Effects Of Submerged Fermentation Pretreatment Of Inonotus Obliquus On Extraction Of Effective Components In Eucommia Ulmoides Leaves And Their Antioxidant Capacity

Eucommia ulmoides,whose scientific name is Eucommia ulmoides Oliver,was once an endemic economic plant in China.Many studies have found that Eucommia ulmoides is rich in bioactive components,which make Eucommia ulmoideshave a variety of pharmacological activities,including antioxidant,antibacterial,anti-inflammatory,lipid-lowering,etc.Therefore,Eucommia ulmoides is widely used in the health care industry.However,the extraction efficiency of active components from Eucommia ulmoides is not high,and the main factor hindering the extraction is the lignocellulose in the cell wall of Eucommia ulmoides.The extraction efficiency of traditional method that organic solvent combined with reflux is not high,and acid-alkali treatment is easy to cause environmental pollution,physical methods such as milling have risks and energy loss in the process of operation.In recent years,microbial pretreatment has gradually emerged,which uses fungi or bacteria to produce enzymes to destroy the structure of lignocellulose.However,it usually takes a long time.Therefore,it is of great significance to find a method which can improve the extraction rate of effective components and reduce the loss and processing cycle.Some studies have found that the fermentation of white rot fungi can produce not only cellulose-degrading enzymes,but also a large number of lignin-degrading enzymes,which can effectively degrade lignocellulose.Therefore,the liquid fermentation of Eucommia ulmoides leaves by Inonotus obliquus was studied in this paper,the content of lignocellulose in Eucommia ulmoides leaves was determined by van Soest method,and the contents of extracellular polyphenols,flavonoids,phenolic acids and iridoids in the fermentation system were determined by ultraviolet spectrophotometry.After 2 days of fermentation,we detected the extract of Eucommia ulmoides leaves by HPLC,and the contents of chlorogenic acid and rutin in dynamic fermentation were determined.Finally,we studied the antioxidant activity of Eucommia ulmoides leaf extract.The results of the study are as follows:（1）The addition of Eucommia ulmoides leaves powder provided nutrition for the growth of Inonotus obliquus mycelium,the mycelium biomass of treated group was significantly higher than fermentation control group.With the progress of fermentation,the quality of Eucommia ulmoides leaves decreased continuously,and on day 14 of fermentation,the mass loss reached 45.5%,and the degradation rates of lignin,cellulose and hemicellulose were 42.0%,47.7%and 50.3%.This shows that the pretreatment of Inonotus obliquus has effectively degraded Eucommia ulmoides leaves.（2）The degradation of lignocellulose significantly increased the release of various components in Eucommia ulmoides leaves.The contents of polyphenols（142.56mg GAE/L）and iridoid（32.06 mg/g）in the extracellular fluid of treated group reached the maximum on day 4,while the contents of flavonoids（28.60 mg/L）and phenolic acids（31.53 mg/L）reached the maximum on day 2,so the best fermentation time should be 2-4 days.Considering that on the second day,the contents of flavonoids,phenolic acids and iridoid in the treated group increased the most compared with the aqueous extract control group（56.4%,46.9%and 173.5%）,and only the contents of polyphenols increased significantly on day 4（6.5%）.Finally,it was determined that the best fermentation time was 2 days.（3）The extracellular extract of Eucommia ulmoides leaves by Inonotus obliquus fermentation pretreatment was analyzed and identified by HPLC.It was found that the extract contained about 16 components.By comparing with the liquid phase diagram of the standard product,chlorogenic acid（12.905 min）,rutin（27.062 min）and quercetin（49.955 min）were identified.The contents of chlorogenic acid and rutin（13.28 mg/g and 4.69 mg/g）in the treated group were much higher than those in the aqueous extract control group（2.64 mg/g and 1.18 mg/g）,which increased by 404.30%and 298.50%,which further indicated that the degradation of lignocellulose in Eucommia ulmoides leaves greatly increased the release of active components.（4）The fermentation pretreatment of Eucommia ulmoides leaves by Inonotus obliquus enhances the antioxidant activity of the extract.The scavenging ability of the extracts from the treated group on ABTS free radicals,DPPH free radicals and hydroxyl free radicals(IC₅₀=8.19 mg/mL,3.32 mg/mL,2.50 mg/mL)was stronger than that of the aqueous extract control group(IC₅₀=12.38 mg/mL,22.69 mg/mL,5.02 mg/mL).In this study,the liquid fermentation of Inonotus obliquus was used to pretreat Eucommia ulmoides leaves.The fermentation treatment can effectively degrade the lignocellulose of Eucommia ulmoides leaves within 2 days,thus,a large number of active components in Eucommia ulmoides leaves are released,which greatly shorten the period of microbial pretreatment and reduced the time cost.At the same time,due to the increase in the content of various active ingredients,the antioxidant activity of Eucommia ulmoides leaf extract is also enhanced.