COMPOSITION HAVING FUNCTIONS OF MOISTURIZING, REPAIRING AND WHITENING AND USE THEREOF
20240033315 ยท 2024-02-01
Inventors
- Kaiye Yang (Jiangmen, CN)
- Guangrong Liu (Jiangmen, CN)
- Meiling Tai (Jiangmen, CN)
- Jiawen Chen (Jiangmen, CN)
- Shuang Gao (Jiangmen, CN)
- Lusheng Wang (Jiangmen, CN)
- Zhenlin Hu (Jiangmen, CN)
Cpc classification
A61P29/00
HUMAN NECESSITIES
International classification
A61P29/00
HUMAN NECESSITIES
Abstract
Disclosed is a composition having functions of moisturizing, repairing and whitening and use thereof. The composition comprises Dendrobium candidum polysaccharide, a lotus extract and a Glycyrrhiza glabra extract. The lotus extract and the Glycyrrhiza glabra extract can inhibit pigmentation by inhibiting synthesis of melanin, inhibiting transfer of melanin globule, resisting oxidation and improving skin microcirculation, so as to exert the effect of special whitening. Through the synergistic synergy of the three components, and the compounded composition can reduce synthesis of melanin of skin's melanophores, promote expression of keratinocyte cell hydration-related factors, inhibit expression of inflammation-related factors, promote the ability of lipid synthesis and exert multiple effects of whitening, enhancement in moisturizing of the skin, alleviation in inflammation and discomfort of the skin, and improvement in barrier function of epidermis. In addition, the compounded composition may be applied to preparation of skin care products or skin formulation.
Claims
1. A composition, comprising Dendrobium candidum polysaccharide, a lotus extract and a Glycyrrhiza glabra extract.
2. The composition according to claim 1, wherein a mass ratio of the Dendrobium candidum polysaccharide to the lotus extract to the Glycyrrhiza glabra extract is (1-5):(1-3):(1-3).
3. The composition according to claim 1, wherein the mass ratio of the Dendrobium candidum polysaccharide to the lotus extract to the Glycyrrhiza glabra extract is (1-3):(1-3):(1-3).
4. The composition according to claim 1, wherein the mass ratio of the Dendrobium candidum polysaccharide to the lotus extract to the Glycyrrhiza glabra extract is 3:(1-3):(1-3).
5. The composition according to claim 1, wherein the Dendrobium candidum polysaccharide, the lotus extract and the Glycyrrhiza glabra extract are extracted from Dendrobium candidum, lotus and Glycyrrhiza glabra, as raw materials, respectively.
6. A product comprising the composition according to claim 1.
7. A product comprising the composition according to claim 2.
8. A product comprising the composition according to claim 3.
9. A product comprising the composition according to claim 4.
10. A product comprising the composition according to claim 5.
11. The product according to claim 6, wherein the product has at least one function of (a)-(j): (a) reducing a content of melanin; (b) promoting expression of aquaporin AQP3; (c) inhibiting production of reactive oxygen species (ROS); (d) inhibiting expression of inflammatory factors; (e) promoting expression of lipid synthesis-related factors; (f) promoting lipid synthesis; (g) whitening; (h) moisturizing; (i) resisting oxidation; and (j) resisting inflammation.
12. The product according to claim 7, wherein the product has at least one function of (a)-(j): (a) reducing a content of melanin; (b) promoting expression of aquaporin AQP3; (c) inhibiting production of reactive oxygen species (ROS); (d) inhibiting expression of inflammatory factors; (e) promoting expression of lipid synthesis-related factors; (f) promoting lipid synthesis; (g) whitening; (h) moisturizing; (i) resisting oxidation; and (j) resisting inflammation.
13. The product according to claim 8, wherein the product has at least one function of (a)-(j): (a) reducing a content of melanin; (b) promoting expression of aquaporin AQP3; (c) inhibiting production of reactive oxygen species (ROS); (d) inhibiting expression of inflammatory factors; (e) promoting expression of lipid synthesis-related factors; (f) promoting lipid synthesis; (g) whitening; (h) moisturizing; (i) resisting oxidation; and (j) resisting inflammation.
14. The product according to claim 9, wherein the product has at least one function of (a)-(j): (a) reducing a content of melanin; (b) promoting expression of aquaporin AQP3; (c) inhibiting production of reactive oxygen species (ROS); (d) inhibiting expression of inflammatory factors; (e) promoting expression of lipid synthesis-related factors; (f) promoting lipid synthesis; (g) whitening; (h) moisturizing; (i) resisting oxidation; and (j) resisting inflammation.
15. The product according to claim 10, wherein the product has at least one function of (a)-(j): (a) reducing a content of melanin; (b) promoting expression of aquaporin AQP3; (c) inhibiting production of reactive oxygen species (ROS); (d) inhibiting expression of inflammatory factors; (e) promoting expression of lipid synthesis-related factors; (f) promoting lipid synthesis; (g) whitening; (h) moisturizing; (i) resisting oxidation; and (j) resisting inflammation.
16. The product according to claim 6, wherein the product is a cosmetic or a drug.
17. The product according to claim 7, wherein the product is a cosmetic or a drug.
18. The product according to claim 8, wherein the product is a cosmetic or a drug.
19. The product according to claim 9, wherein the product is a cosmetic or a drug.
20. The product according to claim 16, wherein the cosmetic further comprises a excipient, wherein the excipient is at least one selected from the group consisting of a preservative, a chelating agent, a flavor, a moisturizer, a colorant, an emulsifier, an antioxidant and a skin conditioner.
Description
BRIEF DESCRIPTION OF DRAWINGS
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DETAILED DESCRIPTION
[0089] Next, the concept and technical effects of the present disclosure will be clearly and completely described in combination with embodiments to sufficiently understand the objectives, features and effects of the present disclosure. Obviously, the described embodiments are only some embodiments of the present disclosure, but not all the embodiments. Based on the embodiments of the present disclosure, other embodiments obtained by those skilled in the art without creative efforts fall into the protection scope of the present disclosure.
[0090] The lotus extract: the lotus extract is purchased from Ashland Group in the United States with a production batch number of 001684RA00, and the extraction process is consistent with the extraction technology described in the US patent document U.S. Pat. No. 8,668,939B2.
