KR101220091B1 - Low molecular weight pectin hydrolysate preparation method and the cosmetic composition containing the same - Google Patents

Abstract

The present invention relates to a low molecular weight pectin degradation product and an anti-aging functional cosmetic composition containing the same as an active ingredient, and more specifically, to a step of decomposing high molecular weight pectin by adding an enzyme, an active low molecular weight pectin decomposition product Method for preparing an active low molecular weight pectin degradate including a process for separating, insoluble precipitate containing unreacted high molecular weight pectin through centrifugation, filtration, concentration under reduced pressure and lyophilization The present invention relates to an anti-aging cosmetic composition containing 0.001 to 30.0% by weight of a molecular weight pectin degradant and having excellent skin light damage protection, moisturizing, skin wrinkle improvement, and skin elasticity improving effect by ultraviolet rays.

Description

Low molecular weight pectin hydrolysate preparation method and the cosmetic composition containing the same}

The present invention relates to a method for producing a low molecular weight pectin degradate and to an anti-aging cosmetic composition containing the same as an active ingredient.

Pectin is a major component of the plant cell wall, a substance present in cell layers such as fruits and vegetables, or in thin layers between cell membranes. It has colloidal properties and gels can be formed when suitable acids and sugars are present. Pectin is a macromolecule (αD-1,4-polygalacturonic acid) in which D-galacturonic acid (galA) is connected by α1,4 bond, and the carboxyl group of D-galacturonic acid is methyl It has been known to be esterified or in salt form or in free radical form.

However, in fact, pectin, which exists in nature, has a much more complicated structure, and most of the pectin molecules are composed of homogalacturonan composed only of galacturonic acid, but are actually branched into various oligos and polysaccharides. It has been reported that rhamnogalacturonans are covalently bound.

The industrial use of pectin is very limited, most of which is derived from the waste skin produced as waste during the juice extraction of citrus fruits. Pectin has long been used mainly for the purpose of thickening and gel forming agents in the food field, but its use is extremely limited, especially in the cosmetic field, the use of pectin is very limited, because of the physicochemical disadvantages and commercial In addition to sexual problems, one of the main reasons is that no noticeable physiological activity has been confirmed by transdermal application such as cosmetic use.

However, in recent years, the encapsulant for encapsulating the drug by the following physico-chemical properties such as the gel forming ability of pectin (S Ahrabi F., G. Madsen, K. Dyrstad, SA Sande and Graffner C., Eur. J. Pharm. Sci , 2000 , 10 (1), 43-52) or its value as a drug carrier (M. Ashford, J. Fell, D. Attwood, H. Sharma and P. Woodhead, J. Control . Release., 1993, 26 ( 3), 213-220,. M. Ashford , J. Fell, D. Attwood, H. Sharma and P. Woodhead, J. Control. Release, 1994, 30 (3), which is recognized 225-232) In addition to these physicochemical characteristics, there are reports that pectin has various physiological activities, and interest in the pharmaceutical field and related fields is increasing. The physiological activity of pectin has been known to include intestinal action, immunostimulatory activity, anti-ulcer activity, anti-tumor activity, anti-cholesterol, etc., mainly through animal experiments such as oral administration, and the pharmacological activities of pectin are mainly pectin It is known to be determined by specific structures or their microstructures such as galacturonan and rhamnogalacturonan in a molecule.

However, pectin is a highly viscous water-soluble polysaccharide component, and the viscosity change is severe due to salt, pH, etc., and is characterized by gelling with alcohols, so cosmetic application is not easy. In addition, pectin has a disadvantage of difficult transdermal penetration due to the hydrophilic high molecular weight molecular structure, and thus various physiological activities that can be expected through oral administration are difficult to appear through transdermal application, which is a form of cosmetic use.

Therefore, the present inventors have determined that cosmetics having excellent physiological activity may be developed when pectin is used as a low molecular weight in order to overcome the disadvantages of pectin as a cosmetic material and utilize various physiological activities.

To prepare low-molecular sugars from high polysaccharides, including pectin, in most cases, acid decomposition methods are used. However, in the case of acid decomposition, since polysaccharides are decomposed in a strong acid solution such as concentrated hydrochloric acid, a large amount of monosaccharides are generated, and thus the conversion efficiency is low. Finally, a neutralization process is required. Salts produced in liquids are known to affect the stability of cosmetics by changing the viscosity of major cosmetic substrates such as carbopol-based thickeners or by putting an emulsification system on them. have. In addition, chemical low-molecular production methods including acid decomposition have the advantage of efficiently producing low-molecular pectin raw materials, but have the disadvantage of random decomposition of pectin by relying on chemical reactions. Thus, with this method it is difficult to reproducibly produce low molecular pectin with specific activity. Therefore, in order to selectively produce low molecular pectin having a specific structure exhibiting bioactivity, it may be said that the application of enzyme and biotechnology is essential, but there are few research results and cosmetic applications.

Enzymes involved in pectin degradation include pectinase (pectinase, EC 3.2.1.15) or polygalacturonase (EC 3.2.1.15) and pectinesterase (EC 3.1.1.11). Can be classified as

Pectinase is an endo-type polymethyl galacturonase that hydrolyzes inside the main chain of highly esterified pectin and an exo-type polymethyl galacturonase that hydrolyzes at the terminal. (polymethyl galacturonase), an endo-type polygalacturonase that hydrolyzes low esterification pectic acid from the inside, and an exo-type polygalacturonase that degrades at its end. Pectin esterase is an enzyme that hydrolyzes methyl ester groups. Pectin lyase (4.2.2.10), which acts on the methyl esterified substrate as a degrading enzyme that acts as an endoform on pectin and pectic acid to produce sugars with unsaturated bonds between non-reducing ends C 4 and C 5 And pectate lyase (EC 4.2.2.2), which acts on free pectic acid.

Commercially available pectin hydrolase is composed of a plurality of enzyme composition among these enzyme groups, and each enzyme activity is relatively different in strength and strength, but an example of producing pectin enzymatic degradation products and using them for cosmetics There has never been.

Studies of pectin enzymes have shown that Pilnik and Voragen de-esterified HE (high ester) pectin with LE (low ester) pectin or pectinic acid, unlike pectin methylesterase (EC 3.1.1.11) mentioned as PMEs. (Food Enzymology, Ed.:PFFox; Elsevier; (1991); pp: 303-337) and Baron (1980 Lebensm. Wiss. Μ-Tecnol. 13pp 330-333) are reported by Aspergillus niger. By separating fungal PME), some PMEs de-esterify the pectin in a block-wise manner, attack the pectin in contact with non-reducing ends or free carboxyl groups and then by a single-chain mechanism Progressing along pectin molecules has been reported to produce non-esterified galacturonic acid unit blocks.

