CN115919722A

CN115919722A - Pandanus communis leaf extract and preparation method and application thereof

Info

Publication number: CN115919722A
Application number: CN202211286012.6A
Authority: CN
Inventors: 张兴琪; 龙艳; 彭春梅; 高标; 苏潇雨; 刘瑞学; 冷群英
Original assignee: Guangdong Bawei Biotechnology Co ltd
Current assignee: Guangdong Bawei Biotechnology Co ltd
Priority date: 2022-08-26
Filing date: 2022-10-20
Publication date: 2023-04-07

Abstract

The invention discloses a preparation method of a Pandanthus miniatus leaf extract, which comprises the following steps: cleaning fresh Pandanum Paniculatum leaves, cutting into segments, drying at low temperature, pulverizing with a pulverizer, and sieving to obtain Pandanum Paniculatum leaf powder; weighing a proper amount of the Pandanus communis leaf powder, adding a solvent with a proper material-liquid ratio for extraction, extracting for 2 times at the temperature of 50-85 ℃ for 1-3h each time, collecting and combining supernate after the extraction is finished, centrifuging for 5min at the speed of 4000r/min, and concentrating the supernate under reduced pressure to 3-5 times of the weight of the Pandanus communis leaf powder to obtain Pandanus communis leaf concentrated solution; then adding a proper amount of stabilizer for redissolution, adding active carbon, standing for 4-8h, and performing membrane filtration to obtain a finished product. Also discloses the Pandanus communis leaf extract obtained by the method and application thereof in cosmetics, and the Pandanus communis leaf extract has good moisturizing and relieving effects while ensuring self-green, natural, safe and stable properties.

Description

Pandanus communis leaf extract and preparation method and application thereof

Technical Field

The invention discloses a plant extract derived from Pandanus of Pandanaceae, in particular to a Pandanus vannamei leaf extract and a preparation method and application thereof, belonging to the technical field of cosmetics.

Background

With the continuous development of the cosmetic industry and the increase of skin problems caused by the pressure of life, more and more people begin to use cosmetics, and in addition, the increase of the types and the selection difference of cosmetics in the industry in recent years, the functional raw materials of the cosmetics gradually get attention of people, and the ingredients of the cosmetics come to emerge. The hot effective components such as nicotinamide and arbutin are widely applied, natural effective plant materials such as liquorice, green tea, centella, aloe and the like are well pursued, and cosmetic innovation gradually changes from product innovation to raw material innovation. In addition, the demand of efficacy end is strong, people pay more attention to the efficacy, new plant raw materials are developed and applied to cosmetics, the activity of industrial products is increased, and the development of companies and researchers is also the key point.

The leaves of the Pandanus amorlifolius (Pandanus amarylifolius Roxb), also called the leaves of the cymbidium goeringii and the leaves of the Vanilla planifolia, are fresh leaves of the Pandanus scolosa of Pandanus of Pandanaceae, are native to Srilan, malaya and Philippine, are widely distributed in southeast Asian regions, are the only plants with aromatic odor in the leaves of Pandanus species, contain a special aroma, namely the leaf aroma, and are introduced and cultivated in China Hainan, yunnan and other places. The Pandanus communis leaves are rich in linoleic acid, squalene, vitamin K3, phytol, estragole and other components, have the effects of reducing blood fat, promoting metabolism of human bodies, accelerating organism tissue repair and the like when applied to the field of food, and are widely applied to the field of food at present. However, in the field of daily chemicals, no study on the moisturizing and relieving effects of the Pandanthus stramineus leaves applied to cosmetics is reported.

CN202010092512.0 discloses a macula She Zhipin and a preparation method thereof, but the invention mainly relates to the field of food processing, optimizes the color-preserving and aroma-enhancing process aiming at a macula leaf raw material, and does not relate to the use in the fields of specific extraction and daily chemicals.

CN202110386460.2 discloses a herba Zeylanicae leaf freeze-dried powder and a preparation process thereof, mainly relates to the technical field of tropical medicinal plants, explains the preparation process of the herba Zeylanicae leaf freeze-dried powder, and does not relate to the efficacy and the use in the field of daily chemicals.

Therefore, the development of the Pandanthus urinaria leaf extract for the field of daily chemical skin care to enhance the innovation of industrial products is very necessary.

Disclosure of Invention

In view of the above disadvantages, the present invention aims to provide a Pandanthus minimus leaf extract, a preparation method and an application thereof, wherein the extract has good skin moistening, hyaluronidase activity inhibiting, inflammatory factor release inhibiting and excellent oxidation resistance, can effectively alleviate symptoms such as skin dryness, skin sensitivity and skin pruritus, and has good moisturizing and soothing effects.

