Identification of potential genes involved in biosynthesis of flavonoid and analysis of biosynthetic pathway in Fagopyrum dibotrys / 中国中药杂志

In order to enrich the transcriptome data of Fagopyrum dibotrys plants, analyze the genes encoding key enzyme involved in flavonoid biosynthesis pathway, and mine their functional genes, in this study, we performed RNA sequencing analysis for the rhizomes, roots, flowers, leaves and stems of F. dibotrys on the BGISEQ-500 sequencing platform. After de novo assembly of transcripts, a total of 205 619 unigenes were generated and 132 372 unigenes were obtained and annotated into seven public databases, of which, 81 327 unigenes were mapped to the GO database and most of the unigenes were annotated in cellular process, biological regulation, binding and catalytic activity. Besides, 86 922 unigenes were enriched in 136 pathways using KEGG database' and we identified 82 unigenes that encodes key enzymes involved in flavonoid biosynthesis. Comparing rhizome with root, flower, leaf or stem in F. dibotrys, 27 962 co-expressed differentially expressed genes(DEGs) were obtained. Among them, 23 515 DEGs of rhizome tissue-specific were enriched into 132 pathways and 13 unigenes were significantly enriched in biosynthesis of flavone and flavonol. In addition, we also identified 3 427 unigenes encoding 60 transcription factor(TFs) families as well as four unigenes encoding bHLH TFs were enriched in flavonoid biosynthesis. Our results greatly enriched the transcriptome database of plants, provided a reference for the analysis of key enzymes involved in flavonoid biosynthesis in plants, and will facilitate the study of the functions and regulatory mechanisms of key enzymes involved in flavonoid biosynthesis in F. dibotrys at the genetic level.

Study on Network Pharmacology of Superior Components of Fagopyrum dibotrys / 中国药房

OBJECTIVE：To study the pharmacological effects and possible molecular mechanism of the superior components in Fagopyrum dibotrys . METHODS ：Based on network pharmacology ，by selecting DL ＞0.18 and OB ＞30％ as criteria ，superior components of F. dibotrys were screened out ，using the traditional Chinese medicine integrated pharmacology （TCMIP）platform. Pharm Mapper database was utilized to obtain the potential targets of each components ；Kyoto gene and genome database （KEGG） signal pathway analysis and gene ontology （GO）bioprocess enrichment analysis were performed for target protein with DAVID database（all using P＜0.05 as criteria ）. RESULTS ：15 kinds of superior components [such as quercetin ，luteolin，procyanidin B 1， （+）catechin and β-sitosterol，etc.] and 114 target proteins （such as estradiol 17-β-dehydrogenase 1，cAMP specific 3，5-cyclic phosphodiesterase 4D，vitamin D 3 receptor，uridine cytidine kinase 2，urokinase triphosphate encoded by HRAS gene ，etc.）were screened out ，involving 34 important pathways ，like MAPK signaling pathway ，VEGF signaling pathway ，chemokine signaling pathway and insulin signaling pathway ；among them ，11 were cancer-related pathways ，7 were metabolic-related pathways ，4 were endocrine-related pathways. The involved molecular functions included steroid receptor activity ，ligand dependent nuclear receptor activity，protein kinase activity ，etc.；cell components included cell fluid ，cell fragment ，soluble part ，etc.；biological processes included the regulation of apoptosis ，the process of organic response ，the process of endogenous stimulation response ，etc. CONCLUSIONS：F. dibotrys may play anti-tumor effects ，anti-inflammatory effects ，regulation of glycolipid metabolism by acting on MAPK signaling pathway ，VEGF signaling pathway ，chemokine signaling pathway and insulin signaling pathway.

