Tuberose extract, preparation method and application thereof

Abstract

The application discloses a tuberose extract and a preparation method and application thereof, and comprises pretreatment, extraction of oil-soluble extract, addition of the pretreated dry flower powder into a solvent, heating extraction, centrifugation, collection of supernatant, filtration, collection of liquid phase, and obtaining of the tuberose extract. The tuberose extract has the functions of antioxidation, inhibition of cortisol and lipid antioxidation, can efficiently remove free radicals at a lower concentration, significantly inhibit lipid peroxidation, help skin resist oxidative damage, effectively regulate cortisol level, and relieve endogenous and exogenous stress. The application has a mild process, extracts fat-soluble active ingredients of tuberose, avoids residue problems of volatile organic solvents, guarantees product safety, and provides a new multifunctional, high-activity and high-safety raw material for the cosmetic industry.

Description

Technical Field

[0001] This invention belongs to the field of cosmetic raw material preparation technology, specifically relating to a tuberose extract, its preparation method, and its application. Background Technology

[0002] Tuberose (Polianthes tuberosa L.), also known as night-blooming jasmine or moon-scented jasmine, is an ornamental plant renowned for its unique and intense fragrance. Its aroma is particularly pronounced from evening to night, making it a highly sought-after ingredient in perfumes, cosmetics, and aromatherapy. Traditionally, the aromatic components of tuberose are primarily extracted to produce essential oils or extracts using steam distillation or organic solvents (such as petroleum ether, ethanol, and hexane). Known volatile components of tuberose mainly include geraniol, eugenol, farnesol, methyl benzoate, methyl anthranilate, and benzyl alcohol.

[0003] While there is relatively much research on the volatile components (essential oil / extract) of tuberose, research on its non-volatile or weakly volatile fat-soluble active components is significantly lacking. Current mainstream techniques still heavily rely on volatile organic solvents (VOCs) for extraction and purification. This solvent-dependent extraction method presents a critical, unavoidable problem: the risk of residual organic solvents.

[0004] Therefore, developing a novel extraction method that can effectively extract the fat-soluble active ingredients of tuberose while avoiding the problem of volatile organic solvent residues and ensuring product safety has become a key technical problem that urgently needs to be solved in this field. This is not only necessary to enhance the utilization value of tuberose resources, but also an inevitable choice to meet the market's growing demand for safe, natural, and efficient cosmetic raw materials. Summary of the Invention

[0005] The purpose of this section is to outline some aspects of the embodiments of the present invention and to briefly describe some preferred embodiments.

[0006] As one aspect of the present invention, the present invention provides a method for preparing tuberose extract, which includes the following steps:

[0007] (1) Pretreatment: Dry and pulverize fresh tuberose flowers to obtain dried flower powder;

[0008] (2) Extraction of oil-soluble extract: The pretreated dried flower powder is added to a solvent and heated at 30-55 °C for 1-4 hours; the solvent is a mixture of meadowfoam seed oil and caprylic / capric triglyceride.

[0009] (3) Centrifuge, collect the supernatant, filter, collect the liquid phase, and obtain the tuberose extract.

[0010] The method for preparing tuberose extract is characterized in that: in step (2), the volume ratio of meadowfoam seed oil and caprylic / capric triglyceride is 2.5~4:1.

[0011] In a preferred embodiment of the preparation method of tuberose extract described in this invention, in step (2), the ratio of dried flower powder to solvent is 1:(4~9) g / mL.

[0012] In one preferred embodiment of the preparation method of tuberose extract described in this invention, in step (2), the ratio of dried flower powder to solvent is 1:(6~7) g / mL.

[0013] In one preferred embodiment of the preparation method of tuberose extract described in this invention, the extraction temperature in step (2) is 40~50 ℃.

[0014] In one preferred embodiment of the preparation method of tuberose extract described in this invention, in step (1), the pulverization is to pulverize fresh tuberose flowers to 50-500 mesh.

[0015] In a preferred embodiment of the preparation method of tuberose extract described in this invention: in step (3), the centrifugation includes centrifugation at 1000~5000 rpm for 5~20 min, and the filtration includes vacuum filtration using a 0.22~0.45 µm filter membrane.

[0016] The present invention also provides a tuberose extract prepared by the method described above.

[0017] The present invention also provides the use of the tuberose extract in the preparation of products that inhibit cortisol.