Modified pectin, which has been recently produced during the decomposition of pectin by enzymes, is applied to oral administration, resulting in detoxification of heavy metals, cholesterol regulation, and probiotic effects. There are also reports of new bioactive actions such as prebiotic effects and anti-cancer actions (Overview on Pectins.Times Food Processing Journal.June-July Issue, p. 44-51 (2006), Bioactive pectic polysaccharides from Glinus opposotofolius (L.) Aug. DC., A Malian medicinal plant, Aulation and partial characterization.J. Of Ethnopharmacrtial.Vrt. 101, p. 204-214 (2005)).

Patent Publication No. 10-0746478 discloses a method for producing a pectin oligomer using radiation characterized in that pectin is low molecular weight by irradiating the pectin, but a method for producing low molecular pectin using radiation is expensive radiation equipment and exposure. It must be equipped with a prevention facility, there is a disadvantage that can not be easily manufactured. In addition, similarly to the chemical decomposition method, there is a problem in that it is not possible to efficiently produce a decomposition product of a specific structure, and there is a possibility of causing vague concerns and avoidance of consumers about radiation.

Photo-aging means that the skin is repeatedly exposed to light such as ultraviolet rays to change the appearance or function of the skin [Ridder GM et al .: J. Am. Acad. Dermatol., 25, 751-760 (1991). More specifically, photoaging is the damage of skin, tissues, and the like by the damage of lipids, proteins, polysaccharides, and nucleic acids, which are the major constituents of the skin, by ultraviolet rays, resulting in skin aging. In particular, collagen, hyaluronic acid, elastin, proteoglycan, and fibronectin, which are the connective tissues of the skin, are severed to interfere with skin elasticity and form deep wrinkles, which are the main phenomenon of photoaging. When this becomes worse, mutations, And the immune function is reduced.

In vivo, the synthesis and degradation of extracellular matrix such as collagen is properly regulated, but as aging progresses, especially in areas exposed to ultraviolet rays, the synthesis decreases and the degradation is accelerated. The expression of matrix metalloproteinase (hereinafter referred to as 'MMP') is promoted, thereby reducing skin elasticity and forming wrinkles.

In order to prevent aging due to UV rays, the most common method is to apply a product containing sunscreen directly to the skin, but in 1988, retinoic acid was reported to be effective in reducing roughness and fine wrinkles of aged skin (KS Weiss et al. , JAMA, 259, 527-532, 1988), research is being actively conducted to develop substances that inhibit or improve skin aging worldwide. Among them, derivatives of vitamins such as vitamins A, C and E and AHA (alpha-hydroxy acid) are representative aging skin improvement substances known to date (Hermitte, Cosmetics & Toiletries, 107, 63-67, 1992, DR Rosenthal et al. , J. Invest.Dermatol., 95, 510-515, 1990, TDDitre et al, J. Invest.Dermatol, 34, 187-195, 1996). However, these materials are very unstable and have problems in use, or are somewhat insufficient to exhibit visible aging improvement effects.

Recently, in order to reduce skin irritation caused by various chemicals and the like, much attention has been focused on functional cosmetics having functionalities such as wrinkle relief and whitening obtained from natural materials. Natural materials have less side effects on the skin, and as the consumer's response to the external skin preparations using the natural materials has increased recently, the development value of the photo-aging anti-aging cosmetic composition is further increased.

Accordingly, the present inventors have studied the possibility of applying to the anti-aging cosmetic composition with respect to plant natural products with excellent safety, the physical and chemical disadvantages and compatibility of the pectin as a cosmetic raw material when decomposing the pectin by enzymatic method and low molecular weight It has been found that the problem can be solved and that the hydrophilic polymer has a disadvantage of notable notable physiological activity due to transdermal application such as cosmetic use.

The present invention provides a process for producing low molecular weight pectin decomposing products which is excellent in protecting against skin damage due to ultraviolet rays, moisturizing, improving skin wrinkles, improving skin elasticity and whitening effect, and having excellent compatibility as a cosmetic raw material and preventing skin aging. It is an object to provide a low molecular weight pectin digest with

In addition, the present invention by using a low-molecular weight pectin degradation product prepared through a low-molecular weight pectin degradation product, by confirming its efficacy as a cosmetic composition and developing a method of using the same, the purpose of increasing the selection range of various cosmetic composition raw materials It is done.

That is, an object of the present invention is to provide a cosmetic composition containing a low molecular weight pectin degradation product to prevent skin cell damage and photoaging caused by ultraviolet light.

In addition, an object of the present invention is to provide a cosmetic composition containing a low molecular weight pectin degradation product exhibiting excellent anti-aging effects, such as reducing MMP-1 biosynthesis and promoting type 1 procollagen biosynthesis, skin wrinkle improvement, skin elasticity improvement effect, etc. .

Accordingly, the inventors of the present invention have invented a method for preparing low molecular weight pectin digests having excellent compatibility as a cosmetic raw material and showing skin anti-aging activity from pectin after a long research. In this way, low molecular weight pectin digests were prepared and prepared low molecular weight pectin. As a result of experiments on skin degradation effect, skin elasticity improvement effect, and skin cell photodamage protection effect on the decomposed product, it was found that the result can be obtained, and the efficacy as a cosmetic composition including anti-aging cosmetics can be expected.

Pectin used as a raw material in the present invention can be obtained from natural products such as citrus fruits containing citrus, citrus fruits such as oranges, lemons, fruit peels and juices such as apples and juice waste including the remaining pulp, or by purchasing a commercial pectin Can be used.

The present invention relates to a low molecular weight pectin degradant having excellent compatibility from pectin as a cosmetic raw material and having skin anti-aging activity.

More specifically, the present invention

Process for preparing pectin solution,

Adding pectin degrading enzyme to the pectin solution prepared in the step and proceeding enzymatic digestion to decompose the pectin,

It provides a low molecular weight pectin digest of molecular weight 1,000 ~ 5,000 Dalton prepared by the method comprising the step of removing the unreacted pectin, low-reacted pectin and pectin degrading enzyme after the pectin decomposition process. If the molecular weight is less than 1,000 Daltons, cytotoxicity is increased. Moreover, when molecular weight exceeds 5,000 Daltons, permeability into skin cells falls.

In addition, the present invention is characterized in that the pectin solution concentration is 0.5% by weight to 10% by weight.

In addition, the pectin degrading enzyme in the present invention is not particularly limited as long as it exhibits pectin-degrading activity. Preferably, the pectin degrading enzyme is maserozyme al-10 ™ (Macerozyme R-10 ™), pectinex ultra SPL ™ (Pectinex Ultra SPL ™), Pectinex 3XL ™ (Pectinex 3XL ™), and one or more selected from Pectolyase Y-23 ™ (Pectolyase Y-23 ™).

In addition, the present invention is characterized in that the pectin degrading enzyme is added within the range of 0.02 to 1.0% by weight of pectin as a substrate.

In addition, in the present invention, the step of removing the unreacted pectin, low-reacted pectin and pectin degrading enzyme is characterized in that it is carried out by adding ethyl alcohol.

In addition, the ethyl alcohol in the present invention is characterized in that the addition of 50 to 500% by weight of the total amount of the reaction solution.