Characterization of the transcriptional profiles in common buckwheat (Fagopyrum esculentum) under PEG-mediated drought stress

BACKGROUND: Common buckwheat (Fagopyrum esculentum) is an important staple food crop in southwest China, where drought stress is one of the largest limiting factors that lead to decreased crop production. To reveal the molecular mechanism of common buckwheat in response to drought stress, we performed a comprehensive transcriptomics study to evaluate gene expression profiles of common buckwheat during PEG-mediated drought treatment. RESULTS: In total, 45 million clean reads were assembled into 53,404 unigenes with an average length of 749 bp and N50 length of 1296 bp. A total of 1329 differentially expressed genes (DEGs) were identified by comparing wellwatered and drought-treated plants, out of which 666 were upregulated and 663 were downregulated. Furthermore, we defined the functional characteristics of DEGs using GO and KEGG classifications. GO enrichment analysis showed that the DEGs were significantly overrepresented in four categories, namely, "oxidoreductase activity," "oxidation­reduction process," "xyloglucan:xyloglucosyl transferase activity," and "apoplast." Using KEGG pathway analysis, a large number of annotated genes were overrepresented in terms such as "plant hormone signal transduction," "phenylpropanoid biosynthesis," "photosynthesis," and "carbon metabolism." Conclusions: These results can be further exploited to investigate the molecular mechanism of common buckwheat in response to drought treatment and could supply with valuable molecular sources for abiotic-tolerant elite breeding programs in the future.

MYB Transcription Factors in Fagopyrum tataricum / 中国实验方剂学杂志

Known as "the king of the five grains",tartary buckwheat (Fagopyrum tataricum) is a medicinal and edible plant with both nutritional and healthcare functions. It contains rich flavonoids, such as rutin,with balanced amino acid composition and multiple effects, like blood fat-lowering,blood giucose-lowering,blood pressure-lowering,anti-oxidation,anti-aging,anti-cancer,anti-cancer,and microcirculation-improving. Transcription factors play important roles in plant growth and development by regulating gene expressions. MYB family is one of the largest transcription factor families in plant,contains the MYB domain,and can be divided into four subfamilies:1R-MYB,R2R3-MYB,3R-MYB,4R-MYB. This family plays various roles in plant growth,plant development and flavonoid biosynthesis of secondary metabolism. In this study,the reported MYB transcription factors in F. tataricum were summarized and systemically clustered,and their interrelationships were defined to provide references for further exploring and cloning MYB transcription factors in F. tataricum. In addition,this study reviewed their regulatory functions of MYB transcription factors in flavonoid biosynthesis pathway,plant hormones pathways and other abiotic stress pathways,and made a conclusion and advances about the future research in F. tataricum. Therefore,this study will provide valuable scientific references for the functional studies of MYB family transcription factors in F. tataricum and its molecular breeding for high-quality varieties.

Comparison of Neck Pain, Shoulder Pain, and Comfort between Buckwheat and Latex Pillows

BACKGROUND: This study aimed to investigate differences in neck pain, shoulder pain, and comfort between buckwheat and latex pillows in elderly sanatorium residents. METHODS: The crossover experimental study was performed with 30 elderly patients in a convalescent hospital for seniors. The buckwheat and latex pillows were used for 2 weeks, the 7 day was assigned as the washout between each one week intervention, respectively. Pre- and posttest visual analog scale (VAS) scores for neck and shoulder pains and the neck disability index (NDI) were investigated. Data were analyzed using paired-samples t test, and their correlations were analyzed using SPSS/WIN 21.0 (IBM Corp., New York, NY, USA). RESULTS: After the 2-week intervention, results showed that the use of buckwheat pillows decreased the VAS scores for neck and shoulder pains and NDI. Further, the use of latex pillows decreased the VAS score for neck pain and NDI, but the VAS score for shoulder pain showed no statistically significant differences (t=1.887, P=0.069). CONCLUSION: In conclusion, using latex and buckwheat pillows is the best option for reducing neck pain. Between the buckwheat and latex pillows, the buckwheat pillow is a better option for decreasing neck and shoulder pains.