Example 1
[0091] Preparation of Dendrobium candidum Polysaccharide [0092] (1) 1 kg of Dendrobium candidum was crushed into crude particles and extracted twice with water, wherein a first solid-liquid ratio (g/mL) was 1:40, and the extraction was conducted for 2 h at 100 C., and a second solid-liquid ratio (g/mL) was 1:10, and the extraction was conducted for 1.5 h at 100 C. The extracting solutions were respectively filtrated by 100 mesh, and the obtained filtrate were combined for tubular centrifugation. The centrifuged extracting solution was concentrated to 1/10 volume of the filtrate at 70-80 C.; absolute ethanol was added to the concentrated solution to adjust a concentration of ethanol to 80%; alcohol precipitation was conducted for 2 h; 300-mesh filtration was performed to collect precipitates; and then the ethanol was evaporated. [0093] (2) The precipitates were added into water for re-dissolution (a concentration was 0.5%) to obtain a polysaccharide solution; the polysaccharide solution was evenly mixed with a sevage reagent [chloroform:n-butanol=4:1 (v:v)] in a ratio of 5:1 (v:v); the above mixed solution was sufficiently shaken for 20 min for standing until obvious layering appeared; the sevage in the lower layer and proteins in the middle layer were released, and such operations were repeated for about 10 times; and the polysaccharide solution was collected for later use. [0094] (3) First, macroporous resin AB-8 was activated by washing with distilled water, then soaking with 3-5% HCl solution and 3-5% NaOH solution for 24 h in sequence; the activated macroporous resin AB-8 was washed with the distilled water to be neutral; then the washing solution was removed by suction filtration. An appropriate amount of activated macroporous resin was added in to the polysaccharide solution, placed in a shaker for shaking for 4 h, and subjected to suction filtration to obtain a polysaccharide solution without pigment. The polysaccharide solution was collected, concentrated to 1/10 volume of the filtrate at 60-70 C. and underwent freeze drying for 48 h at 80 C. to obtain polysaccharide freeze-dried powder. [0095] (4) The polysaccharide freeze-dried powder (20 mg) was weighed and dissolved into 1 mL of distilled water; and a sample was centrifuged for 10 min at 6000 r/min after being completely dissolved into water. The supernatant was taken and loaded. The sample was slowly added to a G100 sephadex column with a dropper, wherein the sample must be slowly added dropwise along the wall of the chromatographic column so as to avoid the situation that uneven gel surface is generated to affect the separation effect. The sample was eluted by using the distilled water as an eluent at a flow rate of 1 mL.Math.min.sup.1, the eluted solution was collected in 6 min/tube, 50 L of solution was taken from each tube until the loading solution was completely collected; the loading solution was concentrated to 1/10 volume of filtrate at 60-70 C.; and then the concentrated solution was subjected to freeze drying for 48 h at 80 to obtain the Dendrobium candidum polysaccharide.
[0096] Detection on Molecular Weight of Dendrobium candidum Polysaccharide [0097] (1) 10 mg of the Dendrobium candidum polysaccharide sample prepared in the above steps were precisely weighed and dissolved into 10 mL of 0.1 M NaNO.sub.3 aqueous solution to prepare an aqueous solution with a final concentration of 1 mg/mL, and the obtained aqueous solution was filtrated through a filter with a pore size of 0.22 m for later use. (2) The filtered Dendrobium candidum polysaccharide solution sample was analyzed by a gel chromatography-differential-multi-angle laser light scattering system, with a injection volume of 100 L, where the chromatographic conditions were as follows: gel size exclusion chromatography columns Ohpak SB-805 HQ (3008 m), Ohpak SB-804 HQ (3008 mm), Ohpak SB-803 HQ (3008 mm) connected in series, a column temperature of 45 C., a mobile phase A (0.1M NaNO.sub.3), a flow rate of 0.4 ml/min, an elution gradient of isocratic elution for 100 min. By detection, the molecular weight of the Dendrobium candidum polysaccharide was shown as 0.3-27 kDa.
[0098] The preparation of the Glycyrrhiza glabra extract may be seen in Chinese patent CN108176079A in details.
[0099] Preparation of a Dendrobium candidum polysaccharide aqueous solution: every 15 mg of dried Dendrobium candidum polysaccharide obtained was dissolved into 1 mL of sterilized deionized water, and the obtained solution was filtered through a 0.22 m microporous membrane to prepare a 15 mg/mL Dendrobium candidum polysaccharide aqueous solution.
[0100] Preparation of a lotus extract aqueous solution: every 15 mg of lotus extract obtained was dissolved into 1 mL of sterilized deionized water, and the obtained solution was filtered through a 0.22 m microporous membrane to prepare a 15 mg/mL lotus extract aqueous solution.
[0101] Preparation of a Glycyrrhiza glabra extract aqueous solution: every 15 mg of dried Glycyrrhiza glabra extract obtained was dissolved into 1 mL of sterilized deionized water, and the obtained solution was filtered through a 0.22 m microporous membrane to obtain a 15 mg/mL Glycyrrhiza glabra extract aqueous solution.
Example 2
[0102] Groups:
[0103] Any treatment was not conducted for a blank group.
[0104] UVB irradiation was conducted only in an ultraviolet irradiator for a UVB group.
[0105] Only 10 L of CaCl.sub.2 aqueous solution (with a concentration of 2 mol/L) was added for a Ca.sup.2+ group.
[0106] Only 2 L of 15 mg/mL Dendrobium candidum polysaccharide aqueous solution was added for administration for a Dendrobium candidum alone group.
[0107] Only 2 L of 15 mg/mL lotus extract aqueous solution was added for administration for a lotus extract alone group.
[0108] Only 2 L of 15 mg/mL Glycyrrhiza glabra aqueous solution was added for administration for a Glycyrrhiza glabra alone group.
[0109] The complexes A1, A2, A3, A4 and A5 were prepared by simply mixing the following three components in the following mass ratios. Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=1:1:1 (Complex A1), 2:2:3 (Complex A2), 2:3:2 (Complex A3), 3:2:2 (Complex A4), and 5:1:1 (Complex A5).
[0110] Complex A1 group: 5 mg of dried Dendrobium candidum polysaccharide, 5 mg of lotus extract and 5 mg of Glycyrrhiza glabra extract were weighted, mixed, dissolved into 1 mL of sterilized deionized water, and filtered via a 0.22 m microporous membrane to obtain a mixture aqueous solution. 2 L of mixture aqueous solution was taken for administration.
[0111] Complex A2 group: 4 mg of dried Dendrobium candidum polysaccharide, 4 mg of lotus extract and 6 mg of Glycyrrhiza glabra extract were weighted, mixed, dissolved into 1 mL of sterilized deionized water, and filtered via a 0.22 m microporous membrane to obtain a mixture aqueous solution. 2 L of mixture aqueous solution was taken for administration.
[0112] Complex A3 group: 4 mg of dried Dendrobium candidum polysaccharide, 6 mg of lotus extract and 4 mg of Glycyrrhiza glabra extract were weighted, mixed, dissolved into 1 mL of sterilized deionized water, and filtered via a 0.22 m microporous membrane to obtain a mixture aqueous solution. 2 L of mixture aqueous solution was taken for administration.
[0113] Complex A4 group: 6 mg of dried Dendrobium candidum polysaccharide, 4 mg of lotus extract and 4 mg of Glycyrrhiza glabra extract were weighted, mixed, dissolved into 1 mL of sterilized deionized water, and filtered via a 0.22 m microporous membrane to obtain a mixture aqueous solution. 2 L of mixture aqueous solution was taken for administration.
[0114] Complex A5 group: 10 mg of dried Dendrobium candidum polysaccharide, 2 mg of lotus extract and 2 mg of Glycyrrhiza glabra extract were weighted, mixed, dissolved into 1 mL of sterilized deionized water, and filtered via a 0.22 m microporous membrane to obtain a mixture aqueous solution. 2 L of mixture aqueous solution was taken for administration.
[0115] The complexes B1, B2, B3, B4 and B5 were prepared by simply mixing the following three components in the following mass ratios. Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=3:2:1 (Complex B1), 3:1:2 (Complex B2), 3:2:2 (Complex B3), 3:3:2 (Complex B4), and 3:2:3 (Complex B5).
[0116] Complex B1 group: 7.5 mg of dried Dendrobium candidum polysaccharide, 5 mg of lotus extract and 2.5 mg of Glycyrrhiza glabra extract were weighted, mixed, dissolved into 1 mL of sterilized deionized water, and filtered via a 0.22 m microporous membrane to obtain a mixture aqueous solution. 2 L of mixture aqueous solution was taken for administration.
