Phenolic Constituents And Biological Activities Of The Stems And Leaves Of Cyperus Esculentus

Cyperus esculentus L.is a multipurpose ecotype cash crop.However,the information on the chemical constituents and biological activities of its stems and leaves is quite limited,though there are extensive and in-depth research reports on its tubers.In order to provide supporting data for the comprehensive research,development and utilization of C.esculentus,this research was carried out to identify and quantify the chemical constituents and evaluate the biological activities of C.esculentus stems and leaves based on a survey of its biological characteristics,resources,chemical components,pharmaceutical applications,and pharmacological research progress.The main contents and results of this dissertation are as follows:Analysis of the chemical constituents and evaluation of the antioxidant and hepatoprotective activity:By ultra-high-performance liquid chromatography-mass spectrometry（UHPLC-MS）,30 compounds were identified from the stems and leaves for the first time,including six pairs of isomers,namely caffeoyl-glucaric acid isomers,feruloyl-glucaric acid isomers,caffeoylquinic acid isomers,coumaroylquinic acid isomers,（epi）gallocatechin and procyanidin B-type isomers.And quantitatively analyzed the dynamic changes in the content of 14 main components in stems and leaves of C.esculentus from June to October.Considering the growth cycle and tiger nut harvest time,extract of the stems and leaves in September（CELE9）was used for in-depth research.The quantitative analysis revealed that gallocatechin（GC,8）,procyanidin B1（B1,15）,catechin（C,16）,chlorogenic acid（19）,orientin（30）,and luteolin 7-O-glucuronide（31）are the major chemical constituents of CELE9.CELE9showed strong antioxidant activity in vitro and DPPH-LC-MS analysis revealed that compounds 8,15,and 19 contributed the most.CELE9showed significant protection against the agricultural fungicide tebuconazole（TEB）-induced developmental toxicity and hepatotoxicity in zebrafish,and also demonstrated glutathione-repleting activity.Investigation on the relationship between the secondary metabolites of the stems and leaves and the tuber morphologies of C.esculentus:The phenotypic parameters of two phenotypes of C.esculentus were compared,and the secondary metabolites in their stems and leaves were qualitatively and quantitatively analyzed by UHPLC-MS.The results showed that the contents of four flavonoids,luteolin 7-O-diglucuronide（29）,30,31 and luteolin（33）,were significantly higher in the stems and leaves of the phenotype with wide-leaf round-tuber than in those with narrow-leaf long-tuber.Four flavonoid differential metabolite biosynthesis pathways have been derived,providing a research basis for screening candidate genes and key enzymes for target flavonoid synthesis pathways from different phenotypes of C.esculentus.Comparative analysis of flavane-3-ols in multi-source plants and purification of procyanidin B1:The flavan-3-ols of 10 plants—primarily food by-products—including Muscat Hamburg grape seed,hawthorn sarcocarp,litchi pericarp,cocoa beans,peanut skin,lotus seedpod,Xinyang Maojian green tea,Cinnamomi cortex,Sargentodoxa cuneata stem and Cyperus esculentus stem and leaf were analyzed from multiple perspectives by UHPLC-MS.C.esculentus stem and leaf was found to contain high content of B1,second only to Muscat Hamburg grape seed.C.esculentus stem and leaf contained primarily procyanidin B1 with few isomers compare to grape seed.B1 was purified at 95%purity by ultrasonic extraction with 70%methanol,followed by an ODS column chromatograph,and 2 runs of Sephadex LH20 column chromatography,the yield was 0.21‰of the C.esculentus stem and leaf.In addition,1 g of C.esculentus stem and leaf yielded about 5.92 mg of epicatechin-cysteine conjugate.Thus,C.esculentus stem and leaf can be used as a source for the preparation of high purity B1 and strong bioactivity flavan-3-ol cysteine derivatives.Investigation on the bioactivities of different flavane-3-ols:The study investigated and compared the bioactivity of C,GC,B1,and procyanidin B2（B2）and their gallates,catechin gallate（CG）,gallocatechin gallate（GCG）,procyanidin B1 3-O-gallate（B1G）and procyanidin B2 3-O-gallate（B2G）.The results showed that galloylation significantly enhanced the bioactivity of flava-3-ols on protection of TEB-induced liver damage and Cu SO₄-induced inflammatory in zebrafish.CG,GCG,B1G and B2G did not transform in the culture medium without zebrafish,but in the medium with zebrafish there was a biotransformation of de-galloyl.At the same time,10 amino acid levels in the pathways of aminoacyl-t RNA biosynthesis,phenylalanine and tyrosine and tryptophan biosynthesis,valine and leucine and isoleucine biosynthesis,and phenylalanine metabolism in zebrafish were significantly up-regulated,suggesting that the alleviation of TEB induced zebrafish injury was related to the biotransformation of these compounds and the up-regulation of the amino acid levels.In summary,30 chemical components,including six pairs of isomers,were identified for the first time from the stems and leaves of C.esculentus.Among the chemical components,the contents of flavan-3-ol were high,and the content of B1 was the highest.An efficient purification process for preparing high-purity B1 from C.esculentus stems and leaves was established.The C.esculentus stem and leave extract,C,GC,and B1showed significant protective effects on the agricultural fungicide tebuconazole-induced hepatotoxicity in zebrafish,and gallate esterification significantly improved the hepatoprotective and anti-inflammatory activities of flavan-3-ol in vivo.This study provides a scientific basis for the innovative utilization of the aboveground stems and leaves of C.esculentus.