Anti-saccharification composition and external skin care product containing same
By using a combination of Portuguese Corynebacterium P8 fermentation extract and erythritol, the expression of DDOST in skin cells was enhanced, solving the problem of skin glycation and achieving a significant anti-glycation effect.
Patent Information
- Application Number
- CN202511502996.0
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-10-17
- Publication Date
- 2026-01-30
AI Technical Summary
The lack of effective methods to enhance DDOST expression levels in existing technologies makes it difficult to effectively address skin glycation problems.
A combination of fermentation extract of Portuguese Corynebacterium P8 and erythritol was used to prepare an anti-glycation composition and a DDOST expression promoter, thereby increasing the expression level of DDOST in skin cells.
It significantly promoted the expression of DDOST in skin cells, achieving an anti-glycation effect and enhancing the skin's anti-glycation ability.
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Figure CN121421886A_ABST
Abstract
Description
Technical Field
[0001] Embodiments of this disclosure relate to the field of cosmetic technology, specifically to anti-glycation compositions and topical skin care products containing the same. Background Technology
[0002] Glycation refers to the non-enzymatic reaction between reducing sugars (such as glucose and fructose) and proteins, lipids, or nucleic acids, forming advanced glycation end products (AGEs). In the skin, glycation primarily affects collagen and elastin in the dermis and is one of the key factors leading to skin aging, dullness, and disease.
[0003] DDOST (Dolichyl-diphosphooligosaccharide-protein glycosyltransferase) is the core catalytic subunit of the oligosaccharide transferase complex in the endoplasmic reticulum. By maintaining normal protein folding and function, DDOST effectively reduces the abnormal accumulation of advanced glycosylation products (AGEs), achieving anti-glycation at the molecular level.
[0004] Existing technologies have investigated *Corynebacterium pachycarpa*, its fermentation filtrate, its preparation methods, and its applications, exploring its antioxidant, anti-aging, and damage repair properties. However, no studies have yet found demonstrating the use of *Corynebacterium pachycarpa* to enhance DDOST expression levels, making this a promising area for further research.
[0005] The information disclosed in this background section is only intended to enhance the understanding of the background of the inventive concept, and therefore may contain information that does not form prior art known to those skilled in the art. Summary of the Invention
[0006] The summary portion of this disclosure is intended to provide a brief overview of the concepts, which will be described in detail in the detailed description portion. This summary portion is not intended to identify key or essential features of the claimed technical solutions, nor is it intended to limit the scope of the claimed technical solutions.
[0007] Some embodiments of this disclosure provide anti-glycation compositions and topical skin care products containing the same to address one or more of the technical problems mentioned in the background section above.
[0008] In a first aspect, some embodiments of this disclosure provide an anti-glycation composition comprising a fermentation extract of *Clavisporalusitaniae* P8, which is classified as *Clavisporalusitaniae* P8 and is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC NO. 31344.
[0009] Secondly, some embodiments of this disclosure provide an anti-glycation composition comprising a fermentation extract of *Clavispora lusitaniae* P8 and erythritol, wherein the mass ratio of the fermentation extract of *Clavispora lusitaniae* P8 to the erythritol is 1-50:1. *Clavispora lusitaniae* P8 is classified as *Clavispora lusitaniae* P8 and is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC NO.31344.
[0010] Optionally, the mass ratio of the fermentation extract of *Corynebacterium lucida* P8 described in the second aspect above to the erythritol can be 10-20:1.
[0011] Thirdly, some embodiments of this disclosure provide the use of a composition in the preparation of a topical skin agent with anti-glycation properties, wherein the composition is an anti-glycation composition as described in the second aspect above.
[0012] Fourthly, some embodiments of this disclosure provide the application of a microbial strain in the preparation of cosmetics with anti-glycation properties. The microbial strain is *Clavispora lusitaniae* P8, which is classified as *Clavispora lusitaniae* P8 and is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC NO.31344.
[0013] Fifthly, some embodiments of this disclosure provide the use of a composition in the preparation of a DDOST expression promoter, the composition comprising a fermentation extract of Corynebacterium lucida P8.
[0014] Sixthly, some embodiments of this disclosure provide the use of a composition in the preparation of a DDOST expression promoter, the composition comprising a fermentation extract of *Corynebacterium lucida* P8 and erythritol, wherein the mass ratio of the fermentation extract of *Corynebacterium lucida* P8 to erythritol is 1 to 50:1.
