CN112972524A

CN112972524A - Application of schisandra chinensis oil in inhibition of sebaceous gland activity

Info

Publication number: CN112972524A
Application number: CN202110368972.6A
Authority: CN
Inventors: 唐丹丹; 杜志云; 周渭; 刘冠廷; 王珊珊
Original assignee: Guangdong University of Technology
Current assignee: Guangdong University of Technology
Priority date: 2021-04-06
Filing date: 2021-04-06
Publication date: 2021-06-18
Anticipated expiration: 2041-04-06
Also published as: CN112972524B

Abstract

The application belongs to the technical field of biological medicines. The application provides application of schisandra chinensis oil in inhibiting sebaceous gland activity. Experimental results show that the schisandra chinensis oil can obviously inhibit the proliferation of sebaceous gland cells, inhibit the lipid synthesis of the sebaceous gland cells and reduce the expression of sebaceous gland cell receptors at the cellular level, and realizes the effects of inhibiting the growth of sebaceous gland plaques and inhibiting the sebum secretion, so that the schisandra chinensis oil has the effects of inhibiting the sebaceous gland activity, controlling oil, regulating the water-oil balance of the face and the like, can be applied to medicaments for treating acne or oil-controlling cosmetics, and has the advantages of safety, high efficiency and low cost.

Description

Application of schisandra chinensis oil in inhibition of sebaceous gland activity

Technical Field

The application belongs to the technical field of biomedicine, and particularly relates to application of schisandra chinensis oil in inhibition of sebaceous gland activity.

Background

Schisandra chinensis (Schisandra chinensis) is a dry mature fruit of Schisandra chinensis (Schisandra chinensis) belonging to Magnoliaceae, and is listed as the top grade in Shen nong Ben Cao Jing. The book of Xin Xiu Ben Cao records that the five flavors of skin and meat are sweet and sour, and bitter in the kernel, all have salty taste, so it is named as Wu Wei Zi. The schisandra fruit has the functions of nourishing and strengthening and has high medicinal and edible values. Chemical research shows that the schisandra chinensis mainly contains lignans, volatile oils, flavonoids, terpenes, organic acids and other active ingredients, and pharmacological research shows that the schisandra chinensis has the effects of protecting liver, tranquilizing and hypnotizing, reducing blood sugar, resisting oxidation, enhancing immunity, resisting cancer, resisting AIDS virus and the like. However, the schisandra chinensis has not been reported to inhibit the sebaceous gland activity.

The main components of sebum include triglyceride, wax ester, squalene, cholesterol ester and cholesterol. Sebaceous glands are a kind of glandular full-plasma secretory tissue distributed in the skin of the whole body except palms and soles, and have the highest density on the scalp and face. The normal lipid on the skin surface can play a barrier role, can moisten the skin, resist infection and maintain the bacteria state balance on the skin surface of a human body. Excessive sebum secretion from sebaceous glands leads to excessive skin oil production and aesthetic problems, and may also promote the development of acne or other diseases, which seriously affect the mental health and quality of life of patients.

The breakdown of keratinocytes and the sebum secreted by the sebaceous glands constitute the major source of lipids on the skin surface. Among them, the breakdown of keratinocytes produces only a small fraction of the lipids, and the excessive sebum secretion from the sebaceous glands is the main cause of the greasiness of the skin. The sebaceous gland function is regulated by a plurality of factors, and androgen is taken as a primary factor influencing sebum secretion and mainly regulates the differentiation and proliferation of the sebaceous gland and the synthesis and secretion of sebum. In addition, sebaceous gland function is also regulated by enzyme systems such as Peroxisome Proliferator Activated Receptors (PPARs), neuropeptides, antimicrobial peptides, inflammatory cytokines, and the like.

At present, western medicines are mainly used for effectively inhibiting sebaceous gland secretion, and have the defects of multiple side effects and strong dependence, so that a safe and effective substitute component is urgently needed.