[0117] Complex B2 group: 7.5 mg of dried Dendrobium candidum polysaccharide, 2.5 mg of lotus extract and 5 mg of Glycyrrhiza glabra extract were weighted, mixed, dissolved into 1 mL of sterilized deionized water, and filtered via a 0.22 m microporous membrane to obtain a mixture aqueous solution. 2 L of mixture aqueous solution was taken for administration.
[0118] Complex B3 group: 6 mg of dried Dendrobium candidum polysaccharide, 4 mg of lotus extract and 4 mg of Glycyrrhiza glabra extract were weighted, mixed, dissolved into 1 mL of sterilized deionized water, filtered via a 0.22 m microporous membrane to obtain a mixture aqueous solution. 2 L of mixture aqueous solution was taken for administration.
[0119] Complex B4 group: 5.25 mg of dried Dendrobium candidum polysaccharide, 5.25 mg of lotus extract and 3.5 mg of Glycyrrhiza glabra extract were weighted, mixed, dissolved into 1 mL of sterilized deionized water, and filtered via a 0.22 m microporous membrane to obtain a mixture aqueous solution. 2 L of mixture aqueous solution was taken for administration.
[0120] Complex B5 group: 5.25 mg of dried Dendrobium candidum polysaccharide, 3.5 mg of lotus extract and 5.25 mg of Glycyrrhiza glabra extract were weighted, mixed, dissolved into 1 mL of sterilized deionized water, and filtered via a 0.22 m microporous membrane to obtain a mixture aqueous solution. 2 L of mixture aqueous solution was taken for administration.
Effect Example
[0121] (1) Determination on Content of Melanin in Cells [0122] A) A375 cells were inoculated in a 6-well plate at 110.sup.5 cells/well, and 2 mL of DMEM culture medium (containing 10% fetal bovine serum) was added to each well for culture. After incubation for 12 h in a CO.sub.2 incubator, a Dendrobium candidum polysaccharide aqueous solution, a lotus extract aqueous solution, a Glycyrrhiza glabra extract aqueous solution and polysaccharide-whitening factor complexes A1-A5 were respectively added according to groups in Example 2, and an untreated group was used as a cell control group (blank group). After incubation for 48 h, the culture medium was discarded, the cells were washed once with PBS. 100 L of cell lysate was added to each well for lysis on ice for 30 min; then the lysed product was centrifuged for 15 min at 12000 r/min; the supernatant was discarded; 150 L of 1 M NaOH solution containing 10% DMSO was added to precipitates; the obtained mixture underwent water bath for 30 min at 80; the supernatant was collected and added to a 96-well plate; and an absorbance value (OD value) was detected at a wavelength of 475 nm with a microplate reader to serve as a detection index of the melanin content. Melanin content=(OD value of assay well-OD value of blank control)/(OD value of cell control group-OD value of blank control). The results were shown in Table 1 and
TABLE-US-00001 TABLE 1 Effects of polysaccharide-whitening factor complexes A1-A5 on the content of melanin in melanocytes (relative value) Group Melanin Blank 1 Dendrobium candidum polysaccharide 0.7867 0.06* Lotus extract 0.8333 0.05 Glycyrrhiza glabra extract 0.8067 0.04* A1 0.6800 0.03** A2 0.7533 0.11** A3 0.6367 0.10**.sup.,b1 A4 0.2800 0.03**.sup.,a2,b2,c2 A5 0.7300 0.14** Note: compared with the blank group, *P < 0.05, **P < 0.01. Compared with the dendrobium candidum polysaccharide alone group, a2 P < 0.01. Compared with the lotus extract alone group, b1 P < 0.05, b2 P < 0.01. Compared with glycyrrhiza glabra extract alone group, c2 P < 0.01.
[0123] It can be seen that compared with the blank control group, all of the polysaccharide-whitening factor complexes A1-A5 can significantly reduce the content of intracellular melanin, indicating that the polysaccharide-whitening factore complexes A1-A5 have a good whitening effect. The Dendrobium candidum polysaccharide alone group and the Glycyrrhiza glabra extract alone group can reduce the content of the melanin in the melanocytes, and have a whitening effect. Where, the polysaccharide-whitening factor complex A3 is significantly better than the lotus extract in the effect of reducing the content of the melanin, and the polysaccharide-whitening factor complex A4 (Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=3:2:2) is significantly better than the Dendrobium candidum polysaccharide alone group, the lotus extract alone group and the Glycyrrhiza glabra extract alone group in the effect of reducing the content of the melanin, indicating that compared with the Dendrobium candidum polysaccharide, the lotus extract or the Glycyrrhiza glabra extract, the polysaccharide-whitening factor complex A4 (Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=3:2:2) has a better whitening effect. The polysaccharide-whitening factor complex A4 (Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=3:2:2) has the best whitening effect. [0124] B) The effects of the polysaccharide-whitening factor complexes B1-B5 on the content of the melanin in the melanocytes were detected according to the above-mentioned specific embodiment.
[0125] The results were shown in Table 2 and
TABLE-US-00002 TABLE 2 Effects of polysaccharide-whitening factor complexes B1-B5 on the content of melanin in melanocytes (relative value) Group Melanin Blank 1 Dendrobium candidum polysaccharide 0.8400 0.09* Lotus extract 0.8733 0.07 Glycyrrhiza glabra extract 0.8500 0.01* B1 0.6400 0.06**.sup.,a2,b2,c2 B2 0.5667 0.03**.sup.,a2,b2,c2 B3 0.2633 0.05**.sup.,a2,b2,c2 B4 0.3900 0.08**.sup.,a2,b2,c2 B5 0.5600 0.08**.sup.,a2,b2,c2 Note: compared with blank group, *P < 0.05, **P < 0.01. Compared with the dendrobium candidum polysaccharide alone group, a2 P < 0.01. Compared with the lotus extract alone group, b2 P < 0.01. Compared with the glycyrrhiza glabra extract alone group, c2 P < 0.01.
[0126] It can be seen that compared with the blank control group, all of the polysaccharide-whitening factor complexes B1-B5 can significantly reduce the content of intracellular melanin, indicating that the polysaccharide-whitening factor complexes B1-B5 have good whitening effect. The Dendrobium candidum polysaccharide alone group and the Glycyrrhiza glabra extract alone group can reduce the content of the melanin in the melanocytes, and have the whitening effect. Where, the polysaccharide-whitening factor complexes B1-B5 are significantly better than the Dendrobium candidum polysaccharide alone group, the lotus extract alone group and the Glycyrrhiza glabra extract alone group in the effect of reducing the content of intracellular melanin, indicating that compared with the Dendrobium candidum polysaccharide, the lotus extract and the Glycyrrhiza glabra extract, the polysaccharide-whitening factor complexes B1-B5 have a better whitening effect. Among them, the polysaccharide-whitening factor complex B3 (Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=3:2:2) has the strongest effect of reducing the content of the intracellular melanin and has the best whitening effect.