[0015] Optionally, in the application of the composition described in the sixth aspect above in the preparation of DDOST expression promoter, the mass ratio of the fermentation extract of *Corynebacterium tumefaciens* P8 to erythritol is 10-20:1.
[0016] In a seventh aspect, some embodiments of this disclosure provide a topical skin care product comprising a composition as described in the first aspect above or a composition as described in the second aspect above.
[0017] Optionally, some embodiments of this disclosure provide topical skin care products including one or more of the following: aqueous solutions, emulsions, creams, ointments, gels, cleansers, and masks.
[0018] The above-described embodiments of this disclosure have the following technical effects: This disclosure verifies the promoting effect of *Corynebacterium tumefaciens* P8 fermentation broth on DDOST expression through Experiment I [detecting the survival rate of human immortalized keratinocytes treated with the following experimental groups] and Experiment II [detecting the expression level of DDOST in human immortalized keratinocytes treated with the following experimental groups]. The addition of *Corynebacterium tumefaciens* P8 fermentation broth significantly increases DDOST expression in glycosylated human immortalized keratinocytes, achieving an anti-glycation effect. Further verification shows that when the mass ratio of *Corynebacterium tumefaciens* P8 fermentation broth to erythritol is 1~50:1, it can also significantly promote the increase in DDOST expression levels, exhibiting a strong anti-glycation effect. In addition, verification shows that when the mass ratio of *Corynebacterium tumefaciens* P8 fermentation broth to erythritol is 10~20:1, the promoting effect on DDOST expression is further enhanced, and the expression of DDOST in human immortalized keratinocytes is further increased, achieving a more significant anti-glycation effect. Attached Figure Description
[0019] Figure 1 This is a schematic diagram illustrating the effect of different concentrations of P8 fermentation broth on cell viability. Figure 2 This is a schematic diagram illustrating the effect of different concentrations of erythritol on cell viability. Figure 3 The effects of samples 4-1, 4-3, and 4-4 on the DDOST expression growth rate were investigated. Figure 4 The effects of samples 4-2, 4-3, and 4-5 on the DDOST expression growth rate were investigated. Figure 5 The effects of samples 4-1, 4-6, and 4-7 on the DDOST expression growth rate were investigated. Figure 6 The effects of samples 4-1, 4-9, and 4-10 on the DDOST expression growth rate were investigated. Figure 7 The effects of samples 4-2, 4-6, and 4-8 on the DDOST expression growth rate were investigated. Figure 8The effects of samples 4-2, 4-9, and 4-11 on the DDOST expression growth rate were investigated. Detailed Implementation
[0020] Embodiments of this disclosure will now be described in more detail with reference to the accompanying drawings. While some embodiments of this disclosure are shown in the drawings, it should be understood that this disclosure can be implemented in various forms and should not be construed as limited to the embodiments set forth herein. Rather, these embodiments are provided to provide a more thorough and complete understanding of this disclosure. It should be understood that the accompanying drawings and embodiments of this disclosure are for illustrative purposes only and are not intended to limit the scope of protection of this disclosure.
[0021] Unless otherwise stated, all technical and scientific terms used herein have the meanings as commonly understood by one of ordinary skill in the art. Generally, the terminology used herein is well-known and conventionally used in the art. When a quantity, concentration, or other value or parameter is expressed as a range, preferred range, or a range defined by a series of upper and lower preferred values, it should be understood as specifically disclosing all ranges formed by any pairing of any upper limit or upper preferred value with any lower limit or right-hand suspending of the lower limit. Whether or not a range is disclosed individually, when a numerical range is described herein, unless otherwise stated, the range should include its endpoints and all integers and fractions within that range.
[0022] It should also be noted that, for ease of description, only the parts relevant to the invention are shown in the accompanying drawings. Unless otherwise specified, the embodiments and features described in this disclosure can be combined with each other.
[0023] It should be noted that the terms "a" and "a plurality of" used in this disclosure are illustrative rather than restrictive, and those skilled in the art should understand that, unless otherwise expressly indicated in the context, they should be understood as "one or more".
[0024] The main terms used in this disclosure are defined as follows: The term "Clavispora lusitaniae P8" is classified as Clavispora lusitaniae P8 and is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC NO.31344.