[0018] The present invention also provides the application of the tuberose extract in the preparation of cosmetics.

[0019] The beneficial effects of this invention are as follows: The tuberose extract of this invention has antioxidant, cortisol-inhibiting, and lipid antioxidant functions. It can effectively scavenge free radicals at low concentrations, significantly inhibit lipid peroxidation, and help the skin resist oxidative damage. At the same time, it effectively regulates cortisol levels and relieves internal and external stress. The process of this invention is gentle, extracting the fat-soluble active ingredients of tuberose while avoiding the problem of volatile organic solvent residues, ensuring product safety, and providing the cosmetics industry with a multifunctional, highly active, and highly safe new raw material. Attached Figure Description

[0020] To more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings used in the description of the embodiments will be briefly introduced below, wherein:

[0021] Figure 1The antioxidant test results are for the tuberose extract prepared in Example 1.

[0022] Figure 2 To investigate the effect of different extraction material-to-liquid ratios on the ABTS free radical scavenging rate in Example 1.

[0023] Figure 3 To investigate the effect of different extraction times on the ABTS radical scavenging rate in Example 2.

[0024] Figure 4 To investigate the effect of different extraction temperatures on the ABTS radical scavenging rate in Example 3.

[0025] Figure 5 To investigate the cortisol-inhibiting effect of tuberose extract at different temperatures.

[0026] Figure 6 The lipid antioxidant effect of the tuberose extract prepared in Example 1.

[0027] Figure 7 The effect of tuberose extract prepared in Example 1 on Hacat cell proliferation.

[0028] Figure 8 Cortisol concentration was tested in the tuberose extracts prepared in Example 1 and Comparative Examples 1-3. Detailed Implementation

[0029] To make the above-mentioned objectives, features and advantages of the present invention more apparent and understandable, the specific embodiments of the present invention will be described in detail below with reference to specific examples.

[0030] Meadowfoam seed oil: purchased from Guangzhou Huashi Cosmetics Technology Co., Ltd., product number: Z20250530-05, INCI name: LIMNANTHES ALBA (MEADOWFOAM) SEED OIL.

[0031] Caprylic / Capric Triglyceride (GTCC): CAS No.: 65381-09-1, Shanghai Yuanye Biotechnology Co., Ltd., Yuanye #S24816-500g, INCI Name: CAPRYLIC / CAPRIC TRIGLYCERIDE.

[0032] Example 1:

[0033] Fresh tuberose flowers were dried in an oven at 50℃ to obtain dried flowers. The dried flowers were pulverized to 350 mesh and added to a solvent consisting of meadowfoam seed oil and a mixture of caprylic / capric triglycerides. The volume ratio of meadowfoam seed oil to caprylic / capric triglycerides was 3:1. The ratio of dried flowers to solvent was 1:6 (g / mL). The mixture was heated to 40℃ and extracted for 1 hour. After extraction, the mixture was centrifuged at 3000 rpm for 5 minutes. The supernatant was collected and further filtered through a 0.45 μm membrane using vacuum filtration to obtain tuberose extract.

[0034] Study Example 1:

[0035] In this study, the effects of different extraction material-to-solvent ratios on the activity of tuberose extract were investigated. The ratios of dried flowers to solvent were selected as 1:1, 1:3, 1:6, 1:10, and 1:20, respectively, and the extraction time and temperature were set at 1 h and 40 °C, respectively.

[0036] Study Example 2:

[0037] In this study, the effects of different extraction times on the activity of tuberose extract were investigated. The extraction times were 0.5h, 1h, 2h, 4h, and 6h, and the extraction material-to-liquid ratio and temperature were set at 1:6 and 40℃, respectively.

[0038] Study Example 3:

[0039] In Example 1, the effects of different extraction temperatures on the activity of tuberose extract were studied. The extraction temperatures were set to 20℃, 30℃, 40℃, 50℃, and 60℃, and the extraction time and material-liquid ratio were set to 1h and 1:6, respectively.

[0040] Comparative Example 1:

[0041] Fresh tuberose flowers were dried in an oven at 50℃ to obtain dried flowers. The dried flowers were pulverized to 350 mesh and added to a solvent, namely meadowfoam seed oil. The ratio of dried flowers to solvent was 1:6 (g / mL). The mixture was heated to 40℃ and extracted for 1 hour. After extraction, the mixture was centrifuged at 3000 rpm for 5 minutes. The supernatant was collected and further filtered through a 0.45 μm membrane using vacuum filtration to obtain tuberose extract.