[0127] (2) Detection of the Expression of HaCaT Cell Hydration Related mRNA Via Real-Time PCR [0128] A) HaCaT cells were inoculated to a 6-well plate at 210.sup.5 cells/well, and 2 mL of DMEM culture medium (containing 10% fetal bovine serum) was added to each well for culture. After incubation for 12 h in a CO.sub.2 incubator, a Dendrobium candidum polysaccharide aqueous solution, a lotus extract aqueous solution, a Glycyrrhiza glabra extract aqueous solution and polysaccharide-whitening factor complexes A1-A5 were respectively added according to groups in Example 2. After treatment for 24 h, the untreated group was used as cell control group (blank group). Except for the blank group, the culture plate was placed in an ultraviolet irradiator for UVB (20 mJ) irradiation and then incubated for 2 h in an incubator, and the cells were lysed for extraction of total RNA. The expression level of mRNA in the HaCaT cell hydration related gene AQP3 was detected via Real-Time PCR to serve as a detection index for hydration. The results were shown in Table 3 and
TABLE-US-00003 TABLE 3 Effects of polysaccharide-whitening factor complexes A1-A5 on the expression of aquaporin AQP3 (relative value) Group AQP3 Blank 1 UVB 0.79 0.04.sup.## Dendrobium candidum polysaccharide 0.79 0.06 Lotus extract 1.03 0.06** Glycyrrhiza glabra extract 0.72 0.04 A1 1.09 0.05.sup.a2,c2 A2 1.24 0.06**.sup.,a2,b2,c2 A3 1.08 0.07**.sup.,a2,c2 A4 2.13 0.07**.sup.,a2,b2,c2 A5 1.23 0.10**.sup.,a2,b2,c2 Note: by comparing UVB group with blank group, .sup.##P < 0.01; and compared with UVB group, **P < 0.01. By comparing the polysaccharide-whitening factor complexes with the dendrobium candidum polysaccharide alone group, a2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the lotus extract alone group, b2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the glycyrrhiza glabra extract alone group, c2P < 0.01.
[0129] It can be seen that compared with the blank control group, the UVB group can significantly reduce the expression level of AQP3, resulting in a drying problem. Compared with the UVB group, the lotus extract alone group can significantly up-regulate the expression level of AQP3, and the polysaccharide-whitening factor complexes A2-A5 can significantly up-regulate the expression the of AQP3, so as to play the moisturizing effect. The effects of polysaccharide-extracting whitening factor complexes A1-A5 in up-regulating the expression level of AQP3 are significantly higher than those of the Dendrobium candidum polysaccharide alone group and the Glycyrrhiza glabra alone group. Compared with the lotus extract alone group, the effects of the polysaccharide-whitening factor complexes A2, A4 and A5 in up-regulating the expression level of AQP3 are significantly higher than those of lotus extract. The above data show that compared with the Dendrobium candidum polysaccharide, the lotus extract or the Glycyrrhiza glabra extract, the polysaccharide-whitening factor complexes A2, A3 and A5 have a better moisturizing effect. Where, the polysaccharide-whitening factor complex A4 (Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=3:2:2) has the strongest effect of up-regulating the expression level of AQP3, indicating that polysaccharide-whitening factor complex A4 (Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=3:2:2) has the best moisturizing effect. [0130] B) The effects of the polysaccharide-whitening factor complexes B1-B5 on the expression of aquaporin AQP3 were detected according to the above-mentioned specific embodiment.
[0131] The results were shown in Table 4 and
TABLE-US-00004 TABLE 4 Effects of polysaccharide-whitening factor complexes B1-B5 on the expression of aquaporin AQP3 (relative value) Group AQP3 Blank 1 UVB 0.80 0.05.sup.## Dendrobium candidum polysaccharide 0.80 0.04 Lotus extract 0.97 0.09* Glycyrrhiza glabra extract 0.71 0.02 B1 1.38 0.06**.sup.,a2,b2,c2 B2 1.75 0.09**.sup.,a2,b2,c2 B3 2.30 0.09**.sup.,a2,b2,c2 B4 2.08 0.03**.sup.,a2,b2,c2 B5 1.79 0.06**.sup.,a2,b2,c2 Note: by comparing the UVB group with the blank group, .sup.##P < 0.01; compared with the UVB group, *P < 0.05, **P < 0.01. By comparing the polysaccharide-whitening factor complexes with the dendrobium candidum polysaccharide alone group, a2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the lotus extract alone group, b2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the glycyrrhiza glabra extract alone group, c2 P < 0.01.
[0132] It can be seen that compared with the blank control group, the UVB group can significantly reduce the expression level of AQP3, resulting in a drying problem. Compared with the UVB group, the lotus extract alone group can up-regulate the expression level of AQP3, and has the moisturizing effect. The polysaccharide-whitening factor complexes B1-B5 can significantly up-regulate the expression level of AQP3, and have the moisturizing effect. The effects of polysaccharide-whitening factor complexes B1-B5 in up-regulating the expression level of AQP3 are significantly higher than those of the Dendrobium candidum polysaccharide alone group, the lotus extract alone group or the Glycyrrhiza glabra alone group, indicating that compared with the Dendrobium candidum polysaccharide, the lotus extract or the Glycyrrhiza glabra extract, the polysaccharide-whitening factor complexes B1-B5 have a better moisturizing effect. Where, the polysaccharide-whitening factor complex B3 (Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=3:2:2) has the strongest effect of promoting the expression of AQP3, indicating that it has the best moisturizing effect.
[0133] (3) Detection of the Yield of Reactive Oxygen Species in HaCaT Cells Via Flow Cytometry [0134] A) HaCaT cells were inoculated into a 6-well plate in 210.sup.5 cells/well, and 2 mL of DMEM culture medium (containing 10% fetal bovine serum) was added to each well for culture. After incubation for 24 h in a CO.sub.2 incubator, a Dendrobium candidum polysaccharide aqueous solution, a lotus extract aqueous solution, a Glycyrrhiza glabra extract aqueous solution and polysaccharide-whitening factor complexes A1-A5 were respectively added according to groups in Example 2. After treatment for 24 h, the untreated group was used as a cell control group (blank group). Except for the blank group, the culture plate was placed in an ultraviolet irradiator for UVB (1J) irradiation, a DCFH-DA fluorescent probe diluted with a serum-free culture medium was added, and then the culture plate was placed in the incubator for incubation for 20 min to load the cells. The cell suspension was collected, and the yield of reactive oxygen species (ROS) in the HaCaT cells was detected by the flow cytometry to serve as a detection index for oxidative stress.
[0135] The results were shown in Table 5 and
TABLE-US-00005 TABLE 5 Effects of polysaccharide-whitening factor complexes A1-A5 on the production of ROS induced by UVB Group ROS Blank 1 UVB 5.34 0.19.sup.## Dendrobium candidum polysaccharide 4.56 0.05** Lotus extract 5.40 0.02 Glycyrrhiza glabra extract 4.82 0.17** A1 4.74 0.06**.sup.,b2 A2 4.75 0.24**.sup.,b2 A3 4.97 0.08*.sup.,b1 A4 2.36 0.12**.sup.,a2,b2,c2 A5 4.76 0.17**.sup.,b2 Note: by comparing the UVB group with the blank group, .sup.##P < 0.01; compared with the UVB group, *P < 0.05, **P < 0.01. By comparing the polysaccharide-whitening factor complexes with the dendrobium candidum polysaccharide alone group, a2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the lotus extract alone group, b1 P < 0.05, and b2 P < 0.01. By the comparing polysaccharide-whitening factor complexes with the glycyrrhiza glabra extract alone group, c2 P < 0.01.