In order to achieve the above object, the first technical solution provided by the present invention is: a preparation method of a Pandanus communis leaf extract is characterized by comprising the following steps:

the method comprises the following steps: cleaning fresh Pandanum Paniculatum leaves, cutting into segments, drying at low temperature, pulverizing with a pulverizer, and sieving to obtain Pandanum Paniculatum leaf powder;

step two: weighing a proper amount of the Pandanus communis leaf powder, adding a solvent with a proper material-liquid ratio for extraction, extracting for 2 times at the temperature of 50-85 ℃ for 1-3h each time, collecting and combining supernate after the extraction is finished, centrifuging for 5min at the speed of 4000r/min, and concentrating the supernate under reduced pressure to 3-5 times of the weight of the Pandanus communis leaf powder to obtain Pandanus communis leaf concentrated solution;

step three: adding appropriate amount of stabilizer, re-dissolving, adding active carbon, standing for 4-8 hr, and membrane filtering to obtain Pandanus communis leaf extract.

Preferably, the low-temperature drying temperature in the first step is 50-70 ℃, and the sieve mesh number is 100-400 meshes.

Preferably, the ratio of the material to the liquid in the second step is 1: (5-30),

further, the extraction solvent is water, ethanol or ethanol water solution.

Further, the extraction mode in the second step is heating stirring extraction or ultrasonic extraction.

As a preference, the first and second liquid crystal compositions are, and the concentrated solution of the leaves of the Chinese mosla herb in the second step is subjected to one or more steps of alcohol precipitation or resin purification before entering the third step.

Further, the alcohol precipitation process comprises the steps of adding 2-4 times of absolute ethyl alcohol, standing at low temperature for 8-16h, collecting a precipitate, dissolving the precipitate with water, and then concentrating under reduced pressure.

Further, the resin purification process comprises the steps of adding the concentrated solution into resin, washing the resin with deionized water until the resin is transparent, eluting the resin with ethanol water, and collecting eluent.

The resin is AB-8 or D101 macroporous resin, the eluent concentration is 30-85% ethanol solution, and then the pressure reduction concentration is carried out.

Preferably, the stabilizer in the third step is one or more of glycerol, butanediol and propylene glycol

Furthermore, the membrane in the third step is a microfiltration membrane, and the aperture is 0.2-1 μm.

Furthermore, the adding proportion of the activated carbon in the third step is 0-1.5%.

The second technical scheme provided by the invention is as follows: a Pandanus communis leaf extract prepared according to any one of the above preparation methods.

The second technical scheme provided by the invention is as follows: the application of the Pandanus communis leaf extract in cosmetics.

Preferably, the cosmetic is a skin external preparation with moisturizing and relieving effects, which is prepared by taking the composition of the leaves of the Chinese mosla herb as an active component and adding conventional pharmaceutical or cosmetic auxiliary materials or auxiliary components.

Specifically, the moisturizing and relieving cosmetic is one or more of astringent, essence, cream, facial mask and gel.

Compared with the prior art, the invention has the following beneficial effects:

1. the Pandanthus minimus leaf extract disclosed by the invention has good skin moistening, hyaluronidase activity inhibiting, inflammatory factor release inhibiting and excellent antioxidant capabilities, can effectively relieve symptoms such as skin dryness, sensitivity and pruritus, has good moisturizing and soothing effects, and really meets the requirement of effective skin care.

2. The Pandanthus minimus leaf extract disclosed by the invention has the advantages of pure nature, no stimulation, safety and no toxicity, can meet the requirements of safety skin care, and has good popularization prospects and wide application markets.

Drawings

Fig. 1 is a graph of the results of the human moisture test in the efficacy test of the present invention.

Detailed Description

The following claims are provided for further explanation of the present invention in conjunction with the detailed description of the invention, but not for any limitation of the present invention, and any limited number of modifications made by anyone within the scope of the claims will still fall within the scope of the claims.

Example 1

The embodiment discloses a preparation method of a Pandanthus urinaria leaf extract, which comprises the following specific steps:

step one, taking fresh leaves of the Pandanus communis, cleaning, cutting into sections, drying at a low temperature of 60 ℃, crushing by a crusher, and sieving by a 400-mesh sieve to obtain powder of the Pandanus communis leaves.

Step two, weighing 10g of Pandanthus minimus leaf powder according to a material-liquid ratio of 1:20 adding deionized water for ultrasonic extraction with ultrasonic power of 400W at 65 deg.C for 2 times, each for 2 hr, centrifuging at 4000r/min for 5min, collecting and mixing the supernatants, concentrating under reduced pressure to obtain 50g water solution, and concentrating to obtain herba Pandani leaf concentrated solution. And step three, adding butanediol with the same mass as the concentrated solution of the leaves of the Pandanus communis to redissolve, and filtering by a 0.45-micrometer membrane to obtain the leaf extract of the Pandanus communis.

Example 2

The preparation method of the pandanus orientalis leaf extract comprises the following specific steps:

step one, taking fresh leaves of the Pandanus communis, cleaning, cutting into sections, drying at a low temperature of 50 ℃, crushing by a crusher, and sieving by a 200-mesh sieve to obtain powder of the Pandanus communis leaves.