Influence of light on gene expression of key synthesis enzyme genes FtANR and FtLAR about proanthocyanidin in seeds of homologous plant of food and medicine Fagopyrum tataricum / 中国中药杂志

Tartary buckwheat Fagopyrum tataricum is an important medicinal and functional herb due to its rich content of flavonoids in the seeds. F.tataricum exhibited good functions for free radicals scavenging， anti-oxidation， anti-aging activities. Although much genetic knowledge of the synthesis， regulation， accumulation of rutin， the genetic basis of proanthocyanidins(PAs) in tartary buckwheat and their related gene expression changes under different lights(blue， red， far red， ultraviolet light) remain largely unexplored. In this study， we cloned one anthocyanidin reductase gene(ANR) and two leucocyanidin reductase gene(LAR) named FtANR，FtLAR1，FtLAR3 involved in formation of(+)-catechin and(-)-epicatechin precusor proanthocyanidin by digging out F. tataricum seed transcriptome data. The expression data showed that the opposite influence of red light on these gene transcript level compared to others lights. The expression levels of FtANR and FtLAR1 decreased and FtLAR3 appeared increment after exposed in the red light， while the expression levels of those genes appeared opposite result after exposed in the blue and far red light.

Content Determination and Analysis of Flavonoids in Buckwheat from Guizhou Province Content Determination and Analysis of Flavonoids in Buckwheat from Guizhou Province / 中国药房

OBJECTIVE:To establish a method for contents determination of rutin,quercetin and total flavonoids in Buck-wheat,and compare the difference of flavone in buckwheat in different varieties and areas. METHODS:HPLC was adopted for content determination of rutin and quercetin:the column was Aglient C18 with mobile phase of methanol-0.4%phosphoric acid(gra-dient elution)at a flow rate of 1 ml/min,detection wavelength was 360 nm,column temperature was 40 ℃,and injection volume was 10 μl;UV spectrophotometry was adopted for content determination of total flavonoids:reference solution was methanol,de-tection wavelength was 360 nm,the standard was rutin. Statistical method was used to analyze the differences among the Fagopy-rum tataricum,F. esculentum and flavonoids of F. tataricum from different areas. RESULTS:The linear range was 0.400 9-16.03 μg for rutin (r=0.999 4) and 0.009 9-0.396 0 μg for quercetin (r=0.999 7),RSDs of precision,stability and reproducibility tests were lower than 2%,recoveries were 99.33%-104.00%(RSD=1.32%,n=9) and 96.92%-101.66%(RSD=1.60%,n=9),re-spectively. The linear range of total flavonoids(recovded by rutin)was 4.14-41.4 mg/L(r=0.999 9),RSDs of precision,stability and reproducibility tests were lower than 2,and recovery was 96.07%-101.96%(RSD=2.63%,n=9). The content of flavonoids on F. esculentum is significantly higher than in F. tataricum from the same area,and F. tataricum in Bijie area is better than other places. CONCLUSIONS:The method is simple,stable,reproducible,and can be used for the contents determination of rutin, quercetin and total flavonoids in F. esculentum. The study can provide theoretical basis for developing and using buckwheat.

Chemical constituents from root tubers of Fagopyrum dibotrys / 中草药

Objective: To study the chemical constituents from the root tubers of Fagopyrum dibotrys. Methods: The compounds were isolated and purified by means of chromatographic techniques and their structures were identified on the basis of spectral features. Results: Fourteen known compounds were isolated from methanol extract in the dry roots of F. dibotrys and thier structures were identified as luteolin (1), tricin (2), luteolin-7,4'-dimethyl ether (3), quercetin (4), genkwanin (5), chrysoeriol (6), protocatechuic acid (7), protocatechuic acid methyl ester (8), p-hydroxybenzaldehyde (9), glutinone (10), glutinol (11), olean-12-ene-3β, 7β, 15α, 28-tetraol (12), juglangenin A (13), and 21β-dihydroxy-olean-12-ene (14). Conclusion: Compounds 5 and 6 are firstly obtained from F. dibotrys. Compounds 12-14 are isolated from the plants of Fagopyrum Mill. for the first time.