[0136] It can be seen that compared with the blank control group, the yield of ROS in the UVB group is significantly increased, indicating that UVB induces excessive production of the ROS, and oxidative damages are caused. Compared with the UVB group, the Dendrobium candidum polysaccharide alone group and the Glycyrrhiza glabra extract alone group can both reduce the production of ROS caused by the UVB, and exert the effect of resisting the oxidative damages, and all the polysaccharide-whitening factor complexes A1-A5 can reduce the production of ROS and exert the effect of resisting the oxidative damages. Compared with the Dendrobium candidum polysaccharide alone group, the lotus extract alone group and the Glycyrrhiza glabra extract alone group, the effect of the polysaccharide-whitening factor complex A4 in inhibiting the production of ROS induced by the UVB is significantly stronger than those of the Dendrobium candidum polysaccharide, the lotus extract and the Glycyrrhiza glabra extract, indicating that compared with the Dendrobium candidum polysaccharide, the lotus extract or the Glycyrrhiza glabra extract, the polysaccharide-whitening factor complex A4 (Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=3:2:2) has better antioxidant effect. [0137] B) The effects of the polysaccharide-whitening factor complexes B1-B5 on the production of ROS induced by the UVB were detected according to the above specific embodiments.
[0138] The results were shown in Table 6 and
TABLE-US-00006 TABLE 6 Effects of polysaccharide-whitening factor complexes B1-B5 on the production of ROS induced by UVB Group ROS Blank 1 UVB 5.44 0.09.sup.## Dendrobium candidum polysaccharide 4.58 0.03** Lotus extract 5.44 0.05 Glycyrrhiza glabra extract 4.92 0.26** B1 3.95 0.08**.sup., a2,b2,c2 B2 3.50 0.08**.sup., a2,b2,c2 B3 2.26 0.12**.sup., a2,b2,c2 B4 3.62 0.04**.sup., a2,b2,c2 B5 3.94 0.19**.sup., a2,b2,c2 Note: by comparing the UVB group with the blank group, .sup.##P < 0.01; compared with the UVB group, *P < 0.01, **P < 0.01. By comparing the polysaccharide-whitening factor complexes with the dendrobium candidum polysaccharide alone group, a2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the lotus extract alone group, b2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the glycyrrhiza glabra extract alone group, c2 P < 0.01.
[0139] It can be seen that compared with the blank control group, the production of ROS in the UVB group is significantly increased, indicating that the UVB induces excessive production of the ROS, and oxidative damages are caused. Compared with the UVB group, the Dendrobium candidum polysaccharide alone group and the Glycyrrhiza glabra extract alone group can both reduce the production of ROS caused by the UVB, and exert the effect of resisting the oxidative damages, and all the polysaccharide-whitening factor complexes B1-B5 can reduce the production of ROS, and exert the effect of resisting the oxidative damages. Compared with the Dendrobium candidum polysaccharide alone group, the lotus extract alone group and the Glycyrrhiza glabra extract alone group, the effects of the polysaccharide-whitening factor complexes B1-B5 in reducing the production of ROS are significantly stronger than those of the Dendrobium candidum polysaccharide, the lotus extract and the Glycyrrhiza glabra extract, indicating that compared with the Dendrobium candidum polysaccharide, the lotus extract or the Glycyrrhiza glabra extract, the polysaccharide-whitening factor complexes B1-B5 have better antioxidant effect. Among them, the polysaccharide-whitening factor complex B3 (Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=3:2:2) has the strongest effect of inhibiting the production of ROS, indicating that it has the best antioxidant effect.
[0140] (4) Detection on the Expression of HaCaT Cell Inflammation Related mRNA Via Real-Time PCR [0141] A) HaCaT cells were inoculated into a 6-well plate in 210.sup.5 cells/well, and 2 mL of DMEM culture medium (containing 10% fetal bovine serum) was added to each well for culture. After incubation for 24 h in a CO.sub.2 incubator, a Dendrobium candidum polysaccharide aqueous solution, a lotus extract aqueous solution, a Glycyrrhiza glabra extract aqueous solution and the polysaccharide-whitening factor complexes A1-A5 were respectively added according to groups in Example 2. After treatment for 24 h, the untreated group was used as cell control group (blank group). Except for the blank group, the culture plate was placed in the ultraviolet irradiator for UVB (15 mJ) irradiation, and the cells were lysed for extraction of total RNA. The expression level of mRNA of HaCaT cell inflammation related factors (IL-1, IL-1, IL-6 and TNF-) was detected via a Real-Time PCR experiment to serve as a detection index for an inflammatory response.
[0142] The results were seen in Table 7,
TABLE-US-00007 TABLE 7 Effects of polysaccharide-whitening factor complexes A1-A5 on UVB-induced inflammatory response Group IL-1 IL-1 IL-6 TNF- Blank 1 1 1 1 UVB 1.86 0.06.sup.## 1.98 0.06.sup.## 7.05 0.46.sup.## 5.22 0.28.sup.## Dendrobium 1.53 0.11** 1.73 0.11** 5.84 0.19** 4.43 0.38** candidum polysaccharide (DCP) Lotus extract 1.61 0.06** 1.94 0.02 6.54 0.62 3.96 0.20** (LE) Glycyrrhiza 1.22 0.12** 1.49 0.11** 7.03 0.11 5.03 0.44 glabra extract (GGE) A1 1.40 0.06**.sup.,b1 1.54 0.10**.sup.,a1,b2 6.72 0.43 4.41 0.08**.sup.,c1 A2 1.20 0.10**.sup., a2,b2 1.73 0.09**.sup.,b1 6.26 0.15.sup.*,c1 3.60 0.16**.sup., a2,c2 A3 1.14 0.07**.sup., a2,b2 1.49 0.03**.sup.,a2,b2 5.35 0.09.sup.**,b2,c2 3.94 0.31**.sup.,c2 A4 0.42 0.06**.sup., a2,b2,c2 0.69 0.04**.sup.,a2,b2,c2 2.44 0.24.sup.**,a2,b2,c2 1.99 0.08**.sup.,a2,b2,c2 A5 1.48 0.03** 1.87 0.05** 5.08 0.32.sup.**,b2,c2 3.30 0.26**.sup.,a2,b1,c2 Note: by comparing the UVB group with blank group, .sup.##P < 0.01; compared with the UVB group, *P < 0.01, **P < 0.01. By comparing the polysaccharide-whitening factor complexes with the dendrobium candidum polysaccharide alone group, a1 P < 0.05, and a2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the lotus extract alone group, b1 P < 0.05, and b2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the glycyrrhiza glabra extract alone group, c1 P < 0.01, and c2 P < 0.01.
[0143] It can be seen that compared with the blank control group, the expression levels of inflammatory factors in the UVB group are significantly increased, indicating that UVB irradiation can lead to cellular inflammatory responses. Compared with the UVB group, the Dendrobium candidum polysaccharide alone group can significantly reduce the levels of inflammatory factors IL-1, IL-1, IL-6 and TNF- and exert the anti-inflammatory effect, and the lotus extract and the Glycyrrhiza glabra extract can significantly reduce the levels of some inflammatory factors and exert the partial anti-inflammatory effect. The polysaccharide-extracting whitening factor complex A1 can significantly reduce the levels of some inflammatory factors, and the polysaccharide-extracting whitening factor complexes A2-A5 can significantly reduce the levels of inflammatory factors IL-1, IL-1, IL-6 and TNF- and exert good anti-inflammatory effect. Compared with the Dendrobium candidum polysaccharide alone group, the lotus extract alone group and the Glycyrrhiza glabra extract alone group, the effects of the polysaccharide-whitening factor complexes A1, A2, A3 and A5 in reducing the expression level of some inflammatory factors are partially stronger than that of the Dendrobium candidum polysaccharide, the lotus extract or the Glycyrrhiza glabra extract. The polysaccharide-whitening factor complex A4 can reduce the expression levels of all inflammatory factors and has the effect stronger than that of the Dendrobium candidum polysaccharide, the lotus extract or the Glycyrrhiza glabra extract, indicating that compared with the Dendrobium candidum polysaccharide, the lotus extract or the Glycyrrhiza glabra extract, the polysaccharide-whitening factor complex A4 (Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=3:2:2) has the best anti-inflammatory effect. [0144] B) The effects of the polysaccharide-whitening factor complexes B1-B5 on inflammatory response caused by UVB were detected according to the above specific embodiment.