In addition, the present invention is characterized in that the low-molecular weight pectin decomposed product prepared as described above containing 0.001 to 30.0% by weight based on the whole composition, anti-aging functional containing low molecular weight pectin decomposed product having a molecular weight of 1,000 ~ 5,000 Dalton as an active ingredient. It provides a cosmetic composition.

In addition, the present invention is characterized in that it contains 0.001 to 30.0% by weight of the low-molecular weight pectin decomposed product prepared as described above with respect to the whole composition, prevention of wrinkles containing a low-molecular weight pectin decomposed product having a molecular weight of 1,000 ~ 5,000 Dalton as an active ingredient It provides a functional cosmetic composition for improvement.

The low molecular weight pectin digest of the present invention

Stage 1; Process for preparing pectin solution,

Step 2; Adding pectin degrading enzyme to the prepared pectin solution and proceeding enzymatic digestion to decompose the pectin,

Step 3; It consists of removing unreacted pectin and pectin degrading enzyme.

More preferably, in the first step of the present invention, the dried crushed pectin is dissolved in purified water and neutral salts such as basic aqueous solution, acidic aqueous solution and sodium chloride are added to adjust pH and salt concentration, and then heated or cooled to 20 to 75 ° C. Prepared with pectin solution with appropriate pH.

More preferably, in the second step of the present invention, the prepared pectin solution contains maserozyme R-10 ™, Pectinex Ultra SPL ™, and Pectinex 3XL ™ (Pectinex 3XL). ™), one or more pectinases selected from Pectolyase Y-23 ™ are added to maintain the reaction temperature and enzymatic decomposition to decompose the high molecular weight pectin.

More preferably, in step 3 of the present invention, ethyl alcohol is added to the prepared pectin enzyme digest to settle the unreacted polymer pectin and pectin enzyme, followed by filtration and centrifugation to remove the precipitate and recover the filtrate. To selectively recover low molecular weight pectin digests.

More preferably, in the fourth step of the present invention, the filtrate obtained in the third step is concentrated under reduced pressure to remove ethyl alcohol to obtain a low molecular weight pectin decomposition product.

The low molecular weight pectin digest prepared as described above may be used as it is, and may be fractionated into pectin digest according to molecular weight using an ultrafiltration apparatus, and the molecular weight may be selected and used according to the purpose of use.

The low-molecular weight pectin degradate may be concentrated and used by vacuum concentration method, and powdery pectin decomposed product may be obtained using a spray dryer, or dried and pulverized using a freeze-drying method to obtain pectin decomposed granules or granules. .

In addition, the pH of the pectin reaction solution in the method for producing a low molecular weight pectin decomposition product of the present invention is characterized in that the pH is 3.5 ~ 7.0, when the pH is out of this range, the reaction rate is reduced, the process time is delayed and economic efficiency is low , Pectin degrading enzyme activity is low, the amount of enzyme is increased, it is difficult to produce the low molecular weight decomposition products required by the present invention, there is a problem that the economic efficiency is lowered.

In addition, in the method for producing a low molecular weight pectin degradant of the present invention, the reaction temperature maintained after adding the pectin degrading enzyme to the pectin reaction solution is characterized in that 40 ~ 60 ℃, when the reaction proceeds at 40 ℃ or less In addition, the microorganisms are more likely to occur, which may cause deterioration in the safety and quality of the produced pectin low molecular weight decomposition products. If the temperature is 60 ° C. or more, the reaction rate is slowed and the enzyme quickly loses activity. This makes it difficult to produce low molecular weight decomposition products.

In addition, the method for producing a low molecular weight pectin digest of the present invention is characterized in that the reaction time maintained after adding the pectin degrading enzyme to the pectin solution is 4 to 48 hours, which is low when the reaction proceeds for less than 4 hours. In order to prepare a molecular weight pectin digest, it is necessary to increase the amount of enzyme addition, and if the reaction proceeds for more than 48 hours, there is a problem that the process cost increases.

In addition, the pectin degrading enzyme used in the method for preparing the low molecular weight pectin degradant of the present invention is not particularly limited in its kind, but maserozyme R-10 ™ (Macerozyme R-10 ™), Pectinex Ultra SPL ™ (Pectinex Ultra SPL) ™), Pectinex 3XL ™ and Pectolyase Y-23 ™, preferably using one or more pectin degrading enzymes. It is characterized in that the range of 0.001 to 0.05% by weight, and the range of 0.02 to 1.0% by weight of the substrate pectin, if the enzyme is less than 0.001% by weight of the total reaction solution, or less than 0.02% by weight of the substrate pectin addition amount The amount of enzyme is too small to efficiently degrade the substrate, the reaction rate is too slow to produce a low molecular weight pectin digest, or the bioactivity is low.

In addition, when the pectin degrading enzyme of the present invention exceeds the range of 0.05% by weight of the total weight of the reaction solution, or exceeds the range of 1.0% by weight of the amount of pectin added as a substrate, the amount of the enzyme is too high, so that the pectin as a substrate has a low molecular weight that is too low. Due to the decomposition, it is difficult to control the reaction time to selectively prepare a low molecular weight pectin digest having high anti-aging activity on the skin.

In addition, the pectin used in the low-molecular weight pectin decomposition product manufacturing method of the present invention is characterized in that the content is dissolved in the range of 0.5 to 10% by weight relative to the dry powder standard reaction liquid, and proceeds to the enzyme reaction, the content of the pectin by weight of the reaction liquid When the concentration is less than 0.5% by weight, the concentration of the pectin decomposition product recovered after the reaction is so low that the concentration of the substrate is too low to be concentrated to a concentration for efficient use in cosmetics, it is not economical, it is difficult to control the reaction rate .

In addition, when the content of the pectin exceeds 10.0% by weight, the viscosity of the reaction solution is too high, the enzyme is uniformly distributed after the addition of the enzyme, and it is difficult to react. There is a disadvantage to supply a positive amount of energy. In addition, it is difficult for the enzyme to proceed evenly with the substrate, so it may be locally overly degraded and locally unreacted.

In addition, the ethyl alcohol which can be used in the step of selectively recovering the low-molecular weight pectin degradate of the three-step low-molecular weight pectin decomposed product of the present invention is added in the range of 50% by weight to 500% by weight relative to the weight of the reaction solution, and thus unreacted pectin. And removing the degrading enzyme and recovering the filtrate. When the amount of ethyl alcohol is less than 50% by weight based on the weight of the reaction solution, it is difficult to efficiently precipitate the unreacted pectin, and the polymer is not reacted with the desired pectin. It is difficult to selectively separate low-molecular weight pectin decomposition products having excellent compatibility as skin raw materials and having anti-aging activity, and the size of the reaction container when adding more than 500% by weight of ethyl alcohol to the weight of the reaction liquid Is too large and the amount of reaction liquid is too high, the process cost is excessive, and the cost of ethyl alcohol removal through decompression concentration is excessive. As well as, it is not economical because it uses an excess of ethyl alcohol.