Disclosure of Invention

In view of the above, the present application provides an application of schizandra chinensis oil in inhibiting sebaceous gland activity, wherein the schizandra chinensis oil can significantly inhibit growth of sebaceous gland plaques and sebum secretion at an animal level.

The specific technical scheme of the application is as follows:

in a first aspect, the application provides the use of schizandra chinensis oil for inhibiting sebaceous gland activity.

Preferably, the inhibition of sebaceous gland activity is specifically inhibition of sebaceous gland cell proliferation.

Preferably, the inhibition of the sebaceous gland activity is specifically inhibition of lipid synthesis by sebaceous gland cells.

Preferably, the inhibition of sebaceous gland activity is specifically the down-regulation of the expression of androgen receptor mRNA of sebaceous gland cells.

Preferably, the schisandra chinensis oil is volatile oil obtained by extracting schisandra chinensis fruits.

Preferably, the step of extracting comprises: crushing fructus schizandrae, sieving with a 14-60-mesh sieve, adding the crushed powder into an aqueous solution of NaCl according to the liquid-material mass ratio of 4-8 times, soaking for 2-4 hours, then carrying out water distillation for 2-5 hours, and collecting the upper essential oil.

More preferably, the step of extracting comprises: pulverizing fructus Schisandrae, sieving with 40 mesh sieve, soaking the pulverized powder in 6 times of distilled water containing 6% NaCl for 3 hr, distilling with water for 3 hr, collecting upper layer essential oil, and adding anhydrous NaSO₄Drying and filtering to obtain the product.

In the application, animal level experiments show that the schisandra chinensis oil can obviously inhibit the proliferation of sebaceous gland cells, inhibit the synthesis of lipid by the sebaceous gland cells and reduce the expression of a sebaceous gland cell receptor at a cell level, so that the effects of inhibiting the growth of sebaceous gland plaques and inhibiting the secretion of sebum are realized, and the schisandra chinensis oil has good development potential.

In a second aspect, the application provides the use of schizandra chinensis oil in the preparation of a medicament for treating acne.

Preferably, the medicament is in the form of emulsion or paste.

In a third aspect, the application provides application of schisandra chinensis oil in preparing oil-control cosmetics.

Preferably, the cosmetic is in the form of water, lotion, essence or facial mask.

In the application, the schisandra chinensis oil can obviously inhibit the growth of sebaceous gland plaques and sebum secretion at the animal level, so that the schisandra chinensis oil has the effects of inhibiting the activity of sebaceous glands, controlling oil, regulating the water-oil balance of the face and the like, can be applied to medicaments for treating acne or oil-control cosmetics, and has the advantages of safety, high efficiency and low cost.

In summary, the application provides an application of schisandra chinensis oil in inhibition of sebaceous gland activity. Experimental results show that the schisandra chinensis oil can obviously inhibit the proliferation of sebaceous gland cells, inhibit the lipid synthesis of the sebaceous gland cells and reduce the expression of sebaceous gland cell receptors at the cellular level, and realizes the effects of inhibiting the growth of sebaceous gland plaques and inhibiting the sebum secretion, so that the schisandra chinensis oil has the effects of inhibiting the sebaceous gland activity, controlling oil, regulating the water-oil balance of the face and the like, can be applied to medicaments for treating acne or oil-controlling cosmetics, and has the advantages of safety, high efficiency and low cost.

Drawings

In order to more clearly illustrate the embodiments of the present application or the technical solutions in the prior art, the drawings needed to be used in the description of the embodiments or the prior art will be briefly introduced below, and it is obvious that the drawings in the following description are only some embodiments of the present application, and it is obvious for those skilled in the art that other drawings can be obtained according to the drawings without inventive exercise.

FIG. 1 is the sebum secretion amount of sebaceous gland plaques of mice with golden yellow color at 0, 10, 20, 30d before and after administration of schizandra chinensis oil (left side) in example 5;

FIG. 2 is the sebum secretion amount of sebaceous gland plaques of mice with golden yellow color at 0, 10, 20, 30d before and after administration of the schisandra fruit oil in example 5 (right side).