[0025] The term "DDOST gene" refers to: Dolichyl-diphosphhooligosaccharide-protein glycosyltransferase (Dolichyl-diphosphhooligosaccharide-protein glycosyltransferase) is a key enzyme that catalyzes the initiation of protein N-glycosylation. Boosting its expression can achieve anti-glycation in two ways: first, it accelerates the transfer of pre-synthesized oligosaccharide chains from the endoplasmic reticulum to specific sites on nascent proteins, reducing non-enzymatic reactions between free proteins and sugars, and decreasing the raw materials for AGEs; second, it promotes correct protein folding, reduces the accumulation of misfolded proteins, and prevents them from abnormally binding with sugars to form AGEs, thereby mitigating AGE damage to cells.
[0026] The term "MGO" refers to methylglyoxal. MGO is one of the strongest precursors for the formation of AGEs. It can rapidly react with proteins, lipids, and DNA within cells to form irreversible AGEs (advanced glycosylation products). Therefore, MGO is often used to simulate the formation of advanced AGEs and to establish cellular glycosylation models.
[0027] The term "MGO solution" refers to a solution with a final concentration of 500 micromoles per liter (μmol / L) obtained by mixing 100 μL of DMEM medium solution containing 10% FBS and 1% penicillin / streptomycin with MGO (methylglyoxal).
[0028] The term "DMEM" refers to: Dulbecco's Modified Eagle Medium is a widely used basal medium for cell culture. It is rich in nutrients and maintains the pH balance of cells, making it suitable for the culture of various cell types and applicable in a wide range of fields.
[0029] The term "skin" includes the skin on the face, scalp, neck, chest, abdomen, back, arms, armpits, hands, and legs.
[0030] The term "erythritol" refers to a naturally occurring sugar alcohol. Erythritol has strong hygroscopic and water-locking abilities, actively absorbing moisture from the environment and reducing skin moisture loss (transdermal water loss, TEWL).
[0031] In a first aspect, the present invention provides an anti-glycation composition comprising a fermentation extract of *Clavispora lusitaniae* P8, which is classified as *Clavispora lusitaniae* P8. It is deposited at the China General Microbiological Culture Collection Center (CGMCC) under accession number CGMCC NO. 31344.
[0032] In a second aspect, the present invention provides an anti-glycation composition comprising a fermentation extract of *Clavispora lusitaniae* P8 and erythritol, wherein the mass ratio of the fermentation extract of *Clavispora lusitaniae* P8 to the erythritol is 1-50:1. *Clavispora lusitaniae* P8 is classified as *Clavispora lusitaniae* P8 and is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC NO.31344.
[0033] Optionally, the mass ratio of the fermentation extract of *Corynebacterium tumefaciens* P8 to the erythritol can be 10-20:1.
[0034] In a third aspect, the present invention provides the use of a composition in the preparation of a topical skin agent having anti-glycation properties, wherein the composition is an anti-glycation composition as described in the second aspect above.
[0035] In a fourth aspect, the present invention provides the application of a microbial strain in the preparation of cosmetics with anti-glycation properties, wherein the microbial strain is Clavispora lusitaniae P8, which is classified as Clavispora lusitaniae P8 and is deposited at the China General Microbiological Culture Collection Center with accession number CGMCC NO.31344.
[0036] In a fifth aspect, the present invention provides the use of a composition in the preparation of a DDOST expression promoter, said composition comprising a fermentation extract of Corynebacterium lucida P8.
[0037] In a sixth aspect, the present invention provides the use of a composition in the preparation of a DDOST expression promoter, the composition comprising a fermentation extract of *Clavisporalusitaniae* P8 and erythritol, wherein the mass ratio of the fermentation extract of *Clavisporalusitaniae* P8 to erythritol is 1-50:1. *Clavisporalusitaniae* P8 is classified as *Clavisporalusitaniae* P8. The preservation number of *Clavisporalusitaniae* P8 is CGMCC NO. 31344.
[0038] Optionally, the mass ratio of the fermentation extract of *Cercospora lucida* P8 to erythritol can be 10-20:1, 10-15:1, 15-20:1, 10:1, or 20:1.
[0039] In a seventh aspect, the present invention provides a topical skin care product comprising a composition as described in the first aspect above or a composition as described in the second aspect above.
[0040] Optionally, the mass fraction of the fermentation extract of Corynebacterium lucida P8 in the topical skin care product is 1-80%.
[0041] Optionally, the erythritol in the topical skin care product has a mass fraction of 0.1-8%.
[0042] Optionally, the fermentation extract of Corynebacterium tumefaciens P8 in the topical skin care product has a mass fraction of 1-80%, and the erythritol has a mass fraction of 0.1-8%.