In addition, the method for producing a low molecular weight pectin degradant of the present invention further comprises the step of separating the pectin having a molecular weight of 500 ~ 5000 Da by membrane separation of the low molecular weight pectin through the enzymatic decomposition process.

The present invention also relates to a cosmetic composition containing a low molecular weight pectin degradant having excellent compatibility as a cosmetic raw material and having skin anti-aging activity.

In addition, the present invention provides a cosmetic composition comprising a low-molecular weight pectin degradant content having excellent compatibility as a raw material for cosmetics and having anti-aging activity on the basis of the freeze-drying weight of the entire cosmetic composition. . When the content of the low molecular weight pectin decomposition product is less than 0.001% by weight, there is almost no skin improvement effect, and when it is 30.0% by weight or more, the degree of increase in the effect of increasing the content is insignificant and economical.

In addition, the present invention is compatible with the cosmetic raw material, characterized in that the cosmetic composition is selected from the lotion, gel, water-soluble liquid, cream, essence, oil-in-water (O / W) type and water-in-oil (W / O) type formulation. The present invention provides a cosmetic composition containing, as a main active ingredient, a low molecular weight pectin degradant having excellent skin anti-aging activity.

The present invention provides a low-molecular weight pectin degradant having a low phototoxicity, high safety for the skin and various physiological activities by providing a method for producing a low molecular weight pectin degradant having excellent compatibility as a cosmetic raw material and having anti-aging activity. It can be used as an external preparation for skin or cosmetics having less skin irritation and exhibiting a predetermined functional effect.

In addition, the low molecular weight pectin digest produced by the method of the present invention can be used as a cosmetic composition exhibiting excellent anti-aging effects such as reducing MMP-1 biosynthesis and promoting type 1 procollagen biosynthesis, improving skin wrinkles, and improving skin elasticity. have.

1 is a molecular weight distribution confirmation test results using gel permeation chromatography of unreacted pectin as a raw material in the present invention.
Figure 2 is a molecular weight distribution confirmation test results using the gel permeation chromatography of the low molecular weight pectin degradation prepared in Example 5 of the present invention.
Figure 3 is a result of comparing the molecular weight distribution of the low molecular weight pectin decomposition products and the unreacted pectin raw material according to Examples 5 and 6 of the present invention using gel permeation chromatography.

Hereinafter, the present invention will be described in more detail with reference to Examples. However, these Examples are only illustrative description of the present invention and the scope of the present invention is not limited to these Examples.

Example  1, 3, 5, 7: using enzyme Low molecule Decomposition  Produce

Pectin was used by purchasing pectin extracted from citrus peel from Sigma-Aldrich. Disperse pectin in pH 4.0 50 mM acetic acid buffer and pH 5.0 50 mM citric acid buffer, heat to 50 ° C., stir to dissolve the pectin sufficiently, and prepare 0.01% by weight of maserozyme R-10 ™. , 0.0075% Pectinex Ultra SPL ™, 0.005% Pectinex 3XL ™, and 0.001% Pectolyase Y-23 ™ in pectin solution, respectively. Was added. Thereafter, the reaction was carried out in a shaking incubator for the final selected reaction time for each enzyme in the range of 50 ° C. and 14 to 20 hours. The reaction proceeded at about 50 ° C. to prevent microbial contamination. After the reaction is completed, the reaction solution is added and stirred with the same amount of ethyl alcohol as the weight of the reaction solution, and then decomposed to unreacted or higher degree of polymerization than required by the present invention through a stationary, centrifugal filtration, etc. in a 4 ° C. low temperature storage tank. Precipitated pectin was removed, and the filtered reaction solution was concentrated under reduced pressure to remove alcohol and freeze-dried to prepare a low molecular pectin digest in powder form (Examples 1, 3, 5 and 7). Production yield was found to be 14.0 ~ 14.6%.

Example  2, 4, 6, 8: Highly polymerized pectin using unreacted pectin and enzyme Decomposition  Produce

In order to prepare pectin digests having low decomposition rate and high molecular weight (high degree of polymerization) compared to the pectin low molecular weight digests of Examples 1, 3, 5, and 7, the supernatant after enzyme treatment in the preparation steps of Examples 1, 3, 5, and 7 The precipitate was recovered to prepare a pectin degradate having a high degree of polymerization.

The pectin of Examples 2, 4, 6, and 8 was prepared by precipitating and recovering the degraded pectin below the level required by the present invention (ie, in a state of high polymerization), and freezing and drying it to form a powder. Production yield was found to be about 44.3 to 53.1% (Table 1).

division Example 1 Example 2 Example 3 Example 4 Example 5 Example 6 Example 7 Example 8 pectin
Degrading enzyme Maserozyme R-10 ™ Maserozyme R-10 ™ Pectinex Ultra SPL ™ Pectinex Ultra SPL ™ Pectex 3XL ™ Teanex 3XL ™ Factorase Y-23 ™ Factorase Y-23 ™ Pectin concentration 5 wt% 5 wt% 5 wt% 5 wt% 5 wt% 5 wt% 5 wt% 5 wt% Reaction solution pH 5.0 100mM
Citric acid
Buffer pH 5.0 100mM
Citric acid
Buffer pH 5.0 100mM
Citric acid buffer pH 5.0 100mM
Citric acid buffer pH 4.0 100 mM
Acetic acid
Buffer pH 4.0 100 mM
Acetic acid
Buffer pH 4.0 100 mM
Acetic acid
Buffer pH 4.0 100 mM
Acetic acid
Buffer Reaction temperature 50 ℃ 50 ℃ 50 ℃ 50 ℃ 50 ℃ 50 ℃ 50 ℃ 50 ℃ Enzyme addition amount
(weight%) 0.01 0.01 0.0075 0.0075 0.005 0.005 0.001 0.001 Reaction time 14 hours 14 hours 18 hours 18 hours 18 hours 18 hours 20 hours 20 hours division ethanol
Sedimentation
Supernatant ethanol
Sedimentation
Redissolve ethanol
Sedimentation
Supernatant Ethanol precipitation
Redissolve ethanol
Sedimentation
Supernatant Ethanol precipitation
Redissolve ethanol
Sedimentation
Supernatant Ethanol precipitation
Redissolve Per 100g of pectin 14.24 24.31 13.96 33.06 14.56 28.18 14.16 30.95

Comparative example  1 to 4: pectin using enzyme Low molecule  plethora resolvent  Produce

In order to prepare pectin low molecular weight overlysate having high decomposition rate and low molecular weight compared to the pectin low molecular weight degradation products of Examples 1, 3, 5, and 7, the amount of enzyme was doubled and compared to the preparation conditions of Examples 1, 3, 5, and 7 The pectin low molecular weight overlysate of Comparative Examples 1 to 4 was prepared by increasing the time by 150%.

