| Kadsura coccinea is an evergreen woody vine wild plant of the Kadsura genusin the Schisandraceae family.In recent years,Kadsura coccinea have attracted much attention due to their diverse biological activities.However,current research on Kadsura coccinea mainly focuses on the biological activity of fruits among different varieties and the exploration of related lignans.There is still a lack of systematic research on the polyphenols in other parts of the Kadsura Therefore,the main focus of this study is on the roots,stems,and leaves of the Kadsura coccinea.Different extraction methods are used to extract polyphenol substances from the three parts of the three varieties,and the polyphenol content measured under different extraction methods is explored,as well as the differences in polyphenol content between different varieties and parts.Research has found that ultrasound extraction has simple steps,short extraction time,and good extraction efficiency.The polyphenol content among different varieties and parts showed the following trend:purple black>pink>bright red,root>stem>leaf.The polyphenol extract content of purple black root extracted by thermal reflux method was the highest,at 40.40 mgGAE/g.At the same time,macroporous resins for polyphenol purification were also screened,and there sults showed that AB-8 had good adsorption and desorption effects on Kadsura coccinea polyphenol extracts.The concentration of polyphenols in the purified roots of the Kadsura coccinea variety purple black is 126.94±0.17 μgGAE/g,the concentration increased by 40.19%;The concentration of polyphenols purified from purple black stems is 58.53±0.13 μgGAE/g,the concentration increased by 37.93%;The concentration of polyphenols purified from purple black leaves is 29.00±0.22 μgGAE/g,the concentration increased by 42.03%;The purity of purified polyphenols has been improved to a certain extent.Finally,the free radical scavenging ability of Kadsura coccinea polyphenol extracts was investigated.The results showed that at high concentrations(1 mg/mL),the DPPH free radical scavenging rate of various varieties and parts of Kadsura coccinea polyphenol extracts reached over 85%,and the ABTS·~+ free radical scavenging rate reached over 90%.Using cell experiments to verify the anti-inflammatory effect of polyphenol extracts from different parts of the Kadsura purple black variety,a RAW246.7 mouse inflammatory model was established.The differences in NO content released by cells and intracellular ROS content between the drug group and the model group were investigated.The differences in protein expression and inflammatory factor release between the drug group and the LPS control group were compared through WB and E-LISA experiments,explore the inhibitory effect of polyphenol extracts from Kadsura coccinea purple black varieties on inflammation models.The results showed that the Kadsura coccinea polyphenol extract was non-toxic to RAW 264.7 cells at a concentration of 6.25-100 μg/mL,and the cell survival rate remained above 90%.The drug group showed a significant decrease in NO release compared to the LPS control group.When the concentration of polyphenol extract was between 50-100μg/mL,the release of NO in the root drug group was significantly lower than that in the stem and leaf drug groups at the same concentration.When the concentration of polyphenol extract from the root of Kadsura coccinea was 100 μg/mL,the content of NO released by the LPS control group was about 2.23 times that of it(P<0.001).The intracellular ROS content in the drug group also showed a downward trend.When the drug concentration in the root experimental group was-50 μg/mL and 100μg/mL,the intracellular ROS level was reduced by 74.10%and 98.95%,respectively,compared to the LPS control group.Compared with the LPS control group,the TNF of each drug group-α、IL-1β、the expression levels of IL-6 and IL-12 proteins were significantly reduced,and the release of inflammatory factors also showed a downward trend.Using ultra-high performance liquid chromate graphy tandem mass spectrometry as the detection method,the qualitative and quantitative analysis of polyphenol metabolites in the root,stem,and leaf of Kadsura coccinea was carried out.A total of 949 metabolites were identified in the root,stem,and leaf of Kadsura coccinea,which can be divided into 14 types of metabolites,including phenolic acids,lignans,flavonols,anthocyanins,and coumarins.Among the three metabolites,phenolic acids,lignans,flavonoids,and flavonols dominate,with 410,149,120,and 101 species,respectively.The OPLS-DA model was used to screen differential metabolites,and the differences in metabolites among different parts were mainly concentrated in phenolic acids,lignans,and flavonoids,while the biological content differences between stilbenes and tannins were relatively small.Further screening identified 249 differentially shared metabolites among the three groups.Among the common differential metabolites in the root of Kadsura coccinea,phenolic acid metabolites mainly showed adownward trend,while lignan metabolites mainly showed an upward trend,which is opposite to the accumulation of metabolites in the stem and leaf parts.Meanwhile,KEGG metabolic pathway analysis was conducted on differential metabolic pathways.It was found that the differential metabolites between different parts are mainly enriched in the biosynthesis of secondary metabolites,flavonoid biosynthesis,and phenylpropanoid biosynthesis metabolic pathways.Studying the enriched metabolites in the phenylalanine metabolism pathway,it was found that phenolic acid precursor metabolites used for the synthesis of lignans have a significant accumulation in the stem,showing a significant upregulation trend compared to the root and leaf parts.This may be due to the fact that more phenolic acid precursor metabolites in the root and stem parts are used for the synthesis of lignans in this pathway. |
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