Step two, weighing 10g of Pandanus communis leaf powder, and adding the powder into the mixture according to the feed liquid ratio of 1:30 adding deionized water for ultrasonic extraction with ultrasonic power of 500W and temperature of 75 deg.C for 2 times, each time for 2h, centrifuging at 4000r/min for 5min, collecting and mixing the supernatants, concentrating under reduced pressure to obtain 50g water solution to obtain herba Pandani leaf concentrated solution. And step three, adding glycerol with the same mass as the concentrated solution of the leaves of the Chinese pandanaceae to redissolve, adding 0.5 percent of activated carbon, standing for 6 hours, and filtering by a 1.0-micron membrane to obtain the Chinese medicinal preparation.

Example 3

Step two, weighing 10g of Pandanus communis leaf powder, and adding the powder into the mixture according to the feed liquid ratio of 1:20 adding deionized water, heating and stirring for extraction at 65 deg.C for 2 times (each for 2 hr), centrifuging at 4000r/min for 5min, collecting and mixing the supernatants, concentrating under reduced pressure to obtain 50g water solution to obtain concentrated solution.

And step three, adding butanediol with the same mass as the concentrated solution of the Pandanus communis leaves for redissolution, adding 0.1% of activated carbon, standing for 4 hours, and filtering by using a 0.45-micrometer membrane to obtain the extract.

Example 4

step one, taking fresh Pandanus communis leaves, cleaning, cutting into sections, drying at a low temperature of 70 ℃, crushing by a crusher, and sieving by a 400-mesh sieve to obtain Pandanus communis leaf powder.

Step two, weighing 10g of the Pandanus communis leaf powder, adding the mixture into the mixture according to the material-liquid ratio of 1:15 adding 50% ethanol, performing ultrasonic extraction at 60 deg.C under 400W for 2 times (2 hr each time), centrifuging at 4000r/min for 5min, collecting supernatant, concentrating under reduced pressure to obtain 50g water solution, and collecting the concentrated solution. And step three, adding propylene glycol and butanediol (the mass ratio of 1:1) which are equal to the mass of the concentrated solution of the Chinese panda leaves for redissolution, adding 1.0% of activated carbon, standing for 8h, and filtering by using a 0.45-micrometer membrane to obtain the Chinese medicinal preparation.

Example 5

Step two, weighing 10g of the Pandanus communis leaf powder, adding the mixture into the mixture according to the material-liquid ratio of 1: adding 70% ethanol into 30, heating and stirring for extraction at 55 deg.C for 2 times, each for 2 hr, centrifuging at 4000r/min for 5min, collecting and mixing the supernatants, concentrating under reduced pressure to obtain 50g water solution to obtain concentrated solution of Pandanum Paniculatum leaf.

And step three, adding propylene glycol with the same mass as the concentrated solution of the leaves of the Pandanus communis to redissolve, adding 0.5% of activated carbon, standing for 6 hours, and filtering by a 1.0-micron membrane to obtain the traditional Chinese medicine.

Example 6

step one, taking fresh leaves of the Pandanus communis, cleaning, cutting into sections, drying at a low temperature of 70 ℃, crushing by a crusher, and sieving by a 100-mesh sieve to obtain powder of the Pandanus communis leaves.

Step two, weighing 10g of Pandanus communis leaf powder, and adding the powder into the mixture according to the feed liquid ratio of 1:10 adding deionized water for ultrasonic extraction, wherein the ultrasonic power is 300W, the temperature is 65 ℃, the extraction is carried out for 2 times, each time lasts for 2 hours, the extraction is finished, centrifuging is carried out for 5 minutes at the speed of 4000r/min, the supernatant is collected and combined, then the supernatant is subjected to reduced pressure concentration, and the concentration is carried out to 50g of water solution, thus obtaining the concentrated solution of the leaves of the Pandanum henryi. And step three, adding glycerol and butanediol (the mass ratio of 1:1) which are equal to the mass of the concentrated solution of the Chinese pandanaceae leaves for redissolution, adding 0.5 percent of activated carbon, standing for 8h, and filtering by a 0.45-micrometer membrane to obtain the Chinese medicinal preparation.

Example 7

step one, taking fresh Pandanus communis leaves, cleaning, cutting into sections, drying at a low temperature of 60 ℃, crushing by a crusher, and sieving by a 400-mesh sieve to obtain Pandanus communis leaf powder.