Chemical constituents from aerial parts of Fagopyrum dibotrys / 中草药

Objective: To study the chemical constituents from the aerial parts of Fagopyrum dibotrys. Methods: The compounds were isolated and purified by means of chromatographic techniques and their structures were identified on the basis of spectral features. Results: Fourteen known compounds were isolated in the methanol extract from the aerial parts of F. dibotrys and their structures were identified as benzoic acid (1), p-hydroxybenzoic acid (2), p-hydroxy benzaldehyde (3), 3,4-dihydroxy benzoic acid (4), succinic acid (5), caffeic acid (6), methyl caffeate acid (7), luteolin (8), tricin (9), quercetin (10), afzelin A (11), 2α,3β,29-trihydroxyolean-12-en-28-oic acid (12), yarumic acid (13), and 3α-hydroxy-urs-12,15-dien (14). Conclusion: Compounds 2-3 and 6-9 are firstly obtained from the aerial parts of F. dibotrys. Compounds 11-14 are isolated from the genus of Fagopyrum Mill. for the first time.

Gene cloning of anthocyanin synthase in Fagopyrum dibotrys and correlation between its expression level and anthocyanin content / 中草药

Objective: To obtain the full-length cDNA sequence of anthocyanins synthase (ANS) gene from Fagopyrum dibotrys (FdANS), and to analyze the expression of FdANS in each tissue and the total anthocyanin content during florescence of F. dibotrys. Methods: The cDNA sequence of ANS gene was cloned by homology cloning from F. dibotrys. The expression of ANS was analyzed by semi-quantitative RT-PCR. The content of total anthocyanin was measured by spectrophotometry. Results: The open reading frame (ORF) of FdANS was 1077 bp and encoded a protein with 358 amino acids named FdANS. Bioinforamtion analysis suggested that the amino acid sequence of FdANS had the higher homology with those in other plants. Both the gene expression FdANS in different tissues during florescence of F. dibotrys and the total anthocyanin content were in the order of flowers > leaves > stems> roots. Conclusion: The cDNA sequence of FdANS is firstly obtained from F. dibotrys and its coding protein has the typical characteristic of ANS homologous protein. The gene expression of FdANS shows the correlation to the total anthocyanins content in the flowers, leaves, stems, and roots of F. dibotrys.

Cloning of flavonoid 3'-hydroxylase gene from Fagopyrum tataricum and its tissue-specific expression under cold stress / 中草药

Objective: To clone and analyze the full-length of flavonoid 3'-hydroxylase (F3'H) gene from tartary buckwheat (Fagopyrum tataricum) and to express it in Escherichia coli. FtF3'H gene expression and anthocyanins accumulation is also to be analyzed in tartary buckwheat sprout under cold stress. Methods: Homology cloning and RACE method were used to obtain FtF3'H gene from flower buds of tartary buckwheat. The recombinant vector pET-30b (+)-FtF3'H was constructed and expressed in E. coli BL21 (DE3). Semi-quantitative RT-PCR was used to analyze FtF3'H gene expression when spectrophotometric method was used to determine anthocyanin content. Results: FtF3'H gene contains an open reading frame (1470 bp) encoding 489 amino acids and belongs to cytochrome P450 family. SDS-PAGE analysis of IPTG induced recombinant E. coli BL21 (DE3) showed that a predicted 54000 Da fusion protein was expressed in the culture. Cold stress significantly enhanced the expression level of FtF3'H and anthocyanin accumulation (P < 0.05). Conclusion: FtF3'H gene could be cloned from F. tataricum and efficiently expressed in E. coli. Under cold stress, FtF3'H gene may enhance its expression level to promote anthocyanin accumulation, by taking part in the process of cold-stress resistance of F. tataricum sprouts.

Alpha and Beta amylase activity of Fagopyrum esculentum (Buckwheat): A Medicinal Plant