[0042] Comparative Example 2:

[0043] Fresh tuberose flowers were dried in an oven at 50℃ to obtain dried flowers. The dried flowers were pulverized to 350 mesh and added to a solvent (caprylic / capric triglyceride). The ratio of dried flowers to solvent was 1:6 (g / mL). The mixture was heated to 40℃ and extracted for 1 hour. After centrifugation at 3000 rpm for 5 minutes, the supernatant was collected and further filtered through a 0.45 μm membrane using vacuum filtration to obtain tuberose extract.

[0044] Comparative Example 3:

[0045] The solvent in Example 1 was replaced with squalane, isononyl isononanoate, or jojoba seed oil, and tuberose extract was prepared in the same manner as in Example 1.

[0046] Test example:

[0047] Antioxidant properties:

[0048] ABTS Method: ABTS Solution Preparation: Dissolve 384 mg of ABTS in 100 mL of deionized water to prepare a 7 mmol / L ABTS solution; dissolve 66 mg of potassium persulfate in 100 mL of deionized water to prepare a 2.45 mmol / L potassium persulfate solution. Then, mix the 7 mmol / L ABTS solution and the 2.45 mmol / L potassium persulfate solution at a volume ratio of 2:1 and allow to react in the dark at room temperature for 16 hours to obtain the ABTS stock solution. Dilute the ABTS stock solution with anhydrous ethanol and n-butanol to achieve an absorbance of approximately 0.5 at 734 nm (dilution ratio: n-butanol: ethanol: ABTS stock solution = 20:20:1.25) to obtain the ABTS working solution. Experimental Group: Add 2 mL of tuberose extract to centrifuge tubes, add 2 mL of ABTS working solution, and bring the volume to 5 mL with n-butanol. Blank Group: Add 2 mL of ABTS working solution to centrifuge tubes, and bring the volume to 5 mL with n-butanol. Solvent group: Add different volumes of tuberose extract to centrifuge tubes, bring the volume to 5 ml with n-butanol, and shake well. After standing for 30 min, measure the absorbance at 734 nm for all three solutions. Calculate the scavenging rate using the formula: ABTS free radical scavenging rate = (A2 - A1 + A3) / A2 × 100%, where A1, A2, and A3 are the absorbances of the experimental group, blank group, and solvent group, respectively.

[0049] DPPH Method: DPPH Solution Preparation: Dissolve 2.5 mg DPPH in 100 mL of anhydrous ethanol to obtain a 0.025 mg / mL solution. Experimental Group: Take different volumes of tuberose extract into centrifuge tubes, add 2 mL of DPPH solution, and dilute to 5 mL with n-butanol. Blank Group: Add 2 mL of DPPH solution into centrifuge tubes and dilute to 5 mL with n-butanol. Solvent Group: Take different volumes of tuberose extract into centrifuge tubes, dilute to 5 mL with n-butanol, and shake well. After reacting the three solutions in the dark for 30 min, measure the absorbance at 517 nm. Calculate the scavenging rate using the formula: DPPH free radical scavenging rate = (A2 - A1 + A3) / A2 × 100%, where A1, A2, and A3 are the absorbances of the experimental group, blank group, and solvent group, respectively.

[0050] Lipid peroxidation experiment:

[0051] Using Fe 2+Experiment on lipid peroxidation catalyzed by egg yolk lecithin: Solution preparation: Prepare 5 mmol / L egg yolk lecithin solution and 20 mmol / L ferrous sulfate solution according to optimal parameters. Experimental group: Take different volumes of tuberose oil-soluble extract into centrifuge tubes, add 1 ml n-butanol, and make up to 5 ml with ethanol. Blank group: Add 1 ml n-butanol into centrifuge tubes, and make up to 5 ml with ethanol. Experimental procedure: Take 0.2 ml of egg yolk lecithin solution into a centrifuge tube, add 0.2 ml of tuberose oil-soluble extract diluent, add 0.2 ml of ferrous sulfate solution, add PBS to make up to 2 ml, incubate at 37℃ for 15 min, add 1 ml of 20% TCA, mix well and let stand for 5 min, add 1 ml of 0.5% TBA, mix well, incubate in boiling water for 10 min, cool with cold water, centrifuge at 8000 r / min for 10 min, and measure the absorbance of the supernatant at 532 nm. The MDA inhibition rate is calculated using the formula: MDA inhibition rate (%) = [(A0−A1) / A0]×100%, where A0 and A1 are the absorbance of the blank group and the experimental group, respectively.

