遠紅光及氮素施用量對組織培養夏枯草生理及二次代謝物之影響

Abstract

夏枯草 (Prunella vulgaris) 為唇形科中藥材植物，有長久的應用歷史，現代醫學也多有關於其藥理價值的研究。遠紅光雖非光合有效輻射的波段範圍，但對植物形態及二次代謝物有所影響。氮素等植物營養對植物的生長與二次代謝物的累積亦有相關。本次研究透過植物組織培養方式，以對照 (CK) 及強遠紅光 (FR) 兩種光照環境，以及全劑量 (1N) 、½倍劑量 (½N) 及¼倍劑量 (¼N) 氮素等三種MS培養基，探討遠紅光與氮素施用量，對組織培養夏枯草形態、光合色素及二次代謝物生合成的影響，並測定夏枯草甲醇萃取物抗氧化能力，分析二次代謝物與抗氧化能力的關聯。結果顯示增加遠紅光無增加夏枯草的葉片對數、鮮重與乾重，而氮素則增加乾重。相同氮素施用量下，強遠紅光會降低葉綠素、胡蘿蔔素與葉綠素崩解產物；同光照處理中，氮素較多的組別則有較高的葉綠素及胡蘿蔔素含量，且兩種環境條件會改變不同光合色素間的比值。二次代謝物中總酚、芸香素與迷迭香酸含量會隨氮素與遠紅光的增加而提升，以FR + 1N有最多，分別有總酚 (44.73 mg Gallic acid equivalent g-1 DW) 、芸香素 (0.18 mg g-1 DW) 及迷迭香酸 (20.43 mg g-1 DW) ，CK + 1N有最多的總類黃酮含量 (105.57 mg Rutin equivalent g-1 DW) ，咖啡酸含量在各組間差異不顯著。以半抑制濃度(IC50)比較各組別的抗氧化能力的結果中，FR + 1N在DPPH清除自由基能力 (1.95 mg ml-1) 與還原能力 (2.33 mg ml-1) 的測定上皆有最低的IC50，表示其有最佳的抗氧化能力。以相關性分析得知此兩種抗氧化能力與酚類和迷迭香酸含量有顯著相關。以上結果顯示，在適量的氮素條件下添加額外的遠紅光，有助於增加夏枯草的二次代謝物含量及其抗氧化活性，增加其價值，可供未來穩定生產夏枯草之環境條件參考。

Prunella vulgaris is a traditional chinese medicinal plant of the Lamiaceae family and being used for a long time. There is many modern medicine research about its pharmacological value. The wavelength of far red light is not in the range of photosynthetically active radiation, but it could influence plant growth and secondary metabolites, so does nitrogen, which is an important plant nutrient. In this study, the effect of two light conditions, control (CK) and high far red light (FR) , and MS medium with three nitrogen levels, full (1N) , half (½N) and quarter nitrogen (¼N) , on morphology, photosynthetic pigments, secondary metabolites of in vitro Prunella vulgaris were explored, the antioxidant activities of its methanol extract were determined, and the relationship between secondary metabolites content and antioxidant activities were also analyzed. The result showed that high far red light did not increase the number of leaf pairs, fresh weight and dried weight. The nitrogen increased dried weight. Within the same nitrogen level, the chlorophylls, carotenoids and chlorophyll degradation pigments content decreased in the high far red light treatment. Under the same lighting condition, there were more chlorophyll and carotenoids content with increasing nitrogen level. The light and nitrogen treatment changed the ratio between different photosynthetic pigments. The content of total phenolic, rutin, and rosmarinic acid were increased with the elevated nitrogen level and the far red light, and the maximum content of these metabolites were observed under FR + 1N, which with total phenolic 44.73 mg Gallic acid equivalent g-1 DW, rutin 0.18 mg g-1 DW, and rosmarinic acid 20.43 mg g-1 DW, respectively. There was maximum content of flavonoid under CK + 1N (105.57 mg Rutin equivalent g-1 DW) . The content of caffeic acid among all groups was insignificantly different. Compared antioxidant activity among groups via half maximal inhibitory concentration (IC50), the result indicated that there were best DPPH free-radical scavenging activity (1.95 mg ml-1) and reducing power (2.33 mg ml-1) in FR + 1N. These two antioxidant activities above showed significantly correlated with the content of total phenolic and rosmarinic acid through correlation analysis. The results of this study showed that under appropriate nitrogen level with high far red light would benefit the increase in secondary metabolites and antioxidant activity in Prunella vulgaris to enhance its value, and could be a reference of the environmental condition when producing Prunella vulgaris stably in the future.