and then it was evaluated whether or not it has an inflammation inhibitory effect.

Cells were seeded at 2 × 10 ⁵ cells per well in a 96-well plate and cultured in a 5% CO ₂ incubator at 37 ° C for 24 hours. LPS (1 ug / mL) and extracts were added at different concentrations and incubated for 24 hours. After incubation, the supernatant was taken and centrifuged at 13,000 rpm for 3 minutes. The supernatant was collected, reacted with Griess reagent 1: 1 and absorbance was measured at 570 nm.

As shown in FIG. 2, the perilla seed extract showed an inhibitory effect on NO production of 18% at a maximum concentration of 2% only in the dark condition.

< Experimental Example  3> invention Perilla sprout  Melanin synthesis of ultrasound extract Low performance  Experiment

murine melanoma (B-16 F1) cells, supplemented with 10% FBS containing a DMEM (Dulbecco's modified Eagle's medium ) in which after a 6 well plate 1 × 10 ⁵ cell is one then 5% CO ₂ incubator inoculated to each well in suspension in Cells were cultured until they were attached to the wells at least 80%. After the culture, the medium was removed, and the perilla sprout ultrasonic extract of the present invention was injected, followed by culturing at 5% CO ₂ at 37 ° C.

Cells were washed with PBS (phosphatized buffer saline) per well and the cells were recovered by trypsinization. The cells were collected and counted using a hematocytometer. The cells were centrifuged at 5,000 ~ 10,000 rpm for 10 minutes, Remove the pellet. The cell pellet was dried at 60 ° C, and then 100 μl of 1 M sodium hydroxide solution containing 10% DMSO or cell lysis buffer was added thereto. The intracellular melanin was obtained in a 60 ° C. thermostat and then absorbance was measured at 405 nm with a microplate reader.

As shown in FIG. 3, melanin synthesis inhibitory effect was shown in all light sources. In particular, it was confirmed that the highest inhibitory effect of extract at 2% treatment was 58% in red LED condition and 50% in dark condition.

INDUSTRIAL APPLICABILITY As described above, the present invention is an industrially advantageous invention because it has an excellent effect of providing an ultrasound extract having an anti-inflammatory effect of a novel perilla sprout which is artificially manipulated with an effective ingredient by cultivating in a dark condition .

Claims (2)

Growing perilla seeds under dark conditions for 12 hours, at 16-20 ° C for 2 days and then for 8 days to obtain perilla sprouts; The perilla sprouts obtained in the above step are shaded, and propanediol is added thereto to ultrasonically extract at 35 to 40 ° C, thereby producing a perilla sprout extract having an anti-inflammatory effect.
The perilla sprout ultrasonic extract according to claim 1, which is characterized in that it has an anti-inflammatory effect.

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