Study On Flowering And Pollination Characteristics And Main Chemical Compositions In Three Variants Of Gastrodia Elata From Southwest Of China

The dry tuber of Gastrodia elata Bl.was one of the valuable traditional Chinese medicines,which distributed mainly in Guizhou,Yunnan,Sichuan,Shanxi,Hubei,and so on.Gastrodia elata Bl.was usually divided into five variants:G.elata Bl.f.elata,G.elata Bl.f.glauca S.Chow,G.elata.Bl.f.viridis Malkino,G.elata Bl.f.flavida S.Chowand G.elata Bl.f.albaS.Chow.Southwest is the main gastrodia producing area,G.elata Bl.f.elata,G.elata Bl.f.elataandG.elata Bl.f.glauca S.Choware the main cultivars.But the difference of the main chemical components among different variants in southwest have not yet been studied systematically.At present,asexual reproduction as the main mode of reproduction have been widely used in cultivation of G.elata.Meanwhile,the long-term asexual reproduction can cause the degradation of G.elata cultivars.Usually,after several generations of asexual reproduction,sexual alternation would be adoptedalternatively,or different variantsG.elatacould be hybridized to make use of heterosis.So far,no systematic reports character the flowering process including pollen viability and stigma receptivity.In this study,on the one hand,pollen viability and stigma receptivity of G.elata in different variants were studied.On the other hand,the polysaccharides,free amino acids,total polyphenols,gastrodin,hydroxybenzyl alcohol and four parishins were detectedand analyzed among different gastrodia variants.It will provide a theoretical basis for further exploitation and evaluation of the quality of G.elata in this area,and to find out the appropriate time of pollination of different types of G.elata,and provide scientific theoretical basis for the further development of new varieties and the breeding of good varieties for G.elata.The main results were as following:1.The viability and life span of pollen were evaluated by peroxidase solution,the stigma receptivity were estimated by benzidine-H₂O₂ method.The results showed that G.elata f.glauca,G.elata f.elata and G.elata f.viridis appeared the same up-and-down trend of the pollen viability,increased firstly and then decreased.The highest viability of pollen appeared at the 2nd day after blooming.The storage temperature and time had significant impact on the pollen viability.With the extension of storage time,the pollen viability decreased.-20?refrigerator storage might extend the pollen viability.The stigma of three G.elata variants had stigmareceptivity at the 1st day.G.elata f.glauca reached the highest level at the 2nd day and had no receptivity at the 7th day after blooming.G.elata f.elata and G.elata f.viridis reached the highest level at the 3rd day and had no receptivity at the 8th day after blooming.Using artificial pollination,it was suggested that the stigma of G.elata f.glauca at the 2nd day and the stigma of G.elata f.elata and G.elata f.viridis at the 3rd day and the pollen at the 2nd day after blooming should be selected for artificial pollination.2.The comparison and principal component analysis were carried out on the main chemical composition contents in three variants of G.elatafrom southwest of China.The resultsindicatedthatthecontentsofmainchemicalcompositionwere differentamongdifferent variants of G.elata.from southwest of China.The coefficient of variation?CV?of polysaccharidecontentswas the smallest with the value of 18.025%,but the CV of parishin B was the biggest with 62.045%.Among the components in this study,the contents of free amino acid,total polyphenol,gastrodine,parishin B and parishin E of G.elata f.glauca from Beichuan in Sichuanprovincewere 2.873%,0.805%,0.862%,1.146%and 2.101%respectively,which wereall extremely higher than that of the other experimental materials.G.elata.f.viridis from Dafang in Guizhouprovince had the highest content of polysaccharide with the value of 29.225%,which had significantlyhigher content of polysaccharide than that of G.elata f.glauca from Beichuan and had extremely significantly higher content of polysaccharides than that of the rest.G.elata f.glauca from Zhaotong in Yunnan province had the highest content of hydroxyphenyl methanol,thenG.elata f.elata from Guangyuan inSichaun province,with no significant difference between them.The contents of parishin A and parishin C of G.elata f.elata from Guangyuan in Sichuan province were 0.877%and0.383%respectively,which were significantly higher than that of the other materials.The content of gastrodine of three variants of G.elata fom Sichuan provinceand Yunnan province had no significant difference,but which were both extremely significantly higher than that of three variants of G.elata fom Guizhou province.Moreover,The contents of parishin B and parishin E of three variants of G.elata fom Sichuan province were significantly higher than that of Yunnan and Guizhou province.However,the contents of other main chemical compositions among three variants of G.elata from Yunnan,Guizhou and Sichuan province had no significant difference.Comprehensively,the main chemical compositions among variants of G.elata.from southwest of China were different,of them,the main chemical composition contents in G.elata from Sichuan province were higher,but the parishin contents had no obvious relation to the variants.In total,the quality of G.elata f.glauca from Beichuan in Sichuanprovince was the best from the perspective of main chemical composition contents.3.In this study,HPLC method for the determination of parishinA,parishin B,parishinC and parishinE in G.elatawere established.Theanalysis was performed on an Agilent 1200 system with a Agilent TC-C₁₈column?4.6 mm×250 mm,5?m?.The four components wereseparated in30 min with gradient mobile phase consisting ofacetonitrile?A?and 0.1%phosphoric acid-H₂O?B?:0-10 min,10%A;10-20 min,10%-25%A;20-30min,25%A.The temperature of column was 30?,and the injection volume was 20?L.The four components had the good linearity?r?0.9990?within the linear ranges.The precision testRSD was 0.306%¹.205%,stability testRSD was 1.007%².142%,repeatability testRSD was 1.067%².162%,the average recovery rate was96.322%-103.105%,the RSD value was 1.713%².797%and all RSD value<3.0%.It revealed that the test has good precision,the sample solution has good stability,and the repeatability and accuracy of the test are all high.4.The hot water extraction,ultrasound extraction and enzymatic hydrolysis were applied to extracting polysaccharides from G.elata Bl.f.glauca S.Chow from Pingwu in Sichuan provence,then the polysaccharides extraction ratioes and their antioxidant activities were compared.The polysaccharides extraction ratioes and antioxidant activities were obviously influenced by the extraction methods.Of them,the extraction ratio of enzymatic hydrolysis was the biggest with the value of 50.315%,which was extremely different from the other extraction methods?P<0.01?.The antioxidant activities were in the following order:ultrasound extraction,enzymatic hydrolysis and hot water extraction by using DPPH free radical scavenge capacity and FRAP values,but the antioxidant activities were in the following order:enzymatic hydrolysis,ultrasound extraction and hot water extraction by using ABTS free radical scavenge capacity.In conclusion,due to the higher extraction rate and antioxidant activity,enzymatic hydrolysis was fit to extract polysaccharides from G.elata.