Research On Flavonoid Metabolomics Of Tartary Buckwheat Seeds

Tartary buckwheat is an annual plant of the genus Fagopyrum and has high edible and medicinal value.Tartary buckwheat seeds are rich in flavonoids,such as rutin,quercetin,catechins,and epicatechins,which has the functions of lowering blood glucose,blood lipid,blood pressure,anti-aging and dredging blood vessel circulation.This research focuses on the research of flavonoids in metabonomics.First,the flavonoids in tartary buckwheat were identified by LC-QTOF-MS,and then the detected flavonoids in the seeds of tartary buckwheat were quantitatively analyzed by LC-QQQ-MS.Lastly,the effects of tartary buckwheat traits on flavonoid composition were clarified,which provided a basis for the selection and identification of tartary buckwheat seeds.The results of this research are as follows:1.Construction of the Flavonoid Spectrum of Tartary Buckwheat SeedsLC-QTOF-MS technology was used to identify the flavonoids in tartary buckwheat seeds.The flavonoids in tartary buckwheat seeds were determined by comparison with reference standards,accurate mass numbers,fragment ion,and the fragmentation pattern of flavonoids.60 types of flavonoids were identified,including flavonols,flavanols and other flavonoids.The flavonoid profile of tartary buckwheat seeds was successfully constructed,which would lay the foundation for the further quantitative research.2.Establishment of Targeted Metabolomics for Tartary Buckwheat SeedsBased on the analytical method of flavonoid in metabolomics using LC-QQQ-MS/MS established earlier in the laboratory,the experimental conditions including Q1(Parent ion),Q3(Product ion),DP(Declustering potential)and CE(Collision energy),were further optimized to obtain a more stable and reliable method for targeting analysis of flavonoid metabolites in tartary buckwheat seeds.3.Effect of Tartary Buckwheat Seed Traits on Flavonoid MetabolitesThe effect of tartary buckwheat seed traits on flavonoid metabolites was studied using LC-MS/MS.60 flavonoid metabolites in 40 types of tartary buckwheat seed coats and seed kernels were analyzed to study the relationship between tartary buckwheat seed traits and the changes of the flavonoid metabolites in tartary buckwheat seeds.Multivariate statistical methods were used to find differential metabolites associated with the color and shape of tartary buckwheat seeds.OPLS-DA and Volcano plot analysis showed that the differential metabolites in the groats of yellow-brown seeds were higher than those in black seeds,while the hulls in tartary buckwheat,the differential metabolites in the hulls of black seeds are higher than those in yellow-brown ones.Different metabolites in hulls or groats showed the different trend.However,the differential metabolites in long tartary buckwheat seeds are higher than those in the short seeds regardless of the hulls and groats.4.Research on the Difference Between Hulls and GroatsThe LC-MS/MS method was used to find the differential flavonoid metabolites between hulls and groats.Sixty flavonoid metabolites in 40 tartary buckwheat between hulls and groats were measured,and the differences between hulls and groats flavonoid metabolites were reflected according to the changes in the content of metabolites.Multivariate statistical methods were used to find out the different metabolites between hulls and groats.Analysis the data by PCA and Volcano plot,Metabolites between groats and groats were also different.There are 9 metabolites higher in hulls,and 43 metabolites higher in groats.5.Qualitative and Quantitative Research of AnthraquinonesAn MS-based analytical method about anthraquinone metabolomics in tartary buckwheat seeds was established with negative ion detection mode.During the research,anthraquinone metabolites in tartary buckwheat were identified based on the fragmentation of fragment ions and 11 types of anthraquinone metabolites were identified.The experimental conditions were further optimized to establish a stable and reliable method for targeting analysis of anthraquinone metabolites in tartary buckwheat seeds.11 anthraquinone metabolites in 40 hulls and 40 groats wereidentified.Multivariate statistical analysis showed that 3 differential metabolites in the groats of the yellow-brown seeds are higher than those in the black seeds,while 2differential metabolites in the hulls of the yellow-brown seeds are lower than those in the black seeds.When taking the color of tartary buckwheat seeds into consideration,metabolites in groats and hulls showed the different trend,while there is no significant difference in differential metabolites when considering the shape.The metabolites between hulls and groats are also different,where 8 metabolites are higher in hulls,and 3 metabolites are higher in groats.6.Molecular Mechanism of Flavonoids in Tartary Buckwheat SeedsTwo pairs of hybrid combinations(TB063 and TB324,TB325 and TB175)were screened by metabonomics of tartary buckwheat immature seeds with rutin as a characteristic compound.At the same time,transcriptome analysis was performed for the two pairs of hybrid combination cyan seeds and compound differential analysis was resolved at the RNA sequence level.