[0043] Optionally, some embodiments of this disclosure provide topical skin care products including one or more of the following: aqueous solutions, emulsions, creams, ointments, gels, cleansers, and masks.
[0044] Topical skin care products can refer to skin care products that are applied to the skin surface through methods such as smearing or covering.
[0045] Optionally, topical skin care products include one or more of the following: aqueous solutions, emulsions, creams, ointments, gels, cleansers, and masks. In practice, aqueous solutions may include one or more of skin care waters, toners, and serums. Emulsions may include lotions. Creams may include face creams. Face creams may be transparent or opaque. Ointments may include skin care creams. Cleansers may include one or more of cleansing foams, cleansing waters, and cleansing liquids. When the topical skin care product is a mask, a commonly used substrate and the skin care composition of this disclosure applied or impregnated onto the substrate may be used. The substrate includes, but is not limited to, silk mask sheets, cotton mask sheets, polyester mask sheets, or blended mask sheets. The selection of the substrate can be made by those skilled in the art according to actual needs. The forms of the above-mentioned masks may include, but are not limited to, gel masks, cream masks, and essential oil masks.
[0046] Alternatively, the screening method for the above-mentioned *Corynebacterium tumefaciens* P8 is as follows: The first step is to crush the grapes and dissolve them in an appropriate amount of water, mixing thoroughly. The amount of water used is not fixed and can be adjusted according to actual needs.
[0047] The second step involves adding 0.1 mL of sample to 0.9 mL of sterile saline to obtain 10... -1 Diluent. It should be noted that the 0.1 mL sample taken was taken from the mixed solution obtained by crushing the grapes and dissolving them in an appropriate amount of water as described in the first step above, and then mixing thoroughly.
[0048] The third step is to aspirate 0.1 mL of 10 -1The diluent was diluted in 0.9 mL of sterile physiological saline to obtain 1 ²Diluent.
[0049] Step 4: Take 0.1 mL of 1 ² Dilute the solution in 0.9 mL of sterile physiological saline to obtain 1 ³Diluent.
[0050] Step 5, take 0.1 mL of 1 ³ Dilute the solution in 0.9 mL of sterile physiological saline to obtain 1 Diluent.
[0051] Step 6: Take 0.1 mL of 1 The diluent was diluted in 0.9 mL of sterile physiological saline to obtain 1 Diluent.
[0052] Step 7: Take 0.1 mL of 1 The diluent was diluted in 0.9 mL of sterile physiological saline to obtain 1 Diluent.
[0053] Step 8: Take 0.1 mL of 1 The diluted solution was spread evenly on YPD solid medium and then incubated upside down at a temperature of 37°C.
[0054] Step nine involves preliminary identification of the bacterial strains 24 hours later through colony morphology and microscopic examination, followed by selection of strains meeting the specified characteristics for streak plating purification. For example... Figure 1 As shown, the selected strains are round, milky white, and opaque, with a diameter of approximately 1 nm to 2 nm. They have smooth edges, a moist and glossy surface, and appear oval under a microscope. The cells are larger than bacteria.
[0055] Step 10: Select a single colony from the last streak purification plate and inoculate it onto 10 mL of liquid culture medium, then incubate for 24 hours. The liquid culture medium is YPD liquid medium.
[0056] Step 11: After the culture is completed, take 1 mL to 2 mL of the bacterial culture after 24 hours of culture and 40% sterile glycerol into a culture preservation tube and store it in a -80℃ refrigerator.
[0057] Step 12: Centrifuge the bacterial culture after 24 hours of cultivation, discard the supernatant, and use this as the first bacterial culture. The centrifugation speed is 6000 r / min, and the centrifugation time is 2 min.
[0058] Step 13: Add 1.5 mL of sterile water to the first bacterial culture, mix thoroughly, and centrifuge. Discard the supernatant to obtain the second bacterial culture. The centrifugation speed is 6000 r / min, and the centrifugation time is 3 min.
[0059] Step fourteen: Add 1.5 mL of sterile water to the second bacterial culture, mix thoroughly, centrifuge, discard the supernatant, and use this as the third bacterial culture. The centrifugation speed is 6000 r / min, and the centrifugation time is 3 min.
[0060] Step 15: Resuspend the third bacterial culture in 1.5 mL of sterile water and mix thoroughly. Use this mixture as a template for bacterial identification and perform PCR (Polymerase chain reaction).