Detailed Description

In order to make the objects, features and advantages of the present application more obvious and understandable, the technical solutions in the embodiments of the present application are clearly and completely described, and it is obvious that the embodiments described below are only a part of the embodiments of the present application, and not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present application.

The starting materials and reagents used in the examples of the present application are commercially available, and the experimental procedures used are routine in the art unless otherwise specified.

The golden hamster of the embodiment of the application is purchased from Guangdong province medical animal experiment center, SZ95 cells are purchased from Hongbao Biotechnology Limited, Shenzhen, propylene glycol is purchased from Tianjin Mao chemical reagent factory, chloral hydrate is purchased from Sigma-Aldrich, DMSO is purchased from Tianjin Mao chemical reagent factory, Nile Red fluorescent dye solution is purchased from Shanghai Huling Biotech Limited, reverse transcription kit is purchased from Promega, Trizol is purchased from Gibco, dNTP is purchased from Guangzhou Dongshan Biotech Limited, MgCl₂Purchased from Shanghai Aladdin Biotechnology Ltd.

Example 1 extraction of Schisandra chinensis oil

Pulverizing fructus Schisandrae, sieving with 40 mesh sieve, soaking the pulverized powder in 6 times of distilled water containing 6% NaCl for 3 hr, distilling with water for 3 hr, collecting upper layer essential oil, and adding anhydrous NaSO₄Drying and filtering to obtain the product.

Example 2 Effect of Schisandra chinensis oil on the proliferation of sebaceous gland cells

(1) Culturing human sebaceous gland cells (SZ95 cells), and digesting SZ95 cells by trypsin to prepare SZ95 cell suspension when the cells grow and fuse to reach 80%;

(2) inoculating the SZ95 cell suspension to a 96-well culture plate, wherein each well comprises 4000 cells, culturing for 48h, removing the supernatant, washing for 2 times by using PBS solution, respectively adding the culture medium containing 0.5, 5, 50 and 500 mug/mL of the schisandra chinensis oil prepared in the example 1, continuously culturing for 48h, removing the supernatant, and washing for 2 times by using the PBS solution;

(3) 0.5mg/mL MTT 100u1 was added, after 4h incubation, MTT was discarded, 150. mu.L DMMS 0 was added to each well and incubated for 10 min. The absorbance was measured using a microplate reader at a wavelength of 490 nm.

3 parallel wells were set for each effect concentration, and medium without schizandra oil was added as a blank group, medium without addition of SZ95 cells and without schizandra oil was added as a blank control group, and medium at each effect concentration was added without addition of SZ95 cells as a zero-adjustment group. The results of the effect of schizandra chinensis oil on the proliferation rate of SZ95 cells in example 2 of the present application are shown in table 1, wherein the proliferation rate of SZ95 cells in the blank group is blank OD-blank OD, and the proliferation rate of SZ95 cells in the experimental group is (experimental OD-zero-adjustment OD)/(blank OD-blank OD).

TABLE 1 Effect of Schisandra chinensis oil on the proliferation Rate of SZ95 cells

Group of	Cell proliferation Rate (%) of SZ95
		Blank group	100±4.8860
0.5. mu.g/ml test group	96.67±4.0787
		5 μ g/ml test group	86.10±4.0398*
50 μ g/ml experimental group	71.09±7.6650***
		500 μ g/ml experimental group	46.71±8.3923***

Note: p < 0.05, P < 0.001, compared to blank

As can be seen from Table 1, 0.5. mu.g/ml schizandra chinensis oil has no significant inhibition effect on the proliferation of sebaceous gland cells (P is more than 0.05); the schisandra chinensis oil with the concentration of 5 mu g/ml has obvious inhibition effect on the proliferation of sebaceous gland cells (P is less than 0.01); 50. the Schisandra chinensis oil with the concentration of 500 mu g/ml has stronger inhibition effect on the proliferation of sebaceous gland cells (P is less than 0.001).

