Study On Preparation Technology And Anti-Aging Effect Of Gastrodia Elata Juice Granule

[Purpose] Fresh Gastrodia elata was studied in this paper.The forming process of Gastrodia juice granules is optimized,and its quality evaluation standard is formulated.Network pharmacology was used to predict the antiaging active components,action targets and related signaling pathways of Gastrodia elata juice..The possible mechanism of action was analyzed by verifying its antioxidant activity and the main active ingredients in Gastrodia juice were verified using Caenorhabditis elegans experiments.This study provide theoretical basis for the development of Gastrodia elata related products and systematic research on the anti-aging effect of Gastrodia elata juice.[Methods] 1.Study on the preparation technology,quality analysis research of Gastrodia juice granules: Spray drying and freeze drying,were selected for research,and the most suitable drying process for Gastrodia elata juice was selected,and its formula was optimized.The optimum preparation process was determined by analyzing the type and ratios of thinner and concentration of wetting agent based on the physical and chemical indicators such as the comprehensive scores of the formability,solubility,hygroscopicity and repose angle of the produced granules.Thin-layer chromatography（TLC）and high-performance liquid chromatography（HPLC）technologies were applied to formulate the basic quality assessment indexes of GJG.2.Analysis of anti-aging mechanism of Gastrodia juice based on network pharmacology: UPLC-MS/MS was used to identify the chemical components in Gastrodia elata juice;The pharmacological targets of the active ingredients in Gastrodia elata juice were obtained by Swiss Target Prediction.The aging related targets were obtained by Gene Cards and HAGR database.The PPI network of common target between active ingredient and aging was analyzed by STRING database then use Cytoscape software to screen and analyze its network nodes.The obtained targets were enriched with GO and KEGG analysis by DAVID database.The five key targets were molecularly docked with the active components of gastrodia juice using the Auto Dock Vina tool.3.Anti-aging efficacy and mechanism of gastrodia juice and its functional components on Caenorhabditis elegans: The antioxidant activity in vitro and in vivo was verified by DPPH and paraquat.Gastrodia juice（4、8、16 mg/ml）、HBA（50μM）、GAS（100 μM）、GP（500μg/ml）was given to C.elegans,and the lifespan of C.elegans was measured,and the anti-oxidative effect of gastrodia juice was confirmed.Reproductive experiments were conducted to take the number of eggs laid as an indicator of the normal growth state of C.elegans.By detecting the level of reactive oxygen species（ROS）、antioxidant enzyme activity、lipofuscin accumulation、expression of age-1,daf-16,sod-3,skn-1 genes in FOXO signaling pathway in the C.elegans to evaluate the protective effect of gastrodia juice on C.elegans.[Results] 1.Study on the preparation technology,quality analysis research of Gastrodia juice granules: The optimized preparation procedure was obtained: blend the freeze-dried powder of gastrodia juice with soluble starch（1:1,weight ratio）,then add appropriate amount of 80% ethanol to make soft material,followed by granulation,and then the wet granules were dried at 50 °C for 4 hours,the whole dried granules were then discharged from mesh holes by frequency conversion shaking that transform materials into the desired size of granule particles.The granulated products should have good formability,fluidity,and water solubility,with low water content,and moisture absorption rate.The establilshed TLC and HPLC analyzing methods were highly specific with good reproducibility.The HPLC analysis results indicated that the P-hydroxybenzyl alcohol was well linear in the range of 0.51 to 204 μg/m L,r~2=0.999 8,the average recovery rate was 104.67 %,and the RSD value was 1.05 %.2.Analysis of anti-aging mechanism of Gastrodia juice based on network pharmacology: Ten chemical constituents in gastrodia juice were analyzed by UPLC-MS/MS.Nameed gastrodin、 p-hydroxybenzyl alcohol、p-hydroxybenzoic acid、 p-hydroxybenzaldehyde、 and 3,4-Dihydroxybenzaldehyde、succinic acid、 parishin A、parishin B、parishin C、and parishin E.Forecast results show there are 22 targets in total,and the top 5 key targets are STAT3,ESR1,SRC,MTOR,CCND1;It is predicted that the anti-aging effect of gastrodia juice mainly affects FOXO,PI3K-Akt,insulin,m TOR and other signaling pathways.The five key targets were molecularly docked with the active components of gastrodia juice using the Auto Dock Vina tool,and it was shown that the active components gastrodin and p-hydroxybenzyl alcohol in the gastrodia juice have good binding activity with the key targets.3.Anti-aging efficacy and mechanism of gastrodia juice and its functional components on Caenorhabditis elegans: The experimental results show that Gastrodia juice、 HBA、GAS、GP can significantly prolong the lifespan of C.elegans,enhance the ability of C.elegans to resist oxidative stress,reduce the levels of ROS and lipofuscin,increase the activity of superoxide dismutase（SOD）,and play an anti-aging effect on the FOXO signaling pathway.However,the anti-aging and antioxidant activities of a single compound were weaker than those of Gastrodia juice,indicating that Gastrodia juice exerts anti-aging effects from multiple components,multiple targets,and multiple factors.[Conclusion]1.The optimal preparation process in this experiment is feasible.Gastrodia elata juice is developed into granules with stable,convenient and safe.At the same time,the formalizing process and corresponding quality control standards of Gastrodia elata juice granules are established.2.Using UPLC-MS/MS to analyze the composition of gastrodia juice;Combined with network pharmacology analysis,it was found that the FOXO pathway may be the main pathway for its anti-aging effect.Experiments on Caenorhabditis elegans showed that Gastrodia juice and HBA,GAS,GP in Gastrodia juice can significantly prolong the lifespan of nematodes,achieve the balance between ROS level and antioxidant enzyme activity in nematodes,improve biological characteristics related to aging,and act on FOXO Signaling pathways play an anti-aging effect.