Step two, weighing 10g of the Pandanus communis leaf powder, adding the mixture into the mixture according to the material-liquid ratio of 1:15 adding deionized water for ultrasonic extraction, wherein the ultrasonic power is 400W, the temperature is 65 ℃, the extraction is carried out for 2 times, each time lasts for 2 hours, the extraction is finished, centrifuging is carried out for 5 minutes at the speed of 4000r/min, the supernatant is collected and combined, then the supernatant is subjected to reduced pressure concentration, and the concentration is carried out to 50g of water solution, thus obtaining the concentrated solution 1 of the leaves of the Pandanum henryi. Adding 3 times of 95% ethanol into the Pandanum leaf concentrated solution 1, precipitating with ethanol, standing at low temperature for 12 hr, filtering to obtain precipitate, adding deionized water to dissolve, and concentrating to 30g water solution to obtain Pandanum leaf concentrated solution 2.

And step three, adding glycerol and butanediol (the mass ratio of 1:1) which are equal to the mass of the Pandanum paniculatum leaf concentrated solution 2 for redissolution, and filtering by using a 0.45-micrometer membrane to obtain the tea.

Example 8

Step two, weighing 10g of Pandanus communis leaf powder, and adding the powder into the mixture according to the feed liquid ratio of 1:15 adding deionized water for ultrasonic extraction, wherein the ultrasonic power is 400W, the temperature is 65 ℃, the extraction is carried out for 2 times, each time lasts for 2 hours, the extraction is finished, centrifuging is carried out for 5 minutes at the speed of 4000r/min, the supernatant is collected and combined, then the supernatant is subjected to reduced pressure concentration, and the concentration is carried out to 50g of water solution, thus obtaining the concentrated solution 1 of the leaves of the Pandanum henryi. Adding the Pandanus communis leaf concentrated solution 1 into AB-8 type macroporous resin, washing with deionized water to be transparent, adding 70% ethanol for elution, collecting eluate, and concentrating under reduced pressure to 30g water solution to obtain Pandanus communis leaf concentrated solution 2.

And step three, adding glycerol and butanediol (the mass ratio of 1:1) which are equal to the mass of the Pandanus communis leaf concentrated solution 2 for redissolution, and filtering by using a 0.45-micrometer membrane to obtain the traditional Chinese medicine.

To demonstrate the effect of the present invention, comparative experimental examples of the present application are given below:

comparative example 1

The preparation method of the Pandanus communis leaf extract comprises the following specific steps:

step one, taking fresh leaves of the Pandanus communis, cleaning, cutting into sections, drying at a low temperature of 60 ℃, and crushing by a crusher to obtain Pandanus communis leaf powder.

Step two, weighing 10g of Pandanus communis leaf powder, and adding the powder into the mixture according to the feed liquid ratio of 1: and 5, adding 70% ethanol, heating, stirring, extracting at 45 deg.C for 2 times (2 hr each time), centrifuging at 4000r/min for 5min, collecting supernatant, concentrating under reduced pressure, and concentrating to 50g water solution to obtain concentrated solution of Pandanus communis leaf.

And step three, adding deionized water with the same mass as the concentrated solution of the Pandanus communis leaves for redissolution, adding 2.0% of activated carbon, standing for 8 hours, and filtering by a 3-micron membrane to obtain the extract.

Comparative example 2

step one, taking fresh leaves of the Pandanus communis, cleaning, cutting into sections, drying at 80 ℃, crushing by a crusher, and sieving by a 100-mesh sieve to obtain powder of the Pandanus communis leaves.

Step two, weighing 10g of Pandanus communis leaf powder, and adding the powder into the mixture according to the feed liquid ratio of 1:10 adding 20% butanediol solution, performing ultrasonic extraction at 40 deg.C under 200W for 1 time (1 hr per time), centrifuging at 4000r/min for 5min, collecting supernatant, concentrating under reduced pressure to obtain 50g water solution, and concentrating to obtain concentrated solution.

And step three, adding glycerol with the same mass as the concentrated solution of the Pandanus communis leaves for redissolution, adding 0.1% of activated carbon, standing for 8 hours, and filtering by a 1.0-micron membrane to obtain the traditional Chinese medicine.

Comparative example 3

firstly, taking fresh leaves of the Pandanus communis, cleaning, cutting into sections, drying at a low temperature of 50 ℃, and crushing by a crusher to obtain Pandanus communis leaf powder.

Step two, weighing 10g of Pandanus communis leaf powder, and adding the powder into the mixture according to the feed liquid ratio of 1: and 8, adding deionized water, heating, stirring and extracting at 90 ℃ for 1 time, extracting for 1 hour each time, finishing extraction, centrifuging at the speed of 4000r/min for 5min, collecting and combining supernate, concentrating the supernate under reduced pressure to obtain 50g of water solution, and thus obtaining the concentrated solution of the Pandanthus fasciatus leaves.

And step three, adding deionized water with the same mass as the concentrated solution of the Pandanus communis leaves for redissolution, adding 2.0% of activated carbon, standing for 1h, and filtering by using a 0.45-micron membrane to obtain the traditional Chinese medicine.