The reaction was carried out at about 50 ℃ conditions to prevent microbial contamination. After the reaction solution was terminated by the addition of the same amount of ethyl alcohol and stirring the reaction solution, the pectin was unreacted or degraded below the level required by the present invention through the stationary, centrifugal filtration, etc. The precipitated solution was removed, and the filtered reaction solution was removed under reduced pressure and freeze-dried to prepare pectin low molecular weight excessive decomposition products of Comparative Examples 1, 2, 3, and 4 in the form of a powder. 42.7% was confirmed (Table 2).

division Comparative Example 1 Comparative Example 2 Comparative Example 3 Comparative Example 4 Degrading enzyme Maserozyme R-10 ™ Pectinex Ultra SPL ™ Pectex 3XL ™ Factorase Y-23 ™ Pectin Concentration 5 wt% 5 wt% 5 wt% 5 wt% Reaction solution pH 5.0 100mM
Citric acid buffer pH 5.0 100mM
Citric acid buffer pH 4.0 100 mM
Acetic acid buffer pH 4.0 100 mM
Acetic acid buffer Reaction temperature 50 ℃ 50 ℃ 50 ℃ 50 ℃ Enzyme addition amount 0.02 wt% 0.015 wt% 0.010 wt% 0.002 wt% Reaction time 21 hours 27 hours 27 hours 30 hours Per 100g of pectin 22.30 32.68 24.63 33.26

Comparative example  5: Pectin with Acid Degradation Low molecule Decomposition  Produce

Pectin was used by purchasing pectin extracted from citrus peel from Sigma-Aldrich. After dissolving and dissolving 50 g of pectin in 1 L of 1N hydrochloric acid solution, the mixture is heated in a closed container for 2 hours at 100 ° C. for the decomposition reaction and cooled. Slowly stir the reaction solution cooled to room temperature, slowly add 2N sodium hydroxide solution to neutralize to pH 6.0, and then add and stir the same amount of ethyl alcohol as the weight of the reaction solution. The filtered reaction solution was lyophilized to prepare 15.7 g of acid-decomposed pectin low molecular weight decomposed product in the form of a powder.

Experimental Example  1: pectin Low molecule Decomposition  Check molecular weight distribution

In order to confirm the molecular weight distribution of the pectin or pectin digests prepared in Examples 1 to 8 and Comparative Examples 1 to 4, molecular weight distribution confirmation test using gel permeation chromatography was performed. Waters (USA) 2695 system was used, and the gel permeation chromatography column, Shodex (Japan), Asahipak GF-510 HQ (7.6mmID X 300mm) was used. Mobile phase used 100% of pH 4.75 acetic acid buffer. The column temperature was 50 ° C., developed under an isocratic condition of 0.6 mL / min, and confirmed by an evaporative light scattering detector (ELSD). In order to analyze the results of pectin degradation through enzymatic reaction, maltose and dextran standards were first analyzed by gel permeation chromatography, and the relationship between molecular weight and retention time was confirmed using glucose units. Glucose, maltose and dextran were found to elute from the gel permeation chromatography column at a constant rate depending on the molecular weight. The estimated degree of polymerization is a value expressed as a positive integer by dividing the estimated molecular weight of the degradation product by 194, which is the molecular weight of D-galacturonic acid, the main monomer of pectin.

division Main peak elution time Main peak estimated molecular weights (daltons) Estimated degree of polymerization (mer) Pectin before reaction 9.512 39,000 201 Example 1 17.373 1,910 10 Example 2 10.700 3,019 156 Example 3 17.139 2,361 12 Example 4 15.840 5,588 29 Example 5 16.925 2,809 14 Example 6 16.765 3,165 16 Example 7 17.146 2,347 12 Example 8 15.814 5,659 29 Comparative Example 1 18.133 713 4 Comparative Example 2 19.084 - One Comparative Example 3 18.214 610 3 Comparative Example 4 18.375 419 2

As shown in Fig. 1 and Table 3, the pectin before the enzymatic digestion reaction showed a major peak of 9.5 minutes for elution time and contained a small amount of low molecular peak near 17 minutes. The estimated molecular weight near 9.5 minutes for elution time was about 39,000 dalton. Indicates the properties of the polymer. In Example 1, the polymer peak with a dissolution time of 9.5 minutes was shifted to around 17.4 minutes, and in Example 3 and Example 7, the polymer peak was moved to near 17.1 minutes. In contrast, in Example 5, the relative resolution was low, and the main peak was found to be around 16.9 minutes. On the other hand, the decomposed product recovered the ethanol precipitates of Examples 2, 4, and 8 showed a broad spread peak over the elution time of 10 to 16 minutes, indicating that the relatively decomposed decomposed products of the decomposed products were selectively separated. . As shown in FIG. 3 comparing Example 5 and Example 6 with the pectin before the reaction, it was confirmed that the low-molecular pectin digest recovery process of the present invention recovered the low-molecular decomposition product with selectivity and efficiency.

As shown in Table 2, the pectin low molecular weight overlysates of Comparative Examples 1 to 4 exhibited a main peak after 18 minutes, and a molecular weight of about 1 to 4 when estimated as D-galacturonic acid from about 200 to around 700. Main peaks appeared in the degree of polymerization. In Comparative Example 2, the pectin was completely decomposed to be identified as the peak center near 19 minutes, which is the elution time of D-galacturonic acid.

That is, the pectin digests of Examples 1, 3, 5, and 7 prepared through the low molecular weight pectin digest production process of the present invention were confirmed to have a main molecular weight distribution in the range of 1,000 to 5,000 daltons.

Experimental Example  2: Cytotoxicity test of the sample

This example was carried out to evaluate the cytotoxicity of skin cells by treatment of the pectin or pectin lysate samples obtained in Examples 1-8 and Comparative Examples 1-5.

Fibroblasts (fibroblasts) were placed in a 24-well test plate 1 x 10 ⁵ and attached for 24 hours. Each well was washed once with PBS (phosphate buffered saline), and 1 ml of cell culture medium (DMEM supplemented with 10% FBS) and each of the samples obtained in Examples 1 to 9 diluted to an appropriate concentration And cultured for 24 hours. After 24 hours, the medium was removed, and 500 μl of the cell culture medium and 2.5 μl / ml of MTT (3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyl tetrazolium bromide) And then cultured in a 37 ° C CO ₂ incubator for 2 hours. The medium was removed and 500 μl of isopropanol-HCl (0.04 N) was added. After shaking for 5 minutes, the cells were lysed and 100 쨉 l of the supernatant was transferred to a 96-well test plate, and the absorbance at 565 nm was measured in a microplate reader. Cell survival rate (%) was measured by Equation 1, and the concentration of the sample that did not affect the survival of the cells was determined.

As can be seen in Table 4, the pectin before the reaction is 100% cell viability concentration 0.5%, the cell viability was 95.8% when 0.5% treatment was confirmed that the cytotoxicity is low, and in the case of galacturonic acid which is the main monomer constituting pectin , 100% cell viability was 0.1%, and the cell viability was 66.7% at 0.5% treatment. In contrast, the pectin low molecular weight digests and pectin low molecular weight ethanol precipitates of Examples 1 to 8 prepared using maserozyme al-10 ™, Pectinex Ultra SPL ™, Pectex 3XL ™ and Pectase Y-23 ™ were all 100% cell viability was observed at 0.25% or more, and cell survival rate was also more than 87% at 0.5% treatment.