Background and Objectives: Fagopyrum esculentum, common buckwheat popularly known as mithe fapar is one of the staple food crops of the mountain region. Traditionally, it is used to treat constipation and bowel upsets. It is also used by diabetic in different parts of Nepal and India. Due to its high nutritive and medicinal value, medical scientist and researchers are interested in developing this as pharmaceutical plant. In this regard department of biochemistry, College of Applied Education and Health Sciences, C.C.S. University, Meerut, India is working to analyse the biochemical composition and benefits of this plant. So, as a part of a multidimensional project of analyzing various components and their impact on health and diseases, here we are reporting the amylase activity during germination of seed in Buckwheat (Fagopyrum esculentum) plant. Methodology: Common buckwheat (Fagopyrum esculentum) seeds were taken and germinated in dark at room temperature from 0 hours to 192 hours. Biochemical analysis for total amylase, alpha and beta amylase activities was measured by the standard method designed by Bernfeld (1955). Results: The seeds of buckwheat showed high level of amylolytic activity during different stages of germination. At 0 hours, negligible amylase activity was found. The first amylase activity was found at 24 hours and increases up to 96 hours. After 96 hours the total amylase activity starts decreasing and becomes almost negligible at 192 hours. Alpha and Beta –Amylase activity is reported separately. Conclusion: The amylases from the buckwheat showed different level of enzymatic activity during seed germination. Alpha amylase contributed a larger account to total amylase activity. The activity starts increasing and becomes maximum at 96 hours and starts decreasing and becomes lowest at 192 hours suggesting that alpha amylase plays a important role in starch metabolism in developing as well as geminating seeds which can be used for the drug discovery and treatment of several diseases like diabetes, polycystic ovary syndrome, constipation, bowel upsets, obesity and others.

Gene cloning and expression level of chalcone isomerase during florescence and content of flavonoids in Fagopyrum dibotrys / 中草药

Objective: To clone and analyze the full-length cDNA of chalcone isomerase (CHI) gene from Fagopyrum dibotrys (FdCHI). To analyze the expression of CHI and the total flavonoids content during florescence of F. dibotrys. Methods: The cDNA sequence of CHI was cloned by homology cloning from F. dibotrys. The expression of CHI was analyzed in the different tissues during florescence by semi-quantitative RT-PCR. The content of total flavonoids was measured by AlCl3 method. Results: The open reading frame (ORF) of FdCHI was 750 bp and encoded a protein with 249 amino acids. Bioinforamtion analysis suggested that the amino acid sequence of FdCHI had the high homology with those in other plants. Gene expression analysis showed that FdCHI was highly expressed in the flowers, followed by the roots and leaves, while lower in the stems. The content of total flavonoids was the highest in the flowers then the leaves and stems, and the lowest was in the roots. Conclusion: The cDNA sequence of FdCHI is firstly obtained from F. dibotrys and its coding protein has the typical characteristic of CHI homologous protein. The gene expression of FdCHI shows the same to the total flavonoids content in the stems, leaves, and flowers, but different in the roots of F. dibotrys.

A Case of Bronchial Constriction Due to Buckwheat Allergy / 대한내과학회지

Buckwheat has been reported as a food allergen that can induce urticaria, angioedema and anaphylaxis. Moreover, it can cause occupational asthma and acute exacerbation in patients with asthma. Here, we present a case of an asthmatic patient with bronchial constriction due to buckwheat, who was treated for bronchial asthma and atopic dermatitis.

Rutin- potent natural thrombolytic agent.

Thrombosis, the formation of blood clots, is a cause not only of heart attacks and strokes, but of deep venous throm-bosis (DVT) and pulmonary embolism as well. The number one killer of Americans is a blood clot that blocks blood flow to the heart or to the brain and approximately half of all morbidity and mortality in the United States can be attributed to heart attack or stroke. All the blood clot related conditions are life-threatening, and so there is a need for safe, effective and preventive treatment. A natural substance rutin, also called rutoside, is a citrus flavonoid glycoside found in Fagopyrum esculentum (buckwheat), the leaves and petioles of Rheum species, and Asparagus. This flavonoid compound has shown effective thrombolytic activity (prevents the formation of blood clots) by blocking the enzyme protein disulfide isomerase (PDI) found in all cells involved in blood clotting. Food and Drug Administration (FDA) has established that rutin is safe and, thus provides a safe and inexpensive drug that could reduce recurrent clots and help save thousands of lives.