To better illustrate the advantages of the present invention, the efficacy test of the extract of leaves of Pandanthus stramineus provided by the present invention is given below:

test 1, moisturizing efficacy research-weighing method

According to the difference of moisture absorption and retention performances of cosmetic components, different humectant molecules have different effects on water molecules and different capacities of absorbing and retaining water. The acting force is large, the binding force to water molecules is strong, the amount of absorbed and kept water is large, the sealing performance is good, and the water loss is small. The adhesive tape is used for simulating biological materials such as horny layer, epidermis and the like, and cosmetics are coated on the adhesive tape to simulate the application condition of the actual cosmetics. Weighing the sample after a certain time under the conditions of constant temperature and constant humidity, calculating the change of the sample mass, and evaluating the moisturizing performance of the sample by comparing the change with a reference substance with a moisturizing effect.

The experimental steps are as follows: the temperature and humidity required by the test are adjusted by the constant temperature and humidity box, and the temperature is as follows: 20 ℃ ± 2 ℃, relative humidity: 60% +/-5%. And (3) taking a glass plate and a 3M adhesive tape, attaching the 3M adhesive tape on the glass plate, weighing, and accurately obtaining 0.0001g, wherein the mark is M0. Taking a proper amount of sample (0.1 g-0.2 g), uniformly coating the sample on a glass plate stuck with a 3M adhesive tape, weighing the sample until the weight is 0.0001g, marking as M1, and placing the sample in a constant temperature and humidity box; after the standing time is 2 hours, 4 hours and 8 hours, the glass plates are respectively taken out, weighed to be accurate to 0.0001g, recorded as (M2), and immediately placed in a constant temperature and humidity box after weighing. The same procedure was taken with the control 10% glycerol. The tests were all run in parallel.

Moisture retention rate calculation formula:

moisture retention rate (%) = [ (M2-M0)/(M1-M0) ] × 100%

In the formula: the mass (g) of the glass plate of the M0-3M adhesive tape; m1, coating the sample, and then coating the 3M adhesive tape on the glass plate by mass (g); m2-mass (g) after placing in a constant temperature and humidity cabinet for several hours. The arithmetic mean of the replicates was taken as the assay result, which retained the latter decimal point with a relative deviation of <5%.

The judgment principle is as follows: and (3) determining the moisture retention rate level of the test sample and the positive control at different time by comparing the test sample with the positive control with the moisture retention efficacy under the set temperature and humidity conditions to confirm whether the product has the moisture retention performance.

The examples and comparative examples were tested for moisturizing efficacy, with the following results in terms of moisturizing ratio (%):

table 1 results of moisture retention test

As can be seen from table 1, the moisture retention rates of examples 1 to 8 and comparative example 2 in 2h, 4h and 6h are all above 40%, which indicates that the prepared panda leaf extract has a good moisture retention effect, and the moisture retention rates of examples 1 to 8 are much higher than that of the glycerin of the positive control 10%, which indicates that the panda leaf extract prepared by the process has a good moisture retention effect. Compared with the comparative examples, the moisture retention rates of all the examples 2h, 4h and 6h are higher than those of the comparative examples 1 and 3, which shows that the extract prepared according to the specified protection process generally has better moisture retention efficacy, and the prepared Pandanum paniculatum leaf extract has higher moisture retention rate and good physical moisture retention effect.

Test 2, moisturizing efficacy study-human testing

The test method comprises the following steps: the moisture retention effect of the product was verified by applying a sample to the inner side of the forearm of 30 subjects, measuring the water content of the stratum corneum of the skin before and after application, and comparing with that before use. The using method comprises the following steps: the sample was applied to one arm at 2mg/cm2. Testing parts: the forearm is flexed. Environmental conditions: the test environment requires the temperature to be 20-22 ℃ and the humidity to be 40-60%. And testing the water content of the horny layer of the skin by using a skin water content tester Corneometer for 3 times, wherein the testing times are respectively 0h, 4h and 8h.

Sample preparation: taking the extract of the leaves of the Pandanus communis Linne of the example 1, adding deionized water to dilute the extract to 5% of the extract, and fully and uniformly stirring the extract to obtain the leaf moisturizing lotion of the Pandanus communis Linne. And the efficacy test of the perfume pocket moisturizing water is carried out according to the method by taking the non-coated area as a blank control.

And (3) data statistics: the statistical analysis software is SPSS, the significance test of data improvement value normal distribution is carried out by adopting Shapiro-Wilktest, and if Sig. (double sides) > 0.01 shows normal distribution, the pairing t test is carried out. (bilateral) < 0.01, the distribution is abnormal, and Wilcoxon test is performed. If P > 0.05, no significant difference is indicated, which is recorded as "n.s."; if P is less than 0.05, the significance difference is shown, and P is more than or equal to 0.01 and less than 0.05, marked by "; p is more than or equal to 0.001 and less than 0.01, and is marked as a star; p < 0.001, note: "***". Rate of change (percentage) = (post-use data mean-pre-use data mean)/pre-use data mean 100%. The test results are shown in fig. 1.