In contrast, the pectin low molecular weight overlysates of Comparative Examples 1 to 4 showed 100% cell viability at 0.05 to 0.1%, and the cell viability at 0.5% treatment ranged from 48 to 78%, indicating relatively high cytotoxicity. It was confirmed that the cytotoxicity was significantly increased compared to the pectin low molecular weight degradation products of Examples 1, 3, 5 and 7. In addition, the low-molecular pectin lysate using acid degradation of Comparative Example 5 showed 100% cell viability at 0.05%, and 0.5% treatment resulted in 42.6% cell viability. It was confirmed that the cytotoxicity was relatively strong compared to galacturonic acid.

Therefore, the molecular weight distribution of pectin degradants with low toxicity to skin cells and low skin irritation potential can be estimated to be 1,000 or more.

Name of sample 100% cell survival concentration (%, W / V) Cell viability (%) at 0.5% treatment Example 1 0.5 102.3 Example 2 0.5 99.4 Example 3 0.25 88.5 Example 4 0.5 94.3 Example 5 0.5 106.1 Example 6 0.5 92.6 Example 7 0.25 87.6 Example 8 0.5 93.5 Comparative Example 1 0.1 77.6 Comparative Example 2 0.05 48.5 Comparative Example 3 0.1 68.4 Comparative Example 4 0.1 72.1 Comparative Example 5 0.05 42.6 pectin 0.5 95.8 D-galacturonic acid 0.1% 66.7

Experimental Example  3: Experiment for confirming skin keratinocyte activation effect of sample

This experimental example was carried out to evaluate the cell activation effect of the skin cells by low molecular weight pectin lysate sample treatment obtained in Examples 1 to 8 and Comparative Example 5.

HaCaT cells, which are human keratinocyte cell lines, were cultured for 24 hours after dispensing 100 μl in 96-well plates at a concentration of 1 × 10 ⁴ / well. Each sample was diluted with various concentrations in serum-free medium and treated for 48 hours. Remove the medium, add 500 μl of cell culture medium and 60 μl of MTT (3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyl tetrazolium bromide) solution (2.5 mg / ml) per well. Incubated for 2 hours in a 37 ℃ CO ₂ incubator. The medium was removed and 500 μl of isopropanol-HCl (0.04 N) was added. After shaking for 5 minutes, the cells were lysed and 100 μl of the supernatant was transferred to a 96-well test plate, and then 565 nm absorbance was measured in a microplate reader, and cell viability (%) was measured by Equation 1.

Name of sample 0.025% survival rate Survival rate at 0.1% treatment Survival rate at 0.5% treatment Example 1 95.3 120.7 122.5 Example 2 105.6 100.5 99.5 Example 3 100.6 113.0 121.7 Example 4 92.6 95.3 94.2 Example 5 98.5 110.6 118.4 Example 6 96.3 101.3 105.5 Example 7 106.1 125.6 129.5 Example 8 100.5 111.2 115.4 Comparative Example 5 91.6 87.3 82.4 pectin 96.8 102.3 96.3 Galacturonic acid 98.6 96.7 90.1

As can be seen in Table 5, in the case of galacturonic acid which is the main monomer constituting pectin and pectin before the reaction, it did not show a concentration-dependent or significant level of cell activation effect in the range of 0.025 to 0.5% treatment concentration. The low-molecular pectin lysate using acid degradation of Comparative Example 5 did not show a cell activation effect in the same concentration range, but rather confirmed that the cell viability was poorly reduced.

In contrast, the pectins of Examples 1, 3, 5 and 7 prepared using Maserozyme R-10 ™, Pectinex Ultra SPL ™, Pectinex 3XL ™ and Pectase Y-23 ™ All of the low molecular weight lysates showed a cell activation effect of increasing concentration-dependently in the range of 0.025 to 0.5% treatment concentration, so that the low molecular pectin lysates produced through enzymatic degradation had skin cell activation effects. In particular, the pectin low-molecular lysate prepared using the ectasease Y-23 of Example 7 showed a cell survival rate of about 129.5% at 0.5% treatment concentration, showing a very good skin cell activation effect of about 30% increase in cell proliferation. In addition, Example 2, Example 4 in which the pectin low-molecular-weight digested product obtained by recovering low-molecules into the ethanol-precipitated supernatant after the enzymatic digestion of Examples 1, 3, 5 and 7 was all prepared with the same enzyme to recover ethanol-precipitated precipitates. In comparison with the pectin enzymatic residue of Example 6 and Example 8, it can be seen that all show an excellent skin cell activation effect. This is a result showing that the low-molecular pectin degradation product with excellent physiological activity was selectively recovered through the low-molecular decomposition product of the present invention.

That is, the low molecular weight pectin oligomer material prepared by the method for producing an active low molecular weight pectin degradant having a main molecular weight distribution in the range of molecular weight 1,000 to 5,000 dalton of the present invention activates skin cells and skin while having high safety and anti-aging. It was confirmed that it is an excellent material that can exhibit effects and skin texture improvement effects.

Experimental Example  4: of the sample by ultraviolet irradiation Phototoxicity  Confirmation experiment

This experimental example was carried out to evaluate the effect on the skin cell phototoxicity and the resulting cell viability by ultraviolet irradiation by the low molecular weight pectin lysate samples obtained in Examples 1 to 8 and Comparative Example 5.

Keratinocyte was added to the 24-well test plate at 1 × 10 ⁵ cells for 24 hours. Each well was washed once with PBS (phosphate buffered saline), and 500 μl of PBS (Phosphate buffered saline) was added to each well. The keratinocytes were irradiated with 10 mJ / cm 2 of ultraviolet light using an ultraviolet B (UVB) lamp (Model: F15T8, UVB 15W, Sankyo Dennki Co., Japan), and then PBS (phosphate buffered saline) 10% FBS added) was added. After the sample was treated, it was cultured for 24 hours. After 24 hours, the medium was removed, and 500 μl of the cell culture medium and 2.5 μl / ml of MTT (3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyl tetrazolium bromide) And then cultured in a 37 ° C CO ₂ incubator for 2 hours. The medium was removed and 500 μl of isopropanol-HCl (0.04 N) was added. After shaking for 5 minutes, the cells were lysed and 100 μl of the supernatant was transferred to a 96-well test plate, and the absorbance was measured at 565 nm in a microplate reader, and cell viability (%) was measured by Equation 1.

First, the survival rate of the cells irradiated with ultraviolet rays was reduced to 57.72% compared with the cells not irradiated with ultraviolet rays. In addition, as can be seen in Table 6, in the case of galacturonic acid which is the main monomer constituting the pectin and pectin before the reaction, it did not show a concentration-dependent or significant level of increase or decrease of cell viability in the 0.1 ~ 0.4% treatment concentration range. On the contrary, the low-molecular pectin lysate using the acid decomposition of Comparative Example 5 did not show skin cell photodamage protection effect by ultraviolet rays in the same concentration range, and when treated with 0.4% concentration, the cell survival rate was 45.21%, which is lower than that of the control group. The result was confirmed.