Example 3 Effect of Schisandra chinensis oil on sebaceous gland cell lipid Synthesis

(1) Culturing SZ95 cells, and digesting the SZ95 cells by trypsin to prepare SZ95 cell suspension when the cells grow and fuse to reach 80%;

(2) SZ95 cells were seeded in 6-well plates at 2X 10 per well⁵After further culturing for 48 hours, medium containing 0.5, 5, 50 and 500. mu.g/mL of the schisandra oil prepared in example 1 was added. Continuing culturing for 48h, adding trypsin for digestion, stopping digestion with a culture medium containing 10% fetal calf serum, and washing with PBS solution for 2 times to prepare single cell suspension;

(3) nile red fluorescent dye solution was added to a final concentration of 100 ng/mL. Incubate at room temperature for 15min, and filter with 300 mesh nylon filter. The fluorescence intensity of 10000 cells per group was measured by flow cytometry. Finally, the mean fluorescence intensity (excitation wavelength 485nm, emission wavelength 565nm) of each cell was calculated.

3 parallel wells were set for each effect concentration, and medium without schizandra oil was added as a blank. The results of the effect of schizandra chinensis oil on the synthesis of SZ95 cell lipids in example 3 of the present application are shown in table 2, wherein the inhibition ratio of the experimental group is (1-experimental group fluorescence unit/blank group fluorescence unit) × 100%.

TABLE 2 Effect of Schisandra chinensis oil on SZ95 cell lipid synthesis

Group of	Mean fluorescence unit	Inhibition ratio (%)
			Blank group	649.33±46.0145	-
0.5. mu.g/ml test group	629.67±28.1484	3.03
			5 μ g/ml test group	574.90±32.4553	11.46
50 μ g/ml experimental group	458.21±52.0152***	29.43
			500 μ g/ml experimental group	497.50±44.9246**	23.38

Note: p < 0.01, P < 0.001, compared to blank

As can be seen from Table 2, 0.5 and 5. mu.g/ml schizandra chinensis oil had insignificant inhibitory effects on sebaceous gland cell lipid synthesis (P > 0.05); the inhibitory effect of 50 and 500 mu g/ml schisandra fruit oil on the proliferation of sebaceous gland cells is obvious (P is less than 0.01 and P is less than 0.001 respectively).

Example 4 Effect of Schisandra chinensis oil on the expression of Androgen Receptor (AR) mRNA of sebaceous gland cells

(1) Culturing SZ95 cells, and digesting SZ95 cells by trypsin to prepare SZ95 cell suspension when the cells grow and fuse to 80%;

(2) SZ95 cells were seeded in 6-well plates at 2X 10 per well⁵Respectively adding into culture medium after culturing for 96h0.5, 5, 50 and 500. mu.g/mL of the medium of Schisandra chinensis oil prepared in example 1. After further incubation for 24h, Trizol was added and total RNA was extracted from SZ95 cells according to the instructions. The cDNA was synthesized by reverse transcription following the reverse transcription kit instructions, followed by in-line amplification of AR and β -actin (β -actin). The primer sequence is as follows: the AR sense strand: 5'-GAAGACCTGCCTGATCTGTG-3', antisense strand: 5 '-AAGCCTCTCCITCCTCCTGT-3' (product length 269 bp); internal reference beta-actin sense strand: 5'-AGAGATGGCCACGGCTGCTT-3', antisense strand: 5'-ATTTGCGGTGGACGATGGAG-3' (product length 445 bp). The reaction system is 25 μ L: wherein PCR 10 Xbuffer 2.5. mu.L, 10mmol/L dNTP mixture 0.5. mu.L, 25mol/L MgCl₂mu.L of 2. mu.L of each of the sense and antisense strands of AR (20pmoL/L), 0.2. mu.L of each of the sense and antisense strands of beta-actin (20pmoL/L), 1.25U of Taq DNA polymerase, 10. mu.L of cDNA, and finally deionized water to a total volume of 25. mu.L. The reaction conditions are as follows: 30s at 94 ℃, 45s at 60 ℃ and 30s at 72 ℃ for 40 cycles. The RT-PCR products were electrophoresed on a 20% agarose gel and stained with 0.5% ethidium bromide. And analyzing the scanning result by adopting a Shanghai energy gel imaging analysis system.