[0061] Step sixteen involves sequencing the PCR products to obtain the sequencing results. The PCR products are then sent to Sangon Biotech (Shanghai) Co., Ltd. for sequencing.
[0062] Step seventeen: After assembling the sequencing results, homology analysis was performed in the NCBI (National Center for Biotechnology Information) database using BLAST (Basic Local Alignment Search Tool). Sequences with ≥98% sequence identity were selected for comparison to obtain the alignment results. Based on the comparison results, the strain was identified as *Corynebacterium tumefaciens* P8.
[0063] The present disclosure will now be described in detail with reference to embodiments.
[0064] Experiment I [Detecting the survival rate of immortalized human keratinocytes treated with the following experimental groups]
[0065] Cell viability can be detected using methods well-known in the field, such as the CCK8 method (Cell Counting Kit-8). The CCK8 method can be performed according to the experimental methods described in "Experimental Techniques in Molecular Biology", "Guide to Molecular Experimental Techniques", "Experimental Tutorial in Cell Biology", or "Experiments in Biochemistry and Molecular Biology".
[0066] Experimental group information: Sample 1-1: Sample 1-1 is a cell culture medium containing P8 fermentation filtrate, wherein the mass concentration of P8 fermentation filtrate is 1%.
[0067] The preparation method for Sample 1-1 is as follows: Mix the P8 fermentation filtrate and cell culture medium to obtain Sample 1-1.
[0068] Samples 1-2 to 1-5: Samples 1-2 to 1-5 are basically the same as sample 1-1, except for the concentration of P8 fermentation filtrate. Therefore, the preparation method can be implemented with reference to sample 1-1.
[0069] The mass concentration of the P8 fermentation filtrate in samples 1-2 was 0.5%; The mass concentration of the P8 fermentation filtrate in samples 1-3 was 0.25%; The mass concentration of the P8 fermentation filtrate in samples 1-4 was 0.13%; The mass concentration of P8 fermentation filtrate in samples 1-5 was 0.06%.
[0070] Sample 2-1: Sample 2-1 is a cell culture medium containing erythritol, wherein the mass concentration of erythritol is 0.25%.
[0071] The preparation method for sample 2-1 is as follows: Mix erythritol and cell culture medium to obtain sample 2-1.
[0072] Samples 2-2 to 2-5: Samples 2-2 to 2-5 are basically the same as sample 2-1, except for the concentration of erythritol. Therefore, the preparation method can be carried out with reference to sample 2-1.
[0073] The mass concentration of erythritol in sample 2-2 was 0.13%; The mass concentration of erythritol in samples 2-3 was 0.06%; The mass concentration of erythritol in samples 2-4 was 0.03%; The mass concentration of erythritol in samples 2-5 was 0.013%.
[0074] Test Results
[0075] In HaCaT cells, neither 0–1% P8 fermentation broth nor 0–0.13% erythritol showed cytotoxicity. Therefore, DDOST expression experiments were conducted using P8 fermentation filtrate at concentrations of 0.1% and 0.2%. Furthermore, erythritol was combined at concentrations of 0.01% and 0.02% to verify the effect of the combination on DDOST expression. Figure 1 As shown, Figure 1 The diagram illustrates the effect of different concentrations of P8 fermentation broth on cell viability. Figure 1 It is known that high concentrations of P8 fermentation broth may inhibit cell survival, while low concentrations have little or no effect on cell survival, or even no significant inhibition. From Figure 2 As shown, Figure 2The diagram illustrates the effect of different concentrations of erythritol on cell viability. Figure 2 It is known that high concentrations of erythritol may inhibit cell survival, while low concentrations have little or no effect on cell survival. When the concentration of erythritol is 0.25%, it has a significant impact on cell survival. Therefore, setting the concentration of erythritol to 0-0.13% is more appropriate.
[0076] Experiment II [Detection of DDOST expression levels in human immortalized keratinocytes treated with the following experimental groups].
[0077] The expression level of DDOST in cells can be detected using methods well-known in the field, such as qPCR. The qPCR method can be performed according to the experimental procedures described in "Experimental Techniques in Molecular Biology," "Guide to Molecular Experimental Techniques," "Experimental Tutorial in Cell Biology," or "Experiments in Biochemistry and Molecular Biology."
[0078] Control: DMEM medium solution containing 10% FBS and 1% penicillin / streptomycin.
[0079] MGO group: MGO (methylglyoxal) was mixed with DMEM medium solution containing 10% FBS and 1% penicillin / streptomycin; the concentration of MGO was 500 μmol / L.