In contrast, the pectin low-molecular lysates of Examples 1, 3, 5, and 7 prepared using pectin degrading enzymes increased cell viability and concentration-dependently in the range of 0.1 to 0.4% treatment concentration. As the cell viability was increased, UV-protective effect was observed on the low-molecular pectin degradant produced by enzymatic digestion. In particular, the pectin low-molecular lysate prepared using the pectinex 3XL of Example 5 exhibited a 74.77% cell viability at 0.4% treatment concentration, which effectively prevented cell damage caused by UV rays compared to the cell survival rate of 57.72% of the UV treatment group. Skin cell photodamage protective effect has a protective effect. In addition, Example 2, Example 4 in which the pectin low-molecular-weight decomposition products in which the low molecules were recovered as the ethanol precipitation supernatant after the enzymatic digestion of Examples 1, 3, 5, and 7 were all manufactured with the same enzyme to recover the ethanol precipitation. In comparison with the pectin enzyme degradation residues of Example 6 and Example 8, it was confirmed that the skin cell photodamage protection effect was excellent. This indicates that the low molecular weight pectin degradant having excellent physiological activity was selectively recovered through the recovery process of the present invention.

That is, the low-molecular weight pectin oligomer material prepared by the method for producing an active low molecular weight pectin degradant having a main molecular weight distribution in the range of 1,000 to 5,000 daltons of the present invention has high safety and protects against optical damage caused by ultraviolet rays of skin cells. It was confirmed that the material is an excellent material that can exhibit the anti-aging function that can delay the photoaging of the skin.

Name of sample Survival rate (%) compared to control in 0.1% treatment Survival rate (%) compared to the control when 0.2% treatment Survival rate (%) compared to the control when 0.4% treatment Example 1 60.41 62.35 66.57 Example 2 58.34 61.24 59.59 Example 3 57.64 59.59 60.12 Example 4 60.11 59.67 61.32 Example 5 64.07 69.57 74.77 Example 6 59.36 60.22 58.63 Example 7 67.19 64.49 66.16 Example 8 60.41 61.35 66.12 Comparative Example 1 60.21 55.45 45.21 pectin 58.24 58.80 60.97 Galacturonic acid 60.32 55.24 57.68

Experimental Example  5: Type 1 Procollagen Biosynthesis Promoting Effect

In this experiment, the procollagen assay was performed to investigate the effect of promoting the biosynthesis of the type 1 procollagen, a skin substrate component, after adding the low molecular weight pectin digests obtained in Examples 1 to 8 and Comparative Example 5. Was carried out in the following manner. Human dermal fibroblasts isolated from neonatal foreskin tissues were purchased from Modern Tissue Technology (MTT, Korea) and supplemented with 10% fetal bovine serum (FBS) in DMEM / F-12 (3: And cultured at a concentration of 1 × 10 ⁴ cells / cm ² . When 70 ~ 80% of the cells were grown, the cells were subcultured at a ratio of 1: 3, and the cells cultured in the third to fourth passages were used for the experiment. For the measurement of the amount of procollagen, the fibroblasts were cultured in a 48-well plate at a concentration of 90% or more, the respective samples were added at an appropriate concentration, and after 24 hours, the amount of procollagen liberated in the medium was measured with procollagen Type 1 C-peptide EIA kit (MK101, Takara, Japan).

As can be seen in Table 7, in the case of galacturonic acid, which is the main monomer constituting pectin and pectin before the reaction, it did not show a significant effect of promoting the biosynthesis of procollagen at 0.25% concentration. In addition, the low-molecular pectin digest using the acid decomposition of Example 13 was found to be a result of procollagen biosynthesis rather decreased due to cytotoxicity at the same concentration, and thus could not confirm an effective result.

In contrast, Examples 1, 3, 5 and 7 prepared using pectin degrading enzymes such as maserozyme al-10, Pectinex Ultra SPL, Pectinex 3XL, and Pectase Y-23 All of the pectin low molecular weight lysates exhibited a biosynthetic effect of more than 22% procollagen compared to the control at 0.25% concentration, so that the low molecular weight pectin lysates produced through enzymatic digestion showed very good skin basal production. In particular, the pectin low molecular weight degradation product of Example 3 exhibited a very good effect of promoting biosynthesis of 36.4% procollagen at 0.25% concentration. In addition, Example 2, Example 4 in which the pectin low-molecular-weight decomposition products in which the low molecules were recovered as the ethanol precipitation supernatant after the enzymatic digestion of Examples 1, 3, 5, and 7 were all manufactured with the same enzyme to recover the ethanol precipitation. In comparison with the pectin enzymatic degradation residues of Example 6 and Example 8, it was confirmed that the procollagen promoting effect of all excellent. This is a result showing that the low-molecular pectin degradation product with excellent physiological activity was selectively recovered through the low-molecular decomposition product of the present invention. In addition, the biosynthesis-promoting effect of procollagen from low molecular weight pectin was lower than the concentration of TGF-β, a cell signaling pure substance used as a positive control group, but showed very good activity in consideration of cytotoxicity and economic efficiency.

That is, the low molecular weight pectin oligomer material prepared by the method for producing an active low molecular weight pectin degradant having a main molecular weight distribution in the range of 1,000 to 5,000 dalton of the present invention promotes collagen biosynthesis of skin cells in the skin tissue while having high safety. The skin aging phenomenon and wrinkles can be improved, thereby confirming that it is an excellent material that can exhibit a very good anti-aging function.

Name of sample Content (%, W / V) Procollagen biosynthesis promotion rate (%) Example 1 0.25 30.5 Example 2 0.25 16.2 Example 3 0.25 36.4 Example 4 0.25 9.4 Example 5 0.25 22.8 Example 6 0.25 11.2 Example 7 0.25 34.6 Example 8 0.25 5.8 Comparative Example 1 0.25 - pectin 0.25 4.2 Galacturonic acid 0.25 2.4 TGF-β 0.001 35.2

Example  9 and Comparative example  6

In this example, the cosmetics containing the low molecular weight pectin decomposed product obtained in Example 3 were prepared, and the skin elasticity improvement effect and the wrinkle improvement effect were evaluated in a comparative experiment with Comparative Example 6 in humans.

The cosmetics used in the comparative experiments are in cream form and their compositions are shown in Table 8. First, the phase b) recorded in Table 8 is heated and stored at 70 ° C. To this, a) phase is added, and after pre-emulsification, it is emulsified uniformly with a homomixer, and then cooled slowly to prepare a cream (Example 9, Comparative Example 6). 20 experimenters (20-35 year old female) were applied to the cream prepared in Example 9 on the right side of the face and the cream prepared in Comparative Example 6 on the left side of the face for 2 consecutive days, 2 times a day. . The skin elasticity was measured by using a skin elasticity meter (SEM 575, C + K Electronic Co., Germany) before and after using the product for 2 months. The experimental results are shown in Table 9 below as the ΔR7 value of Cutometer SEM 575, where the R7 value indicates viscoelasticity of the skin.

