抗褐化剂对天女木兰芽外植体褐化与酚酸氧化的影响

Effects of Browning Inhibitors on Bud Explants Browning and Phenolic Acids Oxidation of Magnolia sieboldii K. Koch

[目的]确定引起天女木兰褐化现象的酚酸种类,筛选出最佳抗褐化剂及使用最适浓度。[方法]在B5培养基中分别添加各种抗褐化剂,芽外植体培养30 d后计算出褐化率,对褐化率结果进行统计分析,并比较三种抗褐化剂的抗褐化效果;另外,在组培过程中定期取样,利用HPLC测定芽外植体中咖啡酸、绿原酸和对香豆酸含量变化。[结果]表明:天女木兰芽外植体褐化过程中咖啡酸和绿原酸含量下降幅度较大,而对香豆酸含量下降幅度较小,说明咖啡酸和绿原酸容易被氧化,而对香豆酸较为稳定;三种抗褐化剂控制褐化现象发生的最佳效果排序为VC>PVP>CA;尽管PVP控制酚酸氧化效果不如CA,但是预防褐化效果确强于CA,其原因是由于二者抗褐化机理不同所致;VC使用的最适浓度为500 mg·L^(-1),如果超过此浓度会导致外植体褐化率增加;PVP最适浓度为1 000~1 500 mg·L^(-1),CA最适浓度为300 mg·L^(-1)。[结论]天女木兰芽外植体褐化底物是咖啡酸和绿原酸,而与对香豆酸无关;VC、PVP和CA三种抗褐化剂抗褐化机理各不相同,综合比较VC抗褐化效果最好,其次是PVP,CA排在最后。

[ Objective ] To identify the types of phenolic acids that result in bud explants browning of Magnolia sieboldii and determine the most effective anti-browning agent at an optimum concentration. [ Method ] Bud ex- plants were cultured in B5 medium for 30 days. At the beginning, three kinds of anti-browning agents were added separately to the B5 medium. The browning rates were calculated and analyzed. Then the effects of the three inhibi- tors on anti-browning were compared on the basis of the browning rates. In addition, the content variations of caffeic acid, chlorogenic acid, and p-coumaric acid in the bud explants were measured with HPLC using samples regularly selected during tissue culturing. [ Resultl The results showed that the contents of caffeic acid and chlorogenic acid decreased sharply while the content of p-coumaric acid experienced a slight decrease, proving that caffeic acid and chlorogenic acid were more likely to be oxidized than p-coumarie acid. Moreover, the effects of the three inhibitors on anti-browning were sorted in a decreasing order of VC, PVP, and CA. Although PVP was inferior to CA with re- spect to controlling phenolic acid oxidation, its anti-browning effect was more efficient than CA. This result was ex- plained by their different mechanisms for anti-browning. Lastly, the optimal concentration for VC was 500 mg · L^-1, and a higher concentration would result in a greater browning rate of the bud explants. Respective optimal con- centrations for PVP and CA were 1 000 - 1 500 mg· L^-1 and 300 mg· L^-1. [ Conclusion ] The browning sub-strates in bud explants of M. sieboldii are caffeic acid and chlorogenic acid rather than p-coumaric acid. VC, PVP, and CA have distinguishable anti-browning mechanisms and the anti-browning effects of them wiil decrease gradual- ly.