[0145] The results were shown in Table 8,
TABLE-US-00008 TABLE 8 Effects of polysaccharide-whitening factor complexes B1-B5 on UVB-induced inflammatory response Group IL-1 IL-1 IL-6 TNF- Blank 1 1 1 1 UVB 1.89 0.06.sup.## 2.00 0.12.sup.## 6.85 0.46.sup.## 5.42 0.28.sup.## Dendrobium 1.56 0.13** 1.77 0.03* 5.67 0.19** 4.57 0.38** candidum polysaccharide (DCP) Lotus extract 1.65 0.05** 1.99 0.06 6.24 0.62 4.03 0.20** (LE) Glycyrrhiza 1.25 0.12** 1.53 0.12** 7.16 0.11 5.20 0.16 glabra extract (GGE) B1 0.98 0.11**.sup.,a2,b2,c2 1.00 0.06**.sup.,a2,b2,c2 4.55 0.31**.sup.,a2,b2,c2 3.14 0.08**.sup.,a2,b2,c2 B2 0.54 0.09**.sup.,a2,b2,c2 0.83 0.10**.sup.,a2,b2,c2 3.29 0.15**.sup.,a2,b2,c2 2.67 0.16**.sup.,a2,b2,c2 B3 0.36 0.04**.sup.,a2,b2,c2 0.49 0.07**.sup.,a2,b2,c2 2.29 0.09**.sup.,a2,b2,c2 1.91 0.31**.sup.,a2,b2,c2 B4 0.67 0.04**.sup.,a2,b2,c2 0.66 0.06**.sup.,a2,b2,c2 3.15 0.25**.sup.,a2,b2,c2 2.36 0.08**.sup.,a2,b2,c2 B5 0.87 0.05**.sup.,a2,b2,c2 0.88 0.03**.sup.,a2,b2,c2 4.32 0.32**.sup.,a2,b2,c2 3.34 0.27**.sup.,a2,b2,c2 Note: by comparing the UVB group with the blank group, .sup.##P < 0.01; compared with the UVB group, *P < 0.05, and ** P < 0.01. By comparing the polysaccharide-whitening factor complexes with the dendrobium candidum polysaccharide alone group, a2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the lotus extract alone group, b2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the glycyrrhiza glabra extract alone group, c2 P < 0.01.
[0146] It can be seen that compared with the blank control group, the expression levels of inflammatory factors in the UVB group are significantly increased, indicating that UVB irradiation may lead to cellular inflammatory responses. Compared with the UVB group, the Dendrobium candidum polysaccharide alone group can significantly reduce the levels of the inflammatory factors IL-1, IL-1, IL-6, TNF-, and exert the anti-inflammatory effect; and the lotus extract and the Glycyrrhiza glabra extract can significantly reduce the levels of some inflammatory factors and exert the partial anti-inflammatory effect. All the polysaccharide-polysaccharide complexes B1-B5 can significantly reduce the expression levels of the inflammatory factors IL-1, IL-1, IL-6 and TNF-, indicating that the polysaccharide-whitening factor complexes B1-B5 have good anti-inflammatory effect. In addition, compared with the Dendrobium candidum polysaccharide alone group, the lotus extract alone group and the Glycyrrhiza glabra extract alone group, the effects of the polysaccharide-whitening factor complexes B1-B5 in reducing the expression levels of all inflammatory factors are stronger than that of the Dendrobium candidum polysaccharide, the lotus extract or the Glycyrrhiza glabra extract, indicating that compared with the Dendrobium candidum polysaccharide, the lotus extract or the Glycyrrhiza glabra extract, the polysaccharide-whitening factor complexes B1-B5 have better anti-inflammatory effects. Among them, the polysaccharide-whitening factor complex B3 (Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=3:2:2) has the strongest effect of reducing the expression level of inflammatory factors, indicating that it has the best anti-inflammatory effect.
[0147] (5) Detection on the Expression of HaCaT Cell Lipid Synthesis Related mRNA Via Real-Time PCR
[0148] HaCaT cells were inoculated into a 6-well plate in 210.sup.5 cells/well, and 2 mL of DMEM culture medium (containing 10% fetal bovine serum) was added to each well for culture. After incubation for 24 h in a CO.sub.2 incubator, a Dendrobium candidum polysaccharide aqueous solution, a lotus extract aqueous solution, a Glycyrrhiza glabra extract aqueous solution and the polysaccharide-whitening factor complexes A1-A5 were respectively added according to groups in Example 2. The untreated group was used as the cell control group (blank group). Except for the blank group, 10 mM Ca.sup.2+ was added to each group. After treatment for 48 h, the cells were lysed for extraction of total RNA. The expression level of mRNA of HaCaT cell lipid synthesis related factors (CerS3, CerS4, FAS, SPT, ELOVL1, ELOVL4 and ELOVL6) were detected via a Real-Time PCR experiment to serve as an index for detecting a lipid content.
[0149] The results were shown in Table 9,
TABLE-US-00009 TABLE 9 Effects of polysaccharide-whitening factor complexes A1-A5 on Ca.sup.2+-induced lipid synthesis factor expression Group Cers3 CerS4 FAS Blank 1 1 1 Ca.sup.2+ 2.04 0.08.sup.## 1.68 0.13.sup.## 1.54 0.02.sup.## Dendrobium candidum polysaccharide 2.12 0.08 2.05 0.32 1.70 0.03* (DCP) Lotus extract (LE) 1.62 0.10 1.67 0.08 1.55 0.08 Glycyrrhiza glabra extract (GGE) 0.93 0.10 1.32 0.13 1.37 0.05 A1 1.73 0.14.sup.,c2 1.55 0.12 1.61 0.05.sup.c2 A2 1.06 0.07 1.00 0.11 1.41 0.02 A3 2.10 0.18.sup.,b2,c2 2.55 0.23**.sup.,a1,b2,c2 1.57 0.08.sup.c1 A4 3.79 0.06**.sup., a2,b2,c2 3.79 0.10**.sup.,a2,b2,c2 2.56 0.11**.sup.,a2,b2,c2 A5 2.77 0.10**.sup.,a2,b2,c2 2.98 0.24**.sup.,a2,b2,c2 1.90 0.05**.sup.,a1,b2,c2 SPT ELOVL1 ELOVL4 ELOVL6 1 1 1 1 1.57 0.02.sup.## 1.37 0.02.sup.## 2.01 0.06.sup.## 1.36 0.03.sup.## 1.62 0.03 1.50 0.12 2.21 0.03** 1.54 0.06* 1.28 0.07 1.65 0.12 1.33 0.05 1.30 0.08 1.25 0.06 1.55 0.15 1.62 0.02 0.95 0.03 1.51 0.09.sup.b2,c2 1.49 0.16 1.91 0.10.sup.b2,c2 1.07 0.08 1.25 0.02 1.62 0.16 1.98 0.06.sup.b2,c2 1.16 0.02.sup.c1 1.31 0.05 1.78 0.02** 1.82 0.06.sup.b2,c2 1.32 0.04.sup.c2 2.41 0.09**.sup.,a2,b2,c2 2.53 0.09**.sup.,a2,b2,c2 3.04 0.07.sup.**,a2,b2,c2 2.52 0.13**.sup.,a2,b2,c2 1.68 0.04.sup.b2,c2 1.60 0.08 2.45 0.04.sup.**,a2,b2,c2 1.35 0.11.sup.c2 Note: by comparing the Ca.sup.2+ group with the blank group, .sup.##P < 0.01; compared with the Ca.sup.2+ group, *P < 0.05, and **P < 0.01. By comparing the polysaccharide-whitening factor complexes with the dendrobium candidum polysaccharide alone group, a1 P < 0.05, and a2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the lotus extract alone group, b2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the glycyrrhiza glabra extract alone group, c1 P < 0.05, and c2 P < 0.01.