As shown in FIG. 1, in 0-8h, the water content of the horny layer in the

blank regions

4h and 8h was reduced by 3.28% and 1.85% respectively, and the water content showed a steady trend of decreasing. The moisture retention area of the Pandanum parthenium moisture retention area shows good moisture retention effect, the water content of the skin cuticle is respectively 27.52 and 27.41 in 4h and 8h, compared with the skin cuticle moisture retention area before use, the moisture content is respectively improved by 16.61 percent and 16.14 percent, and P is less than 0.0001, and the extremely obvious difference shows that the moisture retention water has good moisture retention effect, and the Pandanum parthenium leaf extract has good short-acting moisture retention effect.

Experiment 3 DPPH radical scavenging experiment

Accurately weighing 41.67mg of DPPH reagent, fixing the volume to a 100mL brown volumetric flask, and fixing the volume to a 50mL brown volumetric flask by using anhydrous ethanol for 5mL to obtain DPPH test solution (0.1 mmol/L). The test solutions of 1ml of PPH were collected and the extracts obtained in examples 1 to 8 and comparative examples 1 to 3 were added at a concentration of 0.5ml of 4% as shown in Table 2. Fully mixing, standing for 30min in a dark place, taking absolute ethyl alcohol as a blank control, measuring the absorbance of the blank at 517nm, setting vitamin C as a control, and calculating the clearance rate of 4% concentration of each sample on DPPH according to the following formula:

DPPH clearance (%) = [1- (As-Ar)/A0 ]. Times.100%

In the formula: as is the absorbance of the reaction system of the extract and DPPH; ar-absorbance of extract-absolute ethanol; a0-absorbance of blank control with absolute ethanol.

TABLE 2 reaction solution composition

Absorbance of the solution	Addition amount of extractive solution (mL)	DPPH test solution (mL)	Anhydrous ethanol (mL)
				A0	0	1	0.5
Ar	0.5	0	1
				As	0.5	1	0

Free radicals generated by mitochondria in the process of electron transfer attack cell membranes, proteins and even DNA, so that cells are aged and even killed, and the free radicals are closely related to various diseases. Inhibiting the activity of free radicals can effectively relieve the degradation of cell structure function under the action of excessive free radicals generated after external stimulation, thereby maintaining the normal barrier structure of various organs including skin. The higher the clearance rate, the better the oxidation resistance degree is, and the barrier repair of the skin is promoted.

TABLE 3 DPPH radical scavenging test results

Sample name	DPPH clearance rate
		Vitamin C	85.63％
Example 1	82.24％
		Example 2	80.53％
Example 3	78.65％
		Example 4	83.63％
Example 5	86.26％
		Example 6	76.72％
Example 7	70.24％
		Example 8	87.46％
Comparative example 1	70.34％
		Comparative example 2	55.36％
Comparative example 3	68.57％

As can be seen from Table 3, the toilet bibs prepared in examples 1-8 and comparative examples 1-3 all have DPPH free radical scavenging rate of more than 50%, the general scavenging rate of more than 70%, and the scavenging rate is better, and the scavenging rate of examples 1-8 is higher than that of comparative examples 1-3, which indicates that the extracts prepared by the specific process have better antioxidant effect. The clearance rate of DPPH free radicals of the example 5 and the example 8 is higher than 85 percent and higher than that of a positive control vitamin C, so that the result shows that the Pandanus communis leaf extract has good clearance rate of DPPH free radicals, has good antioxidant capacity, can effectively relieve the degradation of cell structure functions under the action of excessive free radicals generated after external stimulation, and has good effect of promoting skin barrier repair.

Test 4 inhibition of Hyaluronidase Activity

Hyaluronic acid is a component that limits the diffusion of water and other extracellular substances in the tissue matrix, and after being hydrolyzed by hyaluronidase, cells become non-viscous, causing degranulation of cells and the exudation of new synthesized mediators, exerting biological effects, resulting in the occurrence of immediate type anaphylaxis. Hyaluronidase is a participant in allergic reactions, has strong allergic-related effects, and can relieve allergic phenomena and improve skin by inhibiting the activity of hyaluronidase.

The anti-allergic effects in vitro of the experimental examples 1 to 8 and the comparative examples 1 to 3 were examined by the hyaluronidase inhibition method. Taking 0.1mLCaCl2 solution (0.25 mmol/L) and 0.5mL hyaluronidase solution (100U/mL) to react in water bath at 37 ℃ for 20min; adding 0.5mL of the extracting solution, and keeping the temperature for 20min; then 0.5mL sodium hyaluronate solution (0.5 g/L) is added, and after reacting for 30min, the mixture is taken out and cooled for 5min; adding 0.1mL sodium hydroxide solution (4 mol/L) and 0.5mL acetylacetone solution (3.5 mL acetylacetone dissolved in 50mL 1.0mol/L sodium carbonate solution), boiling in water bath for 15min, and immediately transferring to ice water bath for 5min; 1mL of an Ellisib reagent (0.8 g of p-dimethylaminobenzaldehyde dissolved in 15mL of concentrated hydrochloric acid and 15mL of absolute ethanol) was added dropwise, and diluted with 3mL of absolute ethanol, the mixture was left at 25 ℃ for 20min for color development, and the absorbance was measured at 540nm with dipotassium glycyrrhizinate as a control. The hyaluronidase inhibition rate was calculated according to the following formula:

hyaluronidase inhibition = [ (A-B) - (C-D) ]/(A-B) } × 100%

In the formula: a-absorbance of control solution (the sample solution was replaced with acetic acid buffer); b-control blank solution absorbance (replace sample solution and enzyme solution with acetic acid buffer solution); c is absorbance of sample solution; d-absorbance of sample blank solution (acetic acid buffer solution was used instead of enzyme solution).

The inhibition rate of hyaluronidase by 3% of each of the samples of examples and comparative examples was examined by using 0.3% of dipotassium glycyrrhizinate as a positive control, and the results are shown in table 4:

TABLE 4 Hyaluronidase inhibition test results

As shown in Table 4, the Pandanthus communis samples prepared in examples 1-8 and comparative examples 1-3 all have hyaluronidase inhibition effects, and the inhibition rates are above 15%, and the inhibition rates of examples 1-8 are higher than those of comparative examples 1-3, indicating that the extracts prepared by a specific process have better hyaluronidase pathway inhibition effects. In addition, the inhibition rates of the examples 1 to 7 on hyaluronidase are close to 30% and are slightly lower than those of positive control, which shows that the Pandanus communis leaf extract has good inhibition on hyaluronidase, can inhibit degranulation of cells and exudation of synthesized new medium, relieves the phenomenon of allergy, and has good effects of relieving and resisting allergy.

Test 5, in vitro test for soothing efficacy-macrophage inflammation model test

Mouse macrophage is adopted, LPS is utilized to induce RAW264.7 cells to generate anaphylactic reaction, NO (nitric oxide) and inflammatory factor (IL-6) are released, and the relieving effect of the sample is evaluated by detecting the inhibiting effect of the sample on the NO content and the IL-6 content. The composition prepared in example 1 was selected for testing, the specific procedure being as follows:

1. and (3) detecting cytotoxicity: adjusting the concentration of RAW264.7 cell suspension, adding into 96-well cell plate, 37 deg.C, 5% CO ₂ Incubated overnight under conditions for detection. Cell culture medium for test sampleDiluting to different concentrations, adding into cell plate respectively, and adding into 5% CO at 37 deg.C ₂ Culturing for 20-48 h under the condition. Discarding culture solution, washing with PBS for 1-2 times, adding MTT working solution, continuing culturing, discarding culture medium after culturing, adding DMSO, shaking, mixing, measuring absorbance at 570nm, calculating cytotoxicity according to the result, and screening out sample concentration without obvious cytotoxicity for next step experiment.

2. NO release test: adjusting the concentration of RAW264.7 cell suspension, adding into 96-well cell plate, 37 deg.C, 5% CO ₂ Incubated overnight under conditions for detection. Diluting a test sample with a cell culture medium containing LPS to a non-cytotoxic concentration, adding the sample, taking the culture medium containing LPS as a Negative Control (NC), taking the cell culture medium without LPS as a Blank Control (BC), continuously culturing the cell for 20-24 h after adding the sample, taking 50 mu l of culture solution after the action is finished, putting the culture solution into a new cell plate, adding a Griess reagent with the same volume, fully mixing, reacting for 10min in a dark place, and detecting the OD value at 540 nm.

3. IL-6 inflammatory factor assay: the concentration of RAW264.7 cell suspension was adjusted, added to a 96-well cell plate, and incubated overnight at 37 ℃ under 5% CO2 conditions for detection. Diluting a test sample with a cell culture medium containing LPS to a non-cytotoxic concentration, adding the sample, taking the culture medium containing LPS as a Negative Control (NC) and the cell culture medium without LPS as a Blank Control (BC), continuously culturing the cells for 20-24 h after adding the sample, taking the supernatant after finishing the action, and carrying out an experiment according to the operating procedure of an IL-6Elisa kit.

Relative NO content (%) = OD _{Sample (I)} /OD _NC ×100％

NO inhibition (%) = (OD) _NC -OD _{Sample (I)} )/OD _NC ×100％

In the formula, OD _{Sample (I)} Means the mean value of OD values, OD, of the sample group _NC The OD value of the negative control group is the mean value.

The test was established when the relative NO content in the blank control group (BC group) was < 50%. Statistical and significance analysis was performed using GraphPad Prism software to calculate P values, P < 0.01 indicating significant differences. When P is less than 0.01 and NO inhibition rate is more than or equal to 10%, the sample can be judged to have the effect of relieving.