[0080] The preparation method for sample 4-1 is as follows: The P8 fermentation broth, MGO, and DMEM medium solution containing 10% FBS and 1% penicillin / streptomycin were mixed to obtain the prepared sample 4-1; the mass concentration of the P8 fermentation broth was 0.1%, and the concentration of MGO was 500 μmol / L.
[0081] The preparation method for sample 4-2 is as follows: The P8 fermentation broth, MGO, and DMEM medium solution containing 10% FBS and 1% penicillin / streptomycin were mixed to obtain sample 4-2; the mass concentration of the P8 fermentation broth was 0.2%, and the concentration of MGO was 500 μmol / L.
[0082] The preparation method for sample 4-3 is as follows: Erythritol, MGO, and DMEM medium solution containing 10% FBS and 1% penicillin / streptomycin were mixed to obtain sample 4-3; wherein the mass concentration of erythritol was 0.01% and the concentration of MGO was 500 μmol / L.
[0083] The preparation method for sample 4-4 is as follows: The P8 fermentation broth, erythritol, MGO, and DMEM medium solution containing 10% FBS and 1% penicillin / streptomycin were mixed to obtain sample 4-4; wherein the mass concentration of P8 fermentation broth was 0.1%, the mass concentration of erythritol was 0.01%, and the concentration of MGO was 500 μmol / L.
[0084] The preparation methods for samples 4-5 are as follows: The P8 fermentation broth, erythritol, MGO, and DMEM medium solution containing 10% FBS and 1% penicillin / streptomycin were mixed to obtain preparation 4-5; wherein the mass concentration of P8 fermentation broth was 0.2%, the mass concentration of erythritol was 0.01%, and the concentration of MGO was 500 μmol / L.
[0085] The preparation methods for samples 4-6 are as follows: Erythritol, MGO, and DMEM medium solution containing 10% FBS and 1% penicillin / streptomycin were mixed to obtain preparation 4-6; the mass concentration of erythritol was 0.03%, and the concentration of MGO was 500 μmol / L.
[0086] The preparation methods for samples 4-7 are as follows: The P8 fermentation broth, erythritol, MGO, and DMEM medium solution containing 10% FBS and 1% penicillin / streptomycin were mixed to obtain the prepared samples 4-7; the mass concentration of P8 fermentation broth was 0.1%, the mass concentration of erythritol was 0.03%, and the concentration of MGO was 500 μmol / L.
[0087] The preparation methods for samples 4-8 are as follows: The P8 fermentation broth, erythritol, MGO, and DMEM medium solution containing 10% FBS and 1% penicillin / streptomycin were mixed to obtain the prepared samples 4-8; the mass concentration of P8 fermentation broth was 0.2%, the mass concentration of erythritol was 0.03%, and the concentration of MGO was 500 μmol / L.
[0088] The preparation methods for samples 4-9 are as follows: Erythritol, MGO, and DMEM medium solution containing 10% FBS and 1% penicillin / streptomycin were mixed to obtain the prepared samples 4-9; the mass concentration of erythritol was 0.004%, and the concentration of MGO was 500 μmol / L.
[0089] The preparation method for sample 4-10 is as follows: The P8 fermentation broth, erythritol, MGO, and DMEM medium solution containing 10% FBS and 1% penicillin / streptomycin were mixed to obtain the prepared sample 4-10; the mass concentration of P8 fermentation broth was 0.1%, the mass concentration of erythritol was 0.004%, and the concentration of MGO was 500 μmol / L.
[0090] The preparation method for sample 4-11 is as follows: The P8 fermentation broth, erythritol, MGO, and DMEM medium solution containing 10% FBS and 1% penicillin / streptomycin were mixed to obtain sample 4-11. The mass concentration of P8 fermentation broth was 0.2%, the mass concentration of erythritol was 0.004%, and the concentration of MGO was 500 μmol / L.
[0091] Test results: Table 1 Sample Test Results
[0092] The test results are as follows: In HaCaT cells, 500µM MGO induction significantly reduced the transcriptional level of DDOST, and the cellular glycosylation model was successfully established.
[0093] As shown in Table 1, Figure 3 and Figure 4 As shown, using 0.1% P8 fermentation broth alone significantly promoted DDOST expression, with an increase of 212.55% compared to the MGO model group. Using 0.2% P8 fermentation broth alone also significantly promoted DDOST expression, with an increase of 194.15% compared to the MGO model group. This indicates that P8 fermentation broth has a significant promoting effect on DDOST expression in human immortalized keratinocytes and exhibits an anti-glycation effect.