[0150] It can be seen that compared with the blank control group, the expression levels of the lipid synthesis related factors CerS3, CerS4, FAS, SPT, ELOVL1, ELOVL4 and ELOVL6 in the Ca.sup.2+ group are all significantly increased, indicating that Ca.sup.2+ stimulation may induce lipid synthesis. Compared with the Ca.sup.2+ group, the Dendrobium candidum polysaccharide alone group can promote increase in expression levels of Ca.sup.2+-induced FAS, ELOVL4 and ELOVL6, indicating that the Dendrobium candidum polysaccharide alone group can partially promote lipid synthesis, and the polysaccharide-whitening factor complex A3 can increase the expression level of some lipid synthesis related genes CerS4 and ELOVL1, and the polysaccharide-whitening factor complex A5 can increase the expression level of some lipid synthesis related genes CerS3, CerS4, FAS and ELOVL4, indicating that the polysaccharide-whitening factor complexes A3 and A5 can partially promote lipid synthesis. The polysaccharide-whitening factor complex A4 can promote the expression of all the lipid synthesis related factors CerS3, CerS4, FAS, ELOVL4 and ELOVL6, indicating that the polysaccharide-whitening factor complex A4 has the effect of completely promoting lipid synthesis. Compared with the Dendrobium candidum polysaccharide alone group, the effect of the polysaccharide-extracting whitening factor complex A3 in increasing the expression level of the lipid synthesis related gene CerS4 is better than that of the Dendrobium candidum polysaccharide, and the effect of the polysaccharide-extracting whitening factor complex A5 in increasing the expression levels of the lipid synthesis related factors CerS3, CerS4, FAS and ELOVL4 was better than that of the Dendrobium candidum polysaccharide, and the effect of the polysaccharide-whitening factor complex A4 in increasing the expression levels of all the lipid synthesis related factors CerS3, CerS4, FAS, SPT, ELOVL1, ELOVL4 and ELOVL6 is significantly better than that of the Dendrobium candidum polysaccharide, indicating that compared with the Dendrobium candidum polysaccharide, the lotus extract or the Glycyrrhiza glabra extract, the polysaccharide-whitening factor complex A4 (Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=3:2:2) has the best effect of promoting lipid synthesis. [0151] B) The effects of polysaccharide-whitening factor complexes B1-B5 on the expression levels of Ca.sup.2+-induced lipid synthesis factors were detected according to the above-mentioned specific embodiment.
[0152] The results were shown in Table 10,
TABLE-US-00010 TABLE 10 Effects of polysaccharide-whitening factor complexes B1-B5 on the expression levels of Ca.sup.2+-induced lipid synthesis factors Group CerS3 CerS4 FAS Blank 1 1 1 Ca.sup.2+ 1.99 0.08.sup.## 1.60 0.12.sup.## 1.52 0.05.sup.## Dendrobium candidum polysaccharide 2.21 0.03 1.82 0.18 1.70 0.04* (DCP) Lotus extract (LE) 1.58 0.06 1.65 0.08 1.44 0.07 Glycyrrhiza glabra extract (GGE) 1.00 0.05 1.35 0.10 1.30 0.11 B1 2.86 0.13**.sup.,a2,b2,c2 2.50 0.13**.sup.,a2,b2,c2 2.05 0.08**.sup.,a2,b2,c2 B2 3.19 0.19**.sup.,a2,b2,c2 3.15 0.14**.sup.,a2,b2,c2 2.32 0.07**.sup.,a2,b2,c2 B3 3.95 0.08**.sup.,a2,b2,c2 3.83 0.06**.sup.,a2,b2,c2 2.69 0.02**.sup.,a2,b2,c2 B4 3.24 0.25**.sup.,a2,b2,c2 3.16 0.10**.sup.,a2,b2,c2 2.51 0.04**.sup.,a2,b2,c2 B5 2.82 0.06**.sup.,a2,b2,c2 2.67 0.15**.sup.,a2,b2,c2 2.44 0.09**.sup.,a2,b2,c2 SPT ELOVL1 ELOVL4 ELOVL6 1 1 1 1 1.52 0.05.sup.## 1.32 0.03.sup.## 1.91 0.05.sup.## 1.37 0.04.sup.## 1.59 0.08 1.40 0.09 2.17 0.10* 1.62 0.06** 1.30 0.08 1.53 0.04 1.37 0.09 1.27 0.06 1.25 0.04 1.50 0.09 1.56 0.07 0.97 0.05 1.92 0.06**.sup.,a2,b2,c2 2.05 0.11 2.51 0.08**.sup.,a2,b2,c2 1.84 0.07**.sup.,a1,b2,c2 2.21 0.12**.sup.,a2,b2,c2 2.29 0.11**.sup.,a2,b2,c2 2.86 0.07**.sup.,a2,b2,c2 1.91 0.12**.sup.,a2,b2,c2 2.44 0.14**.sup.,a2,b2,c2 2.64 0.07**.sup.,a2,b2,c2 3.07 0.12**.sup.,a2,b2,c2 2.54 0.11**.sup.,a2,b2,c2 2.22 0.08**.sup.,a2,b2,c2 2.44 0.08**.sup.,a2,b2,c2 2.93 0.13**.sup.,a2,b2,c2 1.87 0.09**.sup.,a1,b2,c2 1.80 0.08**.sup.,a1,b2,c2 2.12 0.19**.sup.,a2,b2,c2 2.55 0.09**.sup.,a2,b2,c2 1.83 0.10**.sup.,a1,b2,c2 Note: by comparing the Ca.sup.2+ group with the blank group, .sup.##P < 0.01; compared with the Ca.sup.2+ group, *P < 0.05, and **P < 0.01. By comparing the polysaccharide-whitening factor complexes with the dendrobium candidum polysaccharide alone group, a1 P < 0.05, and a2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the lotus extract alone group, b2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the glycyrrhiza glabra extract alone group, c2 P < 0.01.