Pharmacokinetics of Fagopyrum tataricum Total Flavonoids in Beagle Dogs / 中国药房

OBJECTIVE:To study the pharmacokinetic features of Fagopyrum tataricum total flavonoids. METHODS:6 Beagle dogs were intragastrically given single dose of 80 mg F.tataricum total flavonoids (with content of quercetin at 1.2% and the content of eldrin at 24.7%). HPLC-UV spectrophotometry was adopted to determine the content of total quercetin in the hydrolyzed plasma samples. RESULTS:Taking the total quercetin as the index of the pharmacokinetic features of the F. tataricum total flavonoids in the Beagle dogs,the main pharmacokinetic parameters were as follows:AUC0→t:(9.92?1.49)?g?h?mL-1;AUC0→∞:(10.81?1.86)?g?h?mL-1;Cmax:(1.07?0.19)?g?mL-1. CONCLUSION:The pharmacokinetics of the F.tataricum total flavonoids in the Beagle dogs conforms to two-compartment model,and the concentration-time curves are characterized by multiple peak pharmacokinetics.

Purification and Characterization of a Protease Inhibitor from Fagopyrum tartaricum Gaertn Seeds and Its Effectiveness Against Insects / 中国生物化学与分子生物学报

Protease inhibitors, which are widely distributed in all types of life forms, are generally considered to be one of the most abundant proteins and a defense mechanism. A protease inhibitor from tartary buckwheat seeds (TBTI-Ⅱ ), with specific trypsin-inhibitory activity, was obtained by Resource Q anion-exchange chromatography and Superdex G 75 gel filtration. SDS-PAGE analysis indicated that the approximate molecular weight was 9.0 kD. Amino-acid analysis showed that the TBTI- Ⅱ was composed of 80 amino-acid residues with a high content of glutamate, aspartate and arginine. The inhibitor had high thermostability and retained 67.6% of its activity after heating at 100℃ for 10 min. The inhibition constant Ki was determined to be 1.01× 10-4 mol/L. It was demonstrated that the inhibitor was able to have an effect on the growth of cotton bollworm larva, after being fed with the artificial diets mixed with the target inhibitor. The present study indicates that the trypsin inhibitor from tartary buckwheat seeds could be a new potential anti-insect factor.

Extracting technology of total flavones in powder of Fagopyrum tataricum stem and leaf by enzymatic treatment / 中草药

Objective To optimize the extracting technology of total flavones in powder of Fagopyrum tataricum stem and leaf by enzymatic treatment. Methods The powder was treated by cellulase before extracted using water, the effects of enzyme dosage, treatment temperature, treatment time, and pH value on the extract rate of total flavones were studied. Results The optimum extracting technology was as follows: enzymatic treatment temperature: 55℃, enzyme dosage: 3.0 ?L, pH value: 6.5, treatment time: 90 min, extracting 3 times at: 90℃ for 30 min once. The extract rate of total flavones was 1.47% by this technology. Conclusion Cellulase could be applied in the assisstant extraction of total flavones in powder of F. tataricum stem and leaf.

Kinetics and related thermodynamics on the extraction of flavonoids from Fagopyrum tartarjcum / 中成药

AIM: To explore the kinetics and thermodynamics parameters in the extraction of flavonoids from Fagopyrum tartarjcum . METHODS: Rutin was adopted as marker substance to create a regression equation between Rutin concentration and absorbance by meaus of UV. RESULTS: According to Fick diffusion principle Ⅱ, extraction process was analysed to gain parameters, such as K, Ea, Y, t 1/2 , K ?. CONCLUSION: The method would be useful for the process design and the choice of operation condition to extract flavonoids from Fagopyrum tartarjcum.

Effects of PFEM and SOD on Cerebral Ischemia Followed by Reperfusion in Gerbils / 昆明医科大学学报

By the ligation of gerbils bilateral carotid arteries and then reperfusion, the effects of PFEM and SOD on the stroke index were evaluated after a 45 min cerebral ischemia followed by a 6 hr reperfusion period and on the mortality after a 24 hr reperfusion period. The resulta indicated that both PFEM and SOD could decrease the stroke index and mortality dramatically. We speculated upon the protective effect of PFEM on cerebral damage following ischemia with reperfusion. It might be due to the inhibition of the free radical metabolism.