[0094] As shown in Table 1, Figure 3-8As shown, the combined use of 0.1% P8 fermentation broth + 0.01% erythritol significantly promoted DDOST levels, with an increase of 244.71% compared to the MGO model group; the combined use of 0.2% P8 fermentation broth + 0.01% erythritol significantly promoted DDOST levels, with an increase of 245.82% compared to the MGO model group; the combined use of 0.1% P8 fermentation broth + 0.03% erythritol significantly promoted DDOST levels, with an increase of 193.1% compared to the MGO model group; the combined use of 0.2% P8 fermentation broth + 0.03% erythritol significantly promoted DDOST levels, with an increase of 193.1% compared to the MGO model group; the combined use of 0.2% P8 fermentation broth + 0.03% erythritol significantly promoted DDOST levels. Erythritol significantly promoted DDOST levels, with an increase of 191.7% compared to the MGO model group; the combined use of 0.1% P8 fermentation broth + 0.004% erythritol significantly promoted DDOST levels, with an increase of 187.47% compared to the MGO model group; the combined use of 0.2% P8 fermentation broth + 0.004% erythritol significantly promoted DDOST levels, with an increase of 192.3% compared to the MGO model group. This indicates that the combination of P8 fermentation broth and erythritol significantly promoted DDOST expression in human immortalized keratinocytes. Specifically, a mass ratio of P8 fermentation broth to erythritol of 1–50:1 significantly promoted DDOST expression in human immortalized keratinocytes, demonstrating an anti-glycation effect.
[0095] like Figure 3 As shown, 0.01% erythritol alone had no effect on promoting DDOST levels. The combined use of 0.1% P8 fermentation broth and 0.01% erythritol significantly promoted DDOST levels, with an increase of 244.71% compared to the MGO model group, demonstrating a significantly higher anti-glycation effect than using 0.1% P8 fermentation broth alone. Figure 4 As shown, the combined use of 0.2% P8 fermentation broth and 0.01% erythritol significantly promoted DDOST levels, with an increase of 245.82% compared to the MGO model group, demonstrating a significantly higher anti-glycation effect than using 0.2% P8 fermentation broth alone. This further demonstrates that a mass ratio of P8 fermentation broth to erythritol of 10–20:1 produces a synergistic effect, further promoting DDOST expression levels and achieving a stronger anti-glycation effect.
[0096] The applications of the prepared materials are as follows: 1. The preparation steps of a water-based skin care product are as follows: First, weigh out water (add water to make all ingredients reach 100% by mass) and 0.1% Carbopol U20 by mass and add them to the emulsifying pot in sequence. After stirring thoroughly until there are no particles or lumps, heat the pot to 80-90℃.
[0097] The second step involves adding 6.0% glycerol, 1.0% panthenol, and 0.02% EDTA-2NA to the emulsification pot in sequence, stirring at a uniform speed until a clear solution is obtained, and then cooling to 50-60℃.
[0098] Third, in another container, heat 0.3% styraxone, 0.5% hexanediol, and 0.8% pentanediol to 70-85°C until dissolved into a clear, particle-free solution. Then, cool the solution to 50°C and add it to the emulsification pot mentioned above.
[0099] Fourth step: Take another container and stir 0.08% arginine and 2% water until it is clear and free of particles, then add it to the emulsification pot mentioned above.
[0100] Fifth, after the emulsifying pot cools to 40-45°C, add 0.5% methyl glucetol polyether-20 and 4.8% of the composition by mass, and stir until homogeneous. It should be noted that the 4.8% composition by mass can characterize the composition described in the first aspect or the composition described in the second aspect above.
[0101] Step 6: Take samples for testing; discharge the material after it passes the test.
[0102] 2. The preparation steps of a water-in-oil type skin care product are as follows: First, add water (add water to make the total mass of all ingredients 100%) and Carbopol 940 (0.25% by mass) to the emulsifying pot in the correct amount. After stirring thoroughly until there are no particles or lumps, heat the pot to 80-90℃.
[0103] The second step involves adding 5.0% butanediol and 1.5% betaine to the emulsifying pot and stirring at a constant speed until a transparent solution is obtained. The mixture is then kept warm and stirred.