[0153] It can be seen that compared with the blank control group, the expression levels of the lipid synthesis related factors CerS3, CerS4, FAS, SPT, ELOVL1, ELOVL4 and ELOVL6 in the Ca.sup.2+ group are significantly increased, indicating that Ca.sup.2+ stimulation may induce lipid synthesis. Compared with the Ca.sup.2+ group, the Dendrobium candidum polysaccharide alone group can promote increase in expression levels of Ca.sup.2+-induced FAS, ELOVL4 and ELOVL6, indicating that Dendrobium candidum polysaccharide alone group can partially promote lipid synthesis. The polysaccharide-whitening factor complexes B1-B5 can promote the expression of all the lipid synthesis related factors CerS3, CerS4, FAS, SPT, ELOVL1, ELOVL4 and ELOVL6, indicating that the polysaccharide-whitening factor complexes B1-B5 have the effect of comprehensively promoting lipid synthesis. Compared with the Dendrobium candidum polysaccharide alone group, the effects of the polysaccharide-whitening factor complexes B1-B5 in increasing the expression levels of all the lipid synthesis related factors are significantly higher than that of the Dendrobium candidum polysaccharide, indicating that compared with the Dendrobium candidum polysaccharide, the lotus extract or the Glycyrrhiza glabra extract, the polysaccharide-whitening factor complexes B1-B5 have better effect of promoting lipid synthesis. Among them, the polysaccharide-whitening factor complex B3 (Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=3:2:2) has the strongest effect of promoting lipid synthesis, indicating that it has the best effect of promoting lipid synthesis.
[0154] (6) Detection on the Content of Lipid in HaCaT Cells Via Fluorescence Staining
[0155] HaCaT cells were inoculated into a 6-well plate in 210.sup.5 cells/well, and 2 mL of DMEM culture medium (containing 10% fetal bovine serum) was added to each well for culture. After incubation for 24 h in a CO.sub.2 incubator, a Dendrobium candidum polysaccharide aqueous solution, a lotus extract aqueous solution, a Glycyrrhiza glabra extract aqueous solution and polysaccharide-whitening factor complexes A1-A5 were respectively added according to groups in Example 2. The untreated group was used as the cell control group (blank group). Except for the blank group, 10 mM Ca.sup.2+ was added to each group. After treatment for 48 h, a Hoechst dye with an original concentration of 1.8 mM was diluted with a complete culture medium to obtain a 20 M Hoechst working solution. Nile red dye powder was dissolved into ethanol to obtain a 2 mg/mL Nile red solution, and 10 L of Nile red solution was added to each 4 mL of PBS to prepare a working solution. A Hoechst nucleus dye was added to cells and stained for 15 min in an incubator, then 0.6-1 mL of Nile red staining solution was added, the plate was placed in the incubator for culture for 20 min to label lipid components, and the fluorescence intensity was observed under a fluorescent inverted microscope to serve as an auxiliary detection index for a lipid content.
TABLE-US-00011 TABLE 11 Effects of polysaccharide-whitening factor complexes A1-A5 on intracellular lipid content Group Lipid content Blank 1 Ca.sup.2+ 1.20 0.05.sup.## Dendrobium candidum polysaccharide 1.15 0.05 Lotus extract 0.89 0.06 Glycyrrhiza glabra extract 0.82 0.01 A1 1.22 0.06.sup.b2,c2 A2 0.98 0.06.sup.c1 A3 1.12 0.07.sup.b2,c2 A4 1.79 0.10**.sup.,a2,b2,c2 A5 1.32 0.06.sup.a1,b2,c2 Note: by comparing the Ca.sup.2+ group with the blank group, .sup.##P < 0.01; and compared with the Ca.sup.2+ group, **P < 0.01. By comparing the polysaccharide-whitening factor complexes with the dendrobium candidum polysaccharide alone group, a1 P < 0.05, and a2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the lotus extract alone group, b2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the glycyrrhiza glabra extract alone group, c1 P < 0.05, and c2 P < 0.01.
[0156] The results were shown in Table 11 and
[0157] It can be seen that compared with the blank group, the lipid content of the Ca.sup.2+ group is significantly increased, indicating that Ca.sup.2+ stimulation may induce lipid synthesis. Compared with the Ca.sup.2+ group, treatment with the polysaccharide-whitening factor complex A4 can promote lipid synthesis induced by Ca.sup.2+ stimulation, indicating that the polysaccharide-whitening factor complex A4 has the effect of promoting lipid synthesis. Compared with the Dendrobium candidum polysaccharide alone group, the polysaccharide-whitening factor complexes A4 and A5 have a stronger effect of promoting lipid synthesis. Compared with the lotus extract alone group, the polysaccharide-whitening factor complexes A1, A3, A4, and A5 have a stronger effect of promoting lipid synthesis. Compared with the Glycyrrhiza glabra extract, the polysaccharide-whitening factor complexes A1-A5 have a stronger effect of promoting lipid synthesis. The above results show that the ability of the polysaccharide-whitening factor complex A4 to promote the increase of lipid is significantly better than that of the Dendrobium candidum polysaccharide, the lotus extract or the Glycyrrhiza glabra extract, the polysaccharide-whitening factor complex A4 (Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=3:2:2) has the best effect of promoting lipid synthesis. [0158] B) The effects of the polysaccharide-whitening factor complexes B1-B5 on a content of intracellular lipid were detected according to the above specific embodiment.
[0159] The results were shown in Table 12 and
TABLE-US-00012 TABLE 12 Effects of polysaccharide-whitening factor complexes B1-B5 on the content of intracellular lipid Group Lipid content Blank 1 Ca.sup.2+ 1.21 0.05.sup.# Dendrobium candidum polysaccharide 1.25 0.09 Lotus extract 0.93 0.05 Glycyrrhiza glabra extract 0.88 0.03 B1 1.60 0.09**.sup.,a2,b2,c2 B2 1.71 0.11**.sup.,a2,b2,c2 B3 1.91 0.09**.sup.,a2,b2,c2 B4 1.82 0.10**.sup.,a2,b2,c2 B5 1.55 0.12**.sup.,a2,b2,c2 Note: by comparing the Ca.sup.2+ group with blank group, .sup.##P < 0.05; and compared with the Ca.sup.2+ group, **P < 0.01. By comparing the polysaccharide-whitening factor complexes with the dendrobium candidum polysaccharide alone group, a2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the lotus extract alone group, b2 P < 0.01. By comparing the polysaccharide-whitening factor complexes with the glycyrrhiza glabra extract alone group, c2 P < 0.01.
[0160] It can be seen that compared with the blank group, the lipid content of the Ca.sup.2+ group is significantly increased, indicating that Ca.sup.2+ stimulation may induce lipid synthesis. Compared with the Ca.sup.2+ group, treatment with the polysaccharide-whitening factor complexes B1-B5 can promote the lipid synthesis induced by Ca.sup.2+ stimulation, indicating that the polysaccharide-whitening factor complexes B1-B5 have the effect of promoting lipid synthesis. Compared with the Dendrobium candidum polysaccharide alone group, the lotus extract alone group and the Glycyrrhiza glabra extract alone group, the polysaccharide-whitening factor complexes B1-B5 have a stronger effect of promoting lipid synthesis, indicating that compared with the Dendrobium candidum polysaccharide, the lotus extract or the Glycyrrhiza glabra extract, the polysaccharide-whitening factor complexes B1-B5 have a better effect of promoting lipid synthesis. Among them, the polysaccharide-whitening factor complex B3 (Dendrobium candidum polysaccharide:lotus extract: Glycyrrhiza glabra extract=3:2:2) has the best effect of promoting lipid synthesis, indicating that it has the best effect of promoting lipid synthesis.
[0161] The present disclosure has been illustrated in detail by the above-mentioned specific embodiments, but the present disclosure is not limited to the above-mentioned embodiments, various variations can also be made within the knowledge scope possessed by persons of ordinary skill in the art and without departing from the spirit of the present disclosure. In addition, embodiments of the present disclosure and features in embodiments can be combined with each other without conflict.