Study On The Photoaging Skin Repair Effect Of Ganoderma Lucidum Glycoprotein And Its Microemulsion Preparation Process

Objective:In this study,Ganoderma lucidum glycoprotein（Gl-Gp）was extracted,purified and identified,and then the repair effect of Gl-Gp on photoaging was initially investigated.Gl-Gp is a macromolecular water-soluble substance,it can be formulated into W/O type microemulsion to increase its transdermal absorption rate.Gl-Gp microemulsion can be added as an active ingredient into a cream to provide a basis for the development of related cosmetic.Methods:（1）The optimal extraction method of Gl-Gp was obtained by single-factor examination combined with BBD method,using the protein yield and polysaccharide yield as index,and its structure was verified byβ-elimination,infrared spectroscopy,anti-O-GlcNAc modification sites,and amino acid analysis;（2）The antioxidant activity of Gl-Gp was investigated using DPPH,ABTS⁺radical scavenging capacity,total reducing capacity and anti-lipid peroxidation capacity as index;（3）Rats were modeled by long-term UV irradiation to compare the appearance and skin thickness of rats,and observe the changes of skin structure and elastic fibers by H&E and aldehyde magenta staining;to investigate the mechanism of Gl-Gp on photoaging repair through the determination of SOD,GSH-Px activity,MDA,HYP content and expression levels of related factors in rat dorsal skin tissue homogenates;（4）The pseudo-ternary phase diagram forming microemulsion area size was used as an index for single-factor examination,and EE and DL were used as index for conducting D-optimal mixture design optimization prescription;and the Gl-Gp microemulsions were also evaluated for appearance,type,pH,transmission electron microscopy,stability and transdermal permeability;（5）Gl-Gp microemulsion was added to the cream as an active ingredient and evaluated for sensory index,moisturizing property,pH,heavy metal and microbial content,and stability.Results:（1）The optimal extraction process of Gl-Gp obtained by single-factor examination combined with BBD method was temperature 71℃,ratio of soild to liquid 1:54g ml^-1,oscillation speed 167 r min^-1,oscillation time of 12 h times^-1,ultrasonic time of 30 min times^-1,extraction 2 times with protein yield of（5.219±0.23）%,polysaccharide yield of（1.519±0.09）%,which contained protein content of（45.56±0.79）%,neutral sugar content of（10.18±0.29）%,and acidic sugar content of（22.69±0.22）%;The Gl-Gp contains O-glycosidic bonds with anti-O-GlcNAc modification sites,and its molecular weight was mainly concentrated at 15-25 kD,and infrared spectra indicated the presence of protein and polysaccharide and other characteristic peaks;amino acid composition results of its content of 18 amino acids.（2）The IC50 values of DPPH and ABTS⁺radical scavenging rate of Gl-Gp were 62.80μg mL^-1 and 0.804 mg mL^-1,respectively,and the total reducing capacity and anti-lipid peroxidation capacity were positively correlated with concentration,indicating that Gl-Gp has strong antioxidant properties.（3）Gl-Gp can repair erythematous and ulcerated skin on the back of rats after UV irradiation,restore tissue structure and elastic fibers to orderly arrangement,repair collagen and elastin,and inhibit the expression of inflammatory factors and tumor necrosis factors.（4）With 15 mg mL^-1 Gl-Gp solution as the aqueous phase,isopropyl myristate as the oil phase,soy phospholipid and anhydrous ethanol as the mixed surfactant（Km=1）,and a mass ratio of 0.260:0.365:0.375,the encapsulation efficiency of Gl-Gp microemulsion obtained was（96.17±0.11）%,and the drug loading coefficient was（3.70±0.07）mg g^-1.It was a clarified and transparent W/O microemulsion with good stability and pH in accordance with human requirements,and the cumulative transdermal rate was（20.98±0.34）%within 36 h.（5）Gl-Gp microemulsion cream was creamy white,good texture,with good moisturizing effect.The heavy metal,microbial and pH tests all be up to the cosmetic standards.Conclusion:The optimal extraction process of Gl-Gp was established;the optimized Gl-Gp can effectively repair the damage of photoaging in rat skin;the prepared microemulsion has stable quality and good transdermal properties.Gl-Gp microemulsion is added to the cream as an active ingredient with good moisturizing capacity and stability.