3 parallel wells were set for each effect concentration, and medium without schizandra oil was added as a blank. The absorbance value takes the product of the intensity and the net area as a quantitative standard, and the ratio of the absorbance value of the specific product and the absorbance value of the beta-actin as a semi-quantitative value. The results of the effect of schizandra chinensis oil on the expression of the androgen receptor mRNA of SZ95 cells in example 4 of the present application are shown in table 3, wherein the expression level is experimental group (blank group) absorbance/β -actin absorbance.

TABLE 3 Effect of Schisandra chinensis oil on the expression of Androgen Receptor (AR) mRNA of SZ95 cells

Group of	Amount of expression
		Blank group	0.632±0.0215
0.5. mu.g/ml test group	0.642±0.0210
		5 μ g/ml test group	0.561±0.0331*
50 μ g/ml experimental group	0.464±0.0319****
		500 μ g/ml experimental group	0.532±0.0251**

Note: p < 0.05, P < 0.01, P < 0.0001 compared to blank

As can be seen from Table 3, 0.5. mu.g/ml schizandra oil failed to inhibit the expression of the mRNA of androgen receptor of sebaceous gland cells (P > 0.05); 5. the inhibitory effect of 50 and 500 mu g/ml schisandra fruit oil on the mRNA expression of androgen receptor of sebaceous gland cells is obvious (P is less than 0.05, P is less than 0.0001 and P is less than 0.01 respectively).

Example 5 inhibitory Effect of Schisandra chinensis oil on sebum secretion

(1) Respectively dissolving 25mg and 100mg of the schisandra chinensis oil prepared in the example 1 in propylene glycol, wherein the total volume of the solution is 10mL, and preparing 2.5mg/mL and 10mg/mL schisandra chinensis oil solutions;

(2) 18 male golden yellow hamster are selected, aged for 6 weeks, and the weight of the male golden yellow hamster is 100g +/-20 g, and the male golden yellow hamster is adaptively raised for 3 days before the experiment. The golden hamster was divided into three groups, a blank group (control) was not administered, a low concentration test group (L-SC) was coated with 2.5mg/mL of a schisandra oil solution, and a high concentration test group (H-SC) was coated with 10mg/mL of a schisandra oil solution. 24 hours before application on the first day, the lateral flank hair of each group of golden hamster was shaved and depilatory cream was used to remove hair again, fully exposing the sebaceous glands. Spreading oleum Schisandrae chinensis solution with different concentrations on sebaceous glands of golden hamster 20 μ L each side once a day for 30 days;

(3) after anesthetizing golden hamster by intraperitoneal injection of chloral hydrate at 0, 10, 20 and 30d, respectively, the maximum transverse Diameter (DT) and the maximum longitudinal Diameter (DL) of sebaceous gland plaques were measured with a vernier caliper, and the change of each group of plaques pi × DT × DL was calculated. After anesthetizing the golden hamster by intraperitoneal injection of chloral hydrate for 0, 10, 20 and 30 days, the skin oil and fat were measured by a CK skin tester.

The data are expressed as X + -S, and the difference between groups is compared by t-test. In example 5 of the present application, the comparison of sizes of sebaceous glands of mice with golden yellow color before and after administration of schizandra chinensis oil is shown in table 4 (left side), and the comparison of sizes of sebaceous glands of mice with golden yellow color before and after administration of schizandra chinensis oil is shown in table 5 (right side).

