| On the basis of principle of pharmacy, surfactants often used in animal and cytological experiments, such as solubilizer and suspending agent, were adopted in our research on fat-soluble allelochemical bioassay. Solubilizer and suspending agent were helpful to increase the solubility of fat-soluble materials in the water, and therefore, the process of fat-soluble allelochemical bioassay became more secure and accurate. We investigated and evaluated the safety and applicable concentration of Tween-80, Tween-60, Tween-40, Tween-20, Poloxamer188(F-68), Myri52(S-40) and CMC-Na by the experiment of plant seed germination and seedling growth. The results indicated that the Tween-80, Tween-60, Tween-40, Poloxamer188(F-68), Myri52(S-40) were safer than Tween-20 and CMC-Na which produced inhibition effect on seed germination and seedling growth. Security concentration value was respectively 14～1000mg/L at Tween-80, 28～5000mg/L at Tween-60, 31～1000 mg/L at Tween-40, 0.2～4.0% at Poloxamer188(F-68), 0.5% at Myri52(S-40). After comparing the allelopathy activity of vanillin and ferulic acid with solubilization of the Poloxamer188(F-68), Tween-80 and CMC-Na respectively, experiment results showed that each solution with different solubilizer and suspending agent was stable and the concentration didn't also change during the bioassay process. These characteristics of solubilizer and suspending agent mentioned above were advantageous to quantificational study of the allelopathy activity. Based on results mentioned above, Tween-80, Tween-60, Tween-40, Poloxamer188(F-68), Myri52(S-40) were safer on the security, and Tween-80, Tween-40, Poloxamer188(F-68), Myri52(S-40), CMC-Na were better on the effect of solubilization, but the comprehensive effect of Poloxamer188(F-68) was best in all investigated solubilizers and suspending agents.Through the isolation and bioassay of allelochemicals from pericarp of Phellodendron amurense Rupr, we found that the inhibition of the volatile oil was not significant, but the inhibition of 90% ethanol extracts was extremely significant. The 90% ethanol crude extracts from the pericarp of Phellodendron amurense was eluted ordinally from solvent of hexane, chloroform, acetone, 95% ethanol, 50% ethanol, after the active allelochemicals test, we found that 95% ethanol component which extracted from the portion of 90% ethanol extracts contained active allelochemicals, when the concentrantion of the component in 1mL equivalent to pericarp 10 to 125mg, the component could inhibited seeds germination potential and germination rates significantly, as well as inhibit the shoot height, root length, root and over-ground part weight of plant markedly, and with the increase in concentration, enhanced inhibition intensity, in high concentration, it could completely inhibit seed germination and seedling growth. For the 95% ethanol component, separated it by a silica gel chromatography, and tracking its allelopathy activity. In high polar portion, a large number of single compoundα-D-glucose and portion which contain allelopathy activity was separated out. Testing the content ofα-D-glucose in 95% ethanol component with HPLC-ELSD, the content is about 31.8%. In the biological detection, we discovered that seeds extended time required for seed germination which caused byα-D-glucose existence. Meanwhile, as the high-quality carbon existence, it results in a lot of mildew growth to impact seed germination process as well. This phenomenon explained why many seeds rated and lost activity in the course of Phellodendron amurense fruit releasing seeds in previously studies. |
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