酚類化合物之生物分解

Abstract

植物次階代謝的產物如酚類化合物等,在自然界中常扮演著植物相剋 作用的角色,台灣蔗田土壤中含有ferulic acid、p-hydroxybenzoic acid 、p-coumaric acid、vanillic acid、syringic acid 等酚酸引起蔗作減 產現象。 此外台灣二期稻作減產及蘆筍減產,其缺株率高達40%證實 為3,4-dihydroxybenzoic acid等酚酸所引起,然而甚少有從微生物的觀點 著手,解決土壤中酚類化合物所引起的問題。 本研究的目的即在篩選具有 分解酚酸能力的細菌來解決土壤酚酸所引起的植物相剋作用。 本實驗分 別以 ferulic acid 及p-hydroxybenzoic acid 為唯一碳源所篩選出來的 分解菌,各別為486f、486p菌種;486f菌對於ferulic acid 100 ppm之分解 能力第10天總移出率為86%,第3天殘餘百分率為45%(分解率55%);486p對於 p-hydroxybenzoic acid 100 ppm之分解能力第7天殘餘百分率為51%(分解 率為49%),第12天總移出率為67%。 以Pseudomonas putida作為已知菌種 分別與486f及486p菌種比較其分解率,由實驗顯示在第7天Pseudomonas putida對p-hydroxybenzoic acid之分解率已達100%,而486p之分解率僅 為49%(殘餘百分率為51%),顯然,Pseudomonas putida其分解能力大於486p 。而486f對於ferulic acid之解第7天其殘餘百分率為21%,而Pseudomonas putida為48%,由結果所示486f其分解能力大於Pseudomonas putida。 在 酚酸的代謝實驗中以50, 100, 1000 ppm不同的濃度作為觀察,發現低濃度 下其代謝速率快,且完全分解;高濃度將造成抑制作用,而影響分解作用 在 添加葡萄糖作為外來碳源對酚酸分解作用的影響實驗中,結果顯示:分解菌 能利用葡萄糖作為外來碳源,且加速分解,但不能完全分解;若添加土壤溶 液則不但加速分解,且分解完全,分解速率優於添加葡萄糖,推測為分解菌 與土壤微生物之協同代謝作用之結果。 胞內酚酸測定之實驗結果顯示兩 株分解菌都有胞內酚酸之存在,且486f胞內酚酸測定中發現有vanillic acid之中間代謝產物的存在。酚酸毒性試驗的結果顯示,200 ppm的 ferulic acid已完全抑制萵苣的生長,而p-hydroxybenzoic acid其抑制率 僅達44.4%,得知不同的植物毒物質其毒性將有差異。本實驗中之生物分析 結果顯示,在接菌後第一天種植萵苣已經得到改善的效果,推測此由於二水 土壤有機碳源高,細菌快速分解之下的結果,因此土壤性質與微生物之間的 關係將影響微生物的活動。

The products of botanical serial metabolism, phenolic compounds for instance, often play a role of allelopathy in the nature. The soil of sugar cane field in Taiwan contains phenolic acids, such as ferulic acid, P-hydroxybenzoic acid, p-coumaric acid, vanillic acid, syringic acid which cause decrease in sugar cane production. Besides, it is verified that decreases in the production of semiannual rice crops and asparagus whose rate of null plants reach as heigh as 40%, are caused by phenolic compounds, 3,4-dihydroxybenzoic acid, through few research treat these problems induced by phenolic compounds in the soil from micro-biological points of view. The purpose of this research is to solve the problem of allelopathy brought about by phenolic acids existing in the soil by sifting out bacteria that are capable of resolving phenolic acids. This experiment separately employs resolving bacteria, species of 486f and 486p that are shifted out by way of using ferulic acid and p- hydroxbenzoic acid as the sole carbon source. The total removal rate of the resolving capacity of Bacterium 486f against ferulic acid 100ppm on the 10th day is 86%, on the 3rd the remaining percentage is 45% (resolving rate 55%); the resolving capacity of Bacterium 486p against p-hydroxybenzoic acid 100 ppm has the remaining percentage of 51% on 7th day (resolving rate 49%), and the total removal rate is 67%. The resolving rate of Pseudomonas putida is compared separately with those 486f ande 486p, the afore-mentioned species of bacteria. The experiment inidicates the resolving rate of Pseudomonas putida against p- hydroxybenzoic acid on 7th day reaches 100%, while that of 486p is only 49% (remaining percentage is 51%). The resolving capability of Pseudomonas putida is evidently larger than that of 486p. The resolving power of 486f against ferulic acid shows its remaining percentage at 21% on the 7th day while the Pseudomonas putida is 48% from the results it is known that Pseudomonas putida. In the experiment of phenolic acid metabolism, different densities of 50, 100, and 1000 ppm are separately observed, finding that not only the speed of metabolism is fast at the low density but the resolution is complete; while at the height density the function is refrained and the resolving affected. In the exsperiment using glucose as foreign carbon source to test influence of phenolic acid resolution function,the result shows that the resolving bacteria can make use of glucose as foreign carbon source and accelerate the resolution, through incomplete; then, if added with soil solution, it does not only accelerates the resolution faster but also does the work perfectly. The rate of resolving speeds is evidently superior when glucose is added in, which is inferred to be the consequence of synergistic function between resolving bacteria and soil microorganisms. The result of experiment of endocytic phenolic acid measurement apparently indicates that both the resolving bacteria have endocytic phenolic acid, and in the measurement of 486f endocytic phenolic acid, existence of mid-metabolic procuct of vanillic acid was found. The result of phenolic acid toxicity test indicates that ferulic acid completely suppress growth of lettuce at 200 ppm, and the rate of suppression of p-hydroxybenzoic acid only reaches 44.4%, telling us that the toxicity of different plant toxins differ. The result of the experiment of biological analysis shows that the lettuce gets improvement on the first day it is planted after receiving the bacteria treatment, which is conjectured to be the result of bacteria's fast resolving, for the organic carbon contained in Erhshui Soil is high. Therefore, the relation between the nature of soil and the microorganisms influence the activities of the microorganisms.