[0104] The third step involves adding 1.5% (by mass) of A170, 1.0% (by mass) of Olivem 1000, 4.0% (by mass) of GTCC, 1.0% (by mass) of C16 / 18 alcohol, and 2.7% (by mass) of ININ to an oil pan, heating it to 80-90°C, and stirring it thoroughly to obtain the oil phase.
[0105] The fourth step is to add the treated oil phase to the emulsification pot mentioned above, homogenize for 3-5 minutes, and stir to cool to 45-50℃.
[0106] Fifth, in another container, mix 3% water and 0.2% arginine until the mixture is clear and free of particles, then add it to the emulsification pot mentioned above.
[0107] Step 6: In another container, heat 1.1% pentanediol, 0.6% hexanediol, and 0.5% styrene to 70-85°C until dissolved into a clear, particle-free solution. Then, cool the solution to 40-50°C and add it to the emulsification pot.
[0108] Step 7: After the emulsifying pot cools to 40-45°C, add 0.5% (by mass) of Deminshuhe and 13% (by mass) of the composition to the emulsifying pot, and stir until homogeneous. It should be noted that the 13% (by mass) composition can represent the composition described in the first aspect or the composition described in the second aspect above.
[0109] Step 8: Sampling and testing; discharge the material after it passes the test.
[0110] The embodiments disclosed herein have the following technical effects: This disclosure verifies, through Experiment I [detecting the survival rate of human immortalized keratinocytes treated with the following experimental groups] and Experiment II [detecting the expression level of DDOST in human immortalized keratinocytes treated with the following experimental groups], that the use of P8 fermentation broth alone promotes DDOST expression, and verifies that when the mass ratio of P8 fermentation broth to erythritol is 10-20:1, it can further promote the increase of DDOST expression level, exhibiting a stronger anti-glycation effect. Furthermore, it verifies that in HaCaT cells, 0-1% P8 fermentation broth, 0-0.13% erythritol, and their combination have no cytotoxicity.
[0111] The above description is merely a selection of preferred embodiments of this disclosure and an explanation of the technical principles employed. Those skilled in the art should understand that the scope of the invention involved in the embodiments of this disclosure is not limited to technical solutions formed by specific combinations of the above-described technical features, but should also cover other technical solutions formed by arbitrary combinations of the above-described technical features or their equivalents without departing from the above-described inventive concept. For example, technical solutions formed by substituting the above-described features with (but not limited to) technical features with similar functions disclosed in the embodiments of this disclosure.
Claims
1. An anti-glycation composition comprising a fermentation extract of Clavispora lusitaniae P8, which is classified as Clavispora lusitaniae P8 and deposited with the China General Microbiological Culture Collection Center under the accession number CGMCC NO.31344. 2.An anti-glycation composition comprising a fermentation extract of Clavispora lusitaniae P8 and erythritol, wherein the mass ratio of the fermentation extract of Clavispora lusitaniae P8 to the erythritol is 1-50:1, which is classified as Clavispora lusitaniae P8 and deposited with the China General Microbiological Culture Collection Center under the accession number CGMCC NO.31344. 3.The anti-glycation composition of claim 2, wherein the mass ratio of the fermentation extract of Clavispora lusitaniae P8 to the erythritol is 10-20:
1. 4.Use of a composition in the preparation of a skin external agent for anti-glycation, wherein the composition is the anti-glycation composition of claim 2. 5.Use of a microbial strain in the preparation of a cosmetic for anti-glycation, wherein the microbial strain is Clavispora lusitaniae P8, which is classified as Clavispora lusitaniae P8 and deposited with the China General Microbiological Culture Collection Center under the accession number CGMCC NO.31344. 6.Use of a composition in the preparation of a DDOST expression promoter, wherein the composition comprises a fermentation extract of Clavispora lusitaniae P8. 7.Use of a composition in the preparation of a DDOST expression promoter, wherein the composition comprises a fermentation extract of Clavispora lusitaniae P8 and erythritol, and the mass ratio of the fermentation extract of Clavispora lusitaniae P8 to the erythritol is 1-50:
1. 8.The use of a composition in the preparation of a DDOST expression promoter of claim 7, wherein the mass ratio of the fermentation extract of Clavispora lusitaniae P8 to the erythritol is 10-20:
1. 9.An external skin care product comprising the composition of claim 1 or the composition of claim 2.
10. The external skin care product according to claim 9, wherein, The external skin care product comprises one or more of an aqueous agent, an emulsion, a cream, a paste, a gel, a cleanser, and a mask.