Studys On Relationship Between Rootins And Brownin And Phenolic Acids Of Tree Peony (Paeonia Suffruticosa Andr.) In Vitro

Three cultivars of tree peony (Paeonia suffruticosa Andr.)’Wu Long Peng Sheng’,’Tai Ping Hong’ and ’Feng Dan Bai’ were used as materials, the dynamic changes of phenolic content and related enzymes activities were assessed during the in vitro rooting and browing process of three cultivars of tree peony. Qualitatively and quantitatively of phenolic acids were analyzed of the higher browing degree plantlets and lower browing degree plantlets and the dynamic changes of the main phenolic acids was assessed during the in vitro rooting process of three cultivars of tree peony. The relationship between phenolic acids and related enzymes activities and rooting and browing of three cultivars of tree peony in vitro will be discussed. In order to clear the relationship between phenolic acids, IAA and PPO, the main phenolic acids in tree peony and IAA as materials, the reaction between them using PPO as catalyst was researched and the chemical structures of products were identified. An understanding for physiological and biochemical of plantlets provided is expected to provide vital clues to how better to try and improve tree peony in vitro rooting and browing. The main results were as follows:1. The dynamic changes of phenolic contents and PAL, PPO and POD activities were assessed during the differentiation process of three tree peony (Peaonia suffruticosa Andr.) cultivars grown in vitro. The results were as follows:there is a close relationship between PAL, PPO, POD activity as well as phenolic contents and tissue browning during the differentiation process in tree peony in vitro. The similar regulation was not appeared between the high or low level of PAL activity and tree peony browning degree, as the higher browning degree the cultivars, the lower phenolic contents and POD activities, and the higher PPO activities.2. Chlorogenic acid, gallic acid, catechol,4-methylcatechol, coumalic acid,6-hydroxycoumarin, caffeic acid, cinnamic acid, benzoic acid,4-hydroxy benzoic acid, shikimic acid,4-methoxy-3-hydroxyacetophenone,4-hydroxybenzhydrazide, paeonolum, resveratrol, paeoniflorin,2,3-dihydroxy-4-methoxyacetophenone methyl gallate were used as standard. The LC-MS method was used to qualitative and quantitative analysis of some phenolic acids in higher browing degree plantlets and lower browing degree plantlets of three cultivars of tree peony. The results were as follows:ten phenolic acids were detected in three different cultivars tree peony plantlets, and it is respectively paeoniflorin, methyl gallate, gallic acid, shikimic acid, benzoic acid,4-hydroxy benzoic acid,4-methoxy-3-hydroxyacetophenone, caffeic acid, catechol and chlorogenic acid. The rest of the eight kinds failed to detected. The content of gallic acid,4-hydroxy benzoic acid and caffeic acid in higher browing degree plantlets was lower than in lower browing degree plantlets in three cultivars; The content of paeoniflorin, methyl gallate, benzoic acid, shikimic acid and catechol was higher than in lower browing degree plantlets in three cultivars; The content of gallic acid, shikimic acid and benzoic acid was higher in higher browing degree cultivars, however, the content of caffeic acid was lower in higher browing degree cultivars. So, this study preliminary holds that these nine phenolic acids are related to tissue browing of tree peony, and gallic acid and coffee acid are related to tissue browing closely.3. Enzyme-related activities and total phenol content, which were related to the rooting of tree peony, changed in rooting culture for all three tree peony cultivars. These changes differed over the rooting period and there were significant differences between higher rooting rate cultivar ’Feng Dan Bai’ and lower rooting rate cultivars ’Wu Long Peng Sheng’and’Tai Ping Hong’. POD activity and total phenolic content were higher in the higher rooting rate cultivar’Feng Dan Bai’than in lower rooting rate cultivars’Tai Ping Hong’and’Wu Long Peng Sheng’on the whole, while IAAO activity was lower in’Feng Dan Bai’than in’Tai Ping Hong’and’Wu Long Peng Sheng’throughout the entire experimental period. PPO and PAL activity showed different differences in different period in rooting process of three cultivars.4. During the rooting culture process of tree peony, the phenolic acids in all three cultivars had changed compared with that of in control. And the content of paeoniflorin, benzoic acid and4-hydroxy benzoic acid in’Tai Ping Hong’cultivar was higher than that of in’Wu Long Peng Sheng’and’Feng Dan Bai’. While the gallicin and shikimic acid was lower than that of in’Wu Long Peng Sheng’and’Feng Dan Bai’. However, no significantly different could be drawn for gallic acid in three tree peony cultivars. So, this study preliminary holds that paeoniflorin, gallicin, gallic acid, shikimic acid, benzoic acid and4-hydroxy benzoic acid may be closely related with rooting in vitro of tree peony.5. The ten phenolic acids which were detected in tree peony and IAA as materials, the reaction between them using PPO as catalyst was researched, the results showed that, benzoic acid,4-hydroxy benzoic acid, shikimic acid,4-methoxy-3-hydroxyacetophenone and paeoniflorin were not the appropriate substrate on PPO because they could not be oxidation by PPO Within20hours and could not be reaction with IAA. Gallicin, caffeic acid, chlorogenic acid and gallic acid were the appropriate substrate on PPO because they could be oxidation by PPO Within2-7hours. When IAA and PPO was added to chlorogenic acid or caffeic acid or catechol solution, the mixtures had reacted and produced new compounds. The new compounds were not chlorogenic acid combinating with IAA after identified the chemical structures, but a class of compounds similar with IAA, which may be speculated that when chlorogenic acid, IAA and PPO were presence at the same time, IAA may be reacted with itself and formed a new compounds.