As shown in Table 9, it can be seen that the cream containing the low molecular weight pectin digest of Example 3 (Example 9) has an excellent skin elasticity improvement effect of the applied experimenter.

Example 9 Comparative Example 6 end Stearyl alcohol 8 8 Stearic acid 2 2 Stearic acid cholesterol 2 2 Squalane 4 4 2-octyldodecyl alcohol 6 6 Polyoxyethylene (25 mole addition) alcohol ester 3 3 Glyceryl monostearate Aster 2 2 I Low Molecular Weight Pectin Degradate of Example 2 One - Propylene glycol 5 5 Suitable amount Suitable amount Suitable amount

Note) Unit: wt%

Experimental product Skin elasticity effect (△ R7) Example 9 0.33 Comparative Example 6 0.13

n = 20, p <0.05

20 experimenters (women aged 20-35 years) were applied to the cream prepared in Example 9 on the right side of the face, and the cream prepared in Comparative Example 6 on the left side of the face, twice a day for 2 consecutive months. To evaluate the wrinkle improvement effect of the skin before and after the use of the product after the completion of the experiment, a silicone replica was made and the wrinkle state of the designated area was measured with a visiometer SV60, C + K Electronic Co., Germany). The results are shown in Table 10, which shows the average of the parameter values after 2 months minus the parameter values two months ago. In other words, the negative the value, the higher the wrinkle improvement effect. As shown in Table 10, the cream of Example 9 containing the low molecular weight pectin digest of Example 3 was confirmed that the skin wrinkle improvement effect was greatly enhanced.

Experimental product R1 R2 R3 Example 9 -0.27 -0.21 -0.10 Comparative Example 6 -0.12 -0.10 -0.03

R1: difference between the highest and lowest of the wrinkle contour

R2: The wrinkle contour line is arbitrarily divided into 5 squares, and the average of R1 values

R3: Highest value of R1 divided by 5

Other examples are shown below. That is, a lotion, an emulsion and a cosmetic liquid containing the low molecular weight pectin decomposed product obtained in Example 3 were prepared in Examples 10 to 12. The lotion, milky lotion and essence containing these low-molecular weight pectin degradants show excellent effects in preventing skin photoaging, preventing skin aging and improving skin aging, including free radical scavenging, skin photodamage protection, skin wrinkle improvement, and skin elasticity. It was.

Example  10: Low molecular weight  pectin Decomposition  Containing lotion

To 8 g of 95% ethanol, 0.05 g of polypyrrolidone, 0.1 g of oleyl alcohol, 0.2 g of polyoxyethylene monooleate, 0.2 g of fragrance, 0.1 g of paraoxybenzoic acid methyl ester, a small amount of antioxidant, and a small amount of pigment are dissolved. . 0.05 g of the low molecular weight pectin digest obtained in Example 3 and 5 g of glycerin were dissolved in 85.33 g of purified water, and then the mixed solution was added and stirred to obtain a skin lotion having a skin improvement effect.

Example  11: Low molecular weight  pectin Decomposition  Manufacture of emulsion containing

1.2 g of cetyl alcohol, 10 g of squalane, 2 g of vaseline, 0.2 g of paraoxybenzoic acid ethyl ester, 1 g of glycerin monoesteralide, 1 g of polyoxyethylene (20 mole addition) monooleate and 0.1 g of fragrance were mixed and dissolved at 70 ° C. , 0.5 g of the low molecular weight pectin decomposed product obtained in Example 3, 5 g of dipropylene glycol, 2 g of polyethylene glycol-1500, 0.2 g of triethanolamine, and 76.2 g of purified water were heated to 75 ° C. to dissolve it. Both mixtures were emulsified and cooled to obtain an emulsion having an oil-in-water (O / W) type skin improvement effect.

Example  12: Low molecular weight  pectin Decomposition  Containing Serum  Produce

To 5 g of 95% ethyl alcohol, 1.2 g of polyoxyethylene sorbitan monooleate, 0.3 g of kitulose, 0.2 g of sodium hyaluronate, 0.2 g of vitamin E-acetate, 0.2 g of sodium licorice, 0.1 g of paraoxybenzoic acid ethyl ester 1g of the low molecular weight pectin digest obtained in Example 3 and a suitable amount of pigment were mixed to obtain a cosmetic solution having an effect of improving skin.

Claims (8)

Process for preparing pectin solution,
Adding pectin degrading enzyme to the pectin solution prepared in the step and proceeding enzymatic digestion to decompose the pectin,
It contains 0.001 to 30.0% by weight of the low-molecular weight pectin decomposed product having a molecular weight of 1,000 ~ 5,000 Dalton prepared by the method comprising the step of removing unreacted pectin, low-reacted pectin and pectin degrading enzyme after the pectin decomposition step An anti-aging functional cosmetic composition comprising a low molecular weight pectin degradation product as an active ingredient.
The method according to claim 1,
The pectin solution is characterized in that the concentration of 0.5% by weight to 10% by weight, anti-aging functional cosmetic composition containing a low molecular weight pectin degradation product as an active ingredient.
The method according to claim 1,
The pectin degrading enzyme is characterized in that the range of 0.02 to 1.0% by weight of the substrate pectin, anti-aging functional cosmetic composition containing a low molecular weight pectin digest as an active ingredient.
The method according to claim 1,
The step of removing the unreacted pectin, low-reacted pectin and pectin degrading enzyme is characterized in that the addition of ethyl alcohol, anti-aging functional cosmetic composition containing a low-molecular weight pectin digest as an active ingredient.
Process for preparing pectin solution,
Adding pectin degrading enzyme to the pectin solution prepared in the step and proceeding enzymatic digestion to decompose the pectin,
It contains 0.001 to 30.0% by weight of the low-molecular weight pectin decomposed product having a molecular weight of 1,000 ~ 5,000 Dalton prepared by the method comprising the step of removing unreacted pectin, low-reacted pectin and pectin degrading enzyme after the pectin decomposition step A functional cosmetic composition for preventing or improving wrinkles, comprising a low molecular weight pectin degradate as an active ingredient.
The method according to claim 5,
The pectin solution is characterized in that the concentration is 0.5% by weight to 10% by weight, functional cosmetic composition for preventing or improving wrinkles containing a low molecular weight pectin decomposition product as an active ingredient.
The method according to claim 5,
The pectin degrading enzyme is characterized in that the range of 0.02 to 1.0% by weight of the substrate pectin, functional cosmetic composition for preventing or improving wrinkles containing a low molecular weight pectin digest as an active ingredient.
The method according to claim 5,
The step of removing the unreacted pectin, low-reacted pectin and pectin degrading enzyme is characterized in that the addition of ethyl alcohol, functional cosmetic composition for preventing or improving wrinkles containing a low molecular weight pectin degradation product as an active ingredient.

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