[0052] Evaluation of cortisol-inhibiting efficacy using a cortisone-induced human keratinocyte model:

[0053] 1) Cell culture: DMEM complete medium (containing 10% fetal bovine serum, 1×1) was used. 05 Cells were cultured in an incubator at 37°C, 5% CO2, and saturated humidity with U / L penicillin and 100 mg / L streptomycin. When the cell density reached about 80%, the cells were passaged using trypsin, and passaged every 1-2 days.

[0054] 2) MTT assay to detect the effect of tuberose extract on Hacat cell proliferation

[0055] Add 0.2 ml of tuberose oil-soluble extract and 0.2 ml of DMSO to 9.6 ml of culture medium, and dilute proportionally to obtain samples of different concentrations. Digest Hacat cells in the logarithmic growth phase, count them, and seed them in 96-well plates at a density of 1 × 10⁻⁶ cells per well. 5Cells were seeded at 100 μL / well in 96-well plates and cultured at 37°C in a 5% CO2 incubator for 24 h. Then, 100 μL of pre-prepared samples of different concentrations were added to each well. For the control group, 100 μL of DMEM medium containing the same concentration of DMSO was added to each well. After 16 h of drug addition, the supernatant was discarded. 100 μL of 0.5 mg / mL MTT solution was added to each well, and the plates were incubated for another 4 h, after which the supernatant was discarded. 100 μL of DMSO was added to each well, and the plates were shaken for 10 min in a microplate reader. The absorbance was measured at 490 nm. Cell proliferation rate was calculated using the formula: Cell proliferation rate = A1 / A2 × 100%, where A1 represents the absorbance of the drug group at 490 nm, and A2 represents the absorbance of the control group at 490 nm. The extract concentration (0.05%) with a cell viability greater than 90% was selected for cortisol inhibition experiments.

[0056] 3) Test on the inhibition of cortisol concentration by tuberose oil-soluble extract

[0057] HaCat cells in the logarithmic growth phase were collected and diluted with complete culture medium to a cell density of 1.5 × 10⁻⁶ cells / year. 5 Cells / mL were seeded into 96-well cell culture plates, 100 μL per well. The plates were incubated at 37°C, 5% CO2, and saturated humidity in a CO2 incubator. After 24 h, the old culture medium was removed, and blank control, negative control, positive control, and sample groups were established. The blank control group received only fresh culture medium; the negative control group received fresh culture medium containing cortisone (20 µg / mL); the positive control group received fresh culture medium containing cortisone and the positive control drug memetidine; and the sample groups received fresh culture medium containing cortisone and the drug-concentrated sample, 100 μL per well. Each experimental group had three replicates, and the cells were incubated at 37°C, 5% CO2, and saturated humidity in a CO2 incubator for 24 h. After incubation, the cell supernatant was collected to detect cortisol secretion levels in each group. The Human Cortisol ELISA Kit was used to test the inhibitory effect of tuberose oil-soluble extract on cortisol, thus reflecting the inhibitory effect of tuberose extract on cortisol release.

[0058] Figure 1 The antioxidant test results of the tuberose extract prepared in Example 1 were obtained by using the tuberose extract prepared in Example 1 as the stock solution and diluting the concentration of the obtained tuberose extract samples to 0.2 wt% and 0.5 wt% respectively for testing. Figure 2 To investigate the effect of different extraction material-to-liquid ratios on the ABTS free radical scavenging rate in Example 1, the concentration of the prepared tuberose extract sample was diluted to 0.2 wt% for testing. Figure 3To investigate the effect of different extraction times on the ABTS free radical scavenging rate in Example 2, the concentration of the prepared tuberose extract sample was diluted to 0.2 wt% for testing. Figure 4 To investigate the effect of different extraction temperatures on the ABTS free radical scavenging rate in Example 3, the concentration of the prepared tuberose extract sample was diluted to 0.2 wt% for testing.

