【作者】 宋立岩；

【导师】 花日茂； 岳永德；

【作者基本信息】 安徽农业大学 ， 农药学， 2003， 硕士

【摘要】 本研究取安徽华星化工有限公司精噁唑禾草灵废水处理系统进水口处污泥进行驯化培养，从中筛选出活性菌株，对其中降解精噁唑禾草灵效果最好的活性菌株进行了鉴定以及UV诱变，并以此菌株以及此菌株UV诱变菌株（诱变菌株中降解精噁唑禾草灵效果最好）为出发菌株，运用高效液相色谱分析精噁唑禾草灵残存量的方法，进行了降解动力学以及外源物质对降解菌活性影响的研究，并通过高效薄层色谱方法，对精噁唑禾草灵的降解产物进行了分离。主要研究结果如下： 1．取安徽华星化工有限公司精噁唑禾草灵废水处理系统进水口处污泥进行驯化培养，分离到8株有效菌株，通过高效液相色谱分析这8株菌株降解精噁唑禾草灵的残存量，确定其中降解精噁唑禾草灵最好的一株菌株为本实验菌株，通过传统的微生物鉴定方法，将其鉴定为产碱菌属（Alcaligenes sp），标记为Alcaligenes sp.H。 2．以Alcaligenes sp.H为出发菌株，通过紫外诱变后，从中挑选降解精噁唑禾草灵效果最好的诱变株H8-1为本实验的第二株菌株，标记为Alcaligenes sp.H_uv。在精噁唑禾草灵浓度为5-100mg／L范围内，Alcaligenes sp.H_uv菌降解精噁唑禾草灵的效果优于Alcaligenes sp.H。 3．本实验中Alcaligenes sp.H和Alcaligenes sp.H_uv以及混合菌降解精噁唑禾草灵的降解动力学方程用一级动力学方程模拟，其相关系数r均大于0.90。 C_t=C₀e^-kt 4．Alcaligenes sp.H与Alcaligenes sp.H_uv不同混合比例（2.5：1，1：1，1：2.5）的混合菌降解效果均明显优于Alcaligenes sp.H单菌，其半衰期分别为37.66h、30.39h、23.90h均明显小于Alcaligenes sp.H单菌的半衰期64.17h，而当Alcaligenes sp.H_uv在混合菌体系中所占比例超过一半时，混合菌的降解效果优于Alcaligenes sp.H_uv单菌（半衰期为35.18h），同时，随着Alcaligenes sp.H与Alcaligenes sp.H_uv混合比例的下降，即Alcaligenes sp.H_uv菌的比例增大，降解加快，这一点可以从半衰期的逐渐减小得到反映，说明Alcaligenes sp.H_uv菌在混合菌体系中占主导地位，也间接反映其降解效果优于Alcaligenes sp.H。 5．Alcaligenes sp.H菌量分别为OD₃₃₀=0.2，0.4，0.6时，精噁唑禾草灵的微生物降解半衰期分别为67.28h、35.0h、21.72h，其半衰期与菌量近似成反相关，降解速率常数与Alcaligenes sp.H菌量近似成正比。而Alcaligenes sp.H_uv菌量分别为OD=0.2，0.4，0.6时，精噁唑禾草灵的微生物降解半衰期为35.54h、21.32h、19.58h，虽然其降解速率与Alcaligenes sp.H_uv菌量呈正相关，但其反映的菌量浓度差并没有Alcaligenes sp.H大，可见对于Alcaligenes sp.H_uv来说并非所有的菌都参与了降解(相对OD₃₃₀=0.2)，说明对于一定浓度的精噁唑禾草灵来说，随着降解菌菌量的增加，农药的降解加快，降解速率与菌量基本成正相关，但同样存在一个最佳菌量问题，即当菌量增加到一定程度后，再增加菌量，其降解速率增加趋势不再明显。 6．在精嗯哇禾草灵的浓度从 sing／L到 100mg／卜变化范围内，对于 Alcalfgenes Sp．H来说，其半衰期逐渐增大，分别为47．47h、65．38h、68．6lh、86．63h、135．88h，对于Alcaliggnes sp．Huv来说其半衰期分另为 18．24h、35．54h、46．51h、57．27h、72．19h；半衰期同样逐渐增大。不同精嗯哇禾草灵浓度对降解动力学的影响表明：AICJ人卯们“sp．H及 Alcallee，7es s H。v只对一定浓度的精噬哇禾草灵保持较强的降解作用，A儿7rnes sn·H及 Alcaliernes sn Huv降解高浓度的精曙哩禾草灵的效率下降。 7．本实验所选取的 3种外源物质对Aleal＠nes Sp．H及Alcalwnes Sp．HW降解精嗯哇禾草灵均产生了抑制作用。其中百菌清对 AlcaligeneS SP．HW及 Alcaligenes Sp丑与Alcaligenes Sp．H＊混合菌门：1)降解精嗯哇禾草灵的抑制作用更为显著，而对于AlcaligenesspH其相对抑制作用较小；当外源物质为十二烷基苯磺酸钠和 Cr＋6时，降解曲线形状发生了明显的变化，尤其降解初期十＝烷基苯磺酸钠和Cr＋‘对降解菌的抑制作用尤为明显，降解速度很慢，随着降解菌对十＝烷基苯磺酸钠和 Cr＋0的适应，降解速度逐渐加快。8．在展开剂为甲苯：H氯甲烷（V：V，7：3）条件下，精嗯哇禾草灵降解产物得到很好的分离，通过比较精嗓哇禾草灵标样及精嗯陛禾草灵降解产物的展开距离即R。值，除Rf叫．22（精嗯哇禾草灵标样峰）的峰外，精嗯哩禾草灵降解产物（降解菌为Ale。lfgenes SP．H）在 Rr＝0．14、0．29、0．34、0．39、0．47处有 5个峰；精lg陛禾草灵降解产物（降解菌为 Alcalfgunes