TABLE 4 comparison of size of sebaceous gland plaques of golden hamster before and after administration of Schisandra chinensis oil (left side) (unit: mm)²)

Group of	Number of examples	Area before application	Area	30 days after application
					Blank group	6	36.82±2.7563	56.00±1.8890
Low concentration test group	6	36.25±1.4849	48.00±3.2239**
				High concentration test group	6	35.85±3.7546	46.03±4.9922**

Note: p < 0.05, P < 0.01, compared to blank

TABLE 5 comparison of size of sebaceous gland plaques of golden hamster before and after administration of Schisandra chinensis oil (right side) (unit: mm)²)

Group of	Number of examples	Area before application	Area	30 days after application
					Blank group	6	36.54±1.0738	57.22±4.2068
Low concentration test group	6	36.45±3.3789	53.87±7.8266
				High concentration test group	6	36.50±4.0583	48.41±4.9341*

Note: p < 0.05, P < 0.01, compared to blank

As can be seen from tables 4 and 5, the areas of the groups before administration were comparable without significant difference (P > 0.05). The sebaceous gland plaques of the golden hamster in the blank group, the low-concentration experimental group and the high-concentration experimental group are increased compared with those before treatment, but the sebaceous gland plaques in the low-concentration experimental group and the high-concentration experimental group are reduced compared with those in the blank group, and the inhibition effect of the high-concentration schisandra fruit oil on the sebaceous gland plaque increase is remarkably different (P is less than 0.01).

Sebum secretion amounts of sebaceous gland plaques of golden hamster at 0, 10, 20, 30d before and after administration of schizandra chinensis oil in example 5 of the present application are shown in fig. 1 (left side) and fig. 2 (right side). Fig. 1 and 2 show that sebum secretion from sebaceous plaques on both left and right sides of golden yellow hamster is significantly reduced in the low and high concentration experimental group compared to the blank group after 30 days of administration. Wherein, the high-concentration schisandra chinensis oil has better inhibition effect on sebum secretion of golden hamster.

The experimental results show that the schisandra chinensis oil can inhibit the proliferation of sebaceous gland cells, inhibit the synthesis of lipid by the sebaceous gland cells and reduce the expression of androgen receptor mRNA of the sebaceous gland cells, thereby realizing the effects of inhibiting the growth of sebaceous gland plaques and inhibiting the secretion of sebum and having good development potential.

The above embodiments are only used for illustrating the technical solutions of the present application, and not for limiting the same; although the present application has been described in detail with reference to the foregoing embodiments, it should be understood by those of ordinary skill in the art that: the technical solutions described in the foregoing embodiments may still be modified, or some technical features may be equivalently replaced; and such modifications or substitutions do not depart from the spirit and scope of the corresponding technical solutions in the embodiments of the present application.

Claims

1. Use of Schisandra chinensis oil for inhibiting sebaceous gland activity is provided.

2. Use according to claim 1, wherein the inhibition of sebaceous gland activity is in particular inhibition of sebaceous gland cell proliferation.

3. Use according to claim 1, wherein the inhibition of sebaceous gland activity is in particular inhibition of lipid synthesis by sebaceous gland cells.

4. The use according to claim 1, wherein said inhibition of sebaceous gland activity is in particular down-regulation of the expression of the androgen receptor mRNA of sebaceous gland cells.

5. The use of claim 1, wherein the schizandra chinensis oil is an essential oil extracted from the fruit of schizandra chinensis.

6. The use according to claim 5, wherein the step of extracting comprises: crushing fructus schizandrae, sieving with a 14-60-mesh sieve, adding the crushed powder into an aqueous solution of NaCl according to the liquid-material mass ratio of 4-8 times, soaking for 2-4 hours, then carrying out water distillation for 2-5 hours, and collecting the upper essential oil.

7. Application of Schisandra chinensis oil in preparing medicine for treating acne is provided.

8. The use according to claim 7, wherein the medicament is in the form of an emulsion or an ointment.

9. Application of Schisandra chinensis oil in preparing oil-controlling cosmetics is provided.

10. The use according to claim 9, characterized in that the cosmetic is in the form of water, a lotion, a serum or a mask.