The calculation formula of the IL-6 down-regulation rate of the inflammatory factor is as follows:

down-regulation Rate (%) = (1-T/C). Times.100%

Wherein:

t-average value of inflammatory factor IL-6 in Experimental group

C-mean value of model group inflammatory factor IL-6

Compared with a Negative Control (NC), the IL-6 content of the sample group has a significant difference (P is less than 0.05), and when the IL-6 content of the sample group is lower than that of the Negative Control (NC), the sample can be judged to have the relieving efficacy, otherwise, the sample is judged to have no relieving efficacy.

As proved by cytotoxicity detection, when the sample concentration is 2.0029%, the cell activity is 92.87%, and no obvious cytotoxicity exists, so that the concentration of 2% is selected for carrying out the experiment, and the test results are shown in the following tables 5 and 6:

table 5 sample for NO content test results of inflammation mediators

Group of	Mean value	SD	P value	Significance of	Inhibition ratio (%)
						BC	15.87	0.24	/	/	/
NC	100.00	3.34	＜0.0001	****	0.00
						PC	65.01	0.51	＜0.0001	****	34.99％
Example 1	34.94	1.35	＜0.0001	****	65.06％

Table 5 shows the detection results of the NO release test for inflammatory mediators, and it can be seen from the detection results that P is less than 0.0001 in the NC group and the BC group have a very significant difference, and the relative content of NO in the BC group is less than or equal to 50%, indicating that the test is established. Through tests, compared with the NC group, the inhibition rate of the sample group example 1 on NO is 65.06%, P is less than 0.0001, and the result has significant difference, which indicates that the example 1 has good inhibition on NO release and has good relieving efficacy.

Table 6 results of the experiment on the content of inflammatory factor IL-6 by samples

Table 6 shows the results of the IL-6 release test, and it can be seen from the results that the IL-6 upregulation rate in the NC group is 12035.56% and P is less than 0.005, which indicates that the test is established. Through tests, compared with an NC group, the IL-6 down-regulation rate of the sample group in example 1 is 26.79%, P is less than 0.01, and the results are significantly different, which shows that the example 1 has good inhibition effect on the inflammatory factor IL-6 and has good relieving efficacy.

In conclusion, the invention researches the characteristic plant Pandanus communis leaves in the technical field of cosmetics, and prepares the Pandanus communis leaf extract through extraction and purification technologies such as heating, stirring/ultrasonic extraction, concentration, membrane filtration and the like, and the extract has good effects of moisture preservation, soothing, oxidation resistance and the like, can effectively relieve symptoms such as dry, sensitive and pruritus of skin, improves the skin state deeply and has good effects.

The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such modifications are intended to be included in the scope of the present invention.

Claims

1. A preparation method of a Pandanus communis leaf extract is characterized by comprising the following steps:

2. The method according to claim 1, wherein the low-temperature drying temperature in the first step is 50 to 70 ℃, and the mesh number is 100 to 400.

3. The preparation method according to claim 1, wherein the ratio of the material to the liquid in the second step is 1: (5-30), the extraction solvent is water, ethanol or ethanol water solution, and the extraction method is heating and stirring extraction or ultrasonic extraction.

4. The preparation method according to claim 1, wherein the concentrated solution of the leaves of the Pandanus communis in the second step is subjected to one or more steps of alcohol precipitation or resin purification before entering the third step.

5. The preparation method according to claim 4, wherein the alcohol precipitation process comprises adding 2-4 times of anhydrous ethanol, standing at low temperature for 8-16h, collecting precipitate, dissolving with water, and concentrating under reduced pressure.

6. The preparation method according to claim 4, wherein the resin purification process comprises adding the concentrated solution into the resin, washing the resin to be transparent by deionized water, eluting the resin by ethanol water solution, and collecting the eluate. The resin is AB-8 or D101 macroporous resin, the eluent concentration is 30-85% ethanol solution, and then the pressure reduction concentration is carried out.

7. The preparation method according to claim 1, wherein the stabilizer in step three is one or more of glycerol, butanediol and propylene glycol, the membrane in step three is a microfiltration membrane, the pore diameter is 0.2-1 μm, and the adding proportion of the activated carbon in step three is 0-1.5%.

8. A Pandanus communis leaf extract, characterized in that the Pandanus communis leaf extract is prepared by the preparation method according to any one of claims 1-7.

9. The application of the Pandanus communis leaf extract prepared by the preparation method according to any one of claims 1-7 is characterized in that the Pandanus communis leaf extract is applied to cosmetics, and the cosmetics are skin external preparations with moisturizing and relieving effects, which are prepared by taking the Pandanus communis leaf extract as an active component and adding conventional pharmaceutical or cosmetic auxiliary materials or auxiliary components.

10. The use of claim 9, wherein the moisturizing and soothing cosmetic is one or more of a lotion, a serum, a cream, a mask, and a gel.