[0059] Figure 5 To investigate the cortisol-inhibiting effect of tuberose extract at different temperatures, the concentration of the prepared tuberose extract samples was diluted to 0.1 wt% for testing. It can be seen that the extract prepared at 40℃ exhibits the best cortisol-inhibiting effect.

[0060] Figure 6 To assess the lipid antioxidant effect of the tuberose extract prepared in Example 1, the concentrations of the obtained tuberose extract samples were adjusted to 0.2 wt% and 0.5 wt% for testing. As the concentration of the tuberose oil-soluble extract increased, its lipid antioxidant effect gradually improved. This indicates that the tuberose oil-soluble extract can be incorporated into cosmetic formulations as a promising innovative ingredient, significantly delaying lipid peroxidation.

[0061] Figure 7 To investigate the effect of the tuberose extract prepared in Example 1 on Hacat cell proliferation, the concentrations of the prepared tuberose extract samples were diluted to 0.005 wt%, 0.01 wt%, 0.025 wt%, 0.05 wt%, 0.1 wt%, 0.2 wt%, 0.25 wt%, and 0.5 wt%, respectively, for testing.

[0062] Figure 8 To determine the cortisol concentration of the tuberose extracts prepared in Examples 1 and Comparative Examples 1-3, the concentration of the prepared tuberose extract samples was diluted to 0.05 wt% for testing. The cortisol inhibition rates in Examples 1, 1, and 2 were 52.1%, 32.9%, and 10.5%, respectively.

[0063] The Bliss Score is calculated based on the Bliss independence model: ΔE = E AB -(E A + E B -E A ·E B The criteria for determining ΔE are: ΔE>0: synergistic effect; ΔE=0: additive effect; ΔE<0: antagonistic effect. AB The inhibition rate of cortisol by the extract of meadowfoam seed oil and caprylic / capric triglycerides in Example 1 is shown in E. A E represents the inhibition rate of cortisol by the extract of meadowfoam seed oil as solvent in Comparative Example 1.B The inhibition rate of cortisol by the extract of Comparative Example 2, using caprylic / capric triglyceride as solvent, was determined. Figure 8 The experimental results showed that ΔE = 0.12 > 0, indicating that the combination of meadowfoam seed oil and caprylic / capric triglycerides as a mixed solvent had a synergistic effect on inhibiting cortisol synthesis.

[0064] It should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit it. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention, and all such modifications or substitutions should be covered within the scope of the claims of the present invention.

Claims

1. A method of preparing extract of tuberose, characterized by: The method comprises the following steps, (1) Pretreatment: drying and crushing the fresh flower of Tuberose to obtain dry flower powder; (2) Extracting oil-soluble extract: adding the pretreated dry flower powder into a solvent and heating extraction at 30-55 ℃ for 1-4 hours; the solvent is white pool flower seed oil and caprylic acid / capric acid triglyceride mixture; (3) Centrifuging, collecting supernatant, filtering, collecting liquid phase to obtain the Tuberose extract; In step (2), the volume ratio of white pool flower seed oil and caprylic acid / capric acid triglyceride is 2.5-4:

1.

2. The preparation method of the extract of tuberose according to claim 1, characterized in that: In step (2), the solid-liquid ratio of dry flower powder and solvent is 1:(4-9) g / mL.

3. The preparation method of the extract of tuberose according to claim 1 or 2, characterized in that: In step (2), the solid-liquid ratio of dry flower powder and solvent is 1:(6-7) g / mL.

4. The preparation method of the extract of tuberose according to claim 1 or 2, characterized in that: In step (2), the extraction temperature is 40-50 ℃.

5. The preparation method of the extract of Polyscias fruticosa according to claim 1 or 2, characterized in that: In step (1), the crushing is crushing the fresh flower of Tuberose to 50-500 mesh.

6. The preparation method of the extract of the genus Polyspora according to claim 1 or 2, characterized by: In step (3), the centrifuging comprises 1000-5000 rpm centrifuging for 5-20 min, and the filtering comprises 0.22-0.45 µm filter membrane filtration under reduced pressure.

7. The Tuberose extract prepared by the preparation method of Tuberose extract according to claim 1.

8. The application of the Tuberose extract according to claim 7 in preparing a product for inhibiting cortisol.

9. The application of the Tuberose extract according to claim 7 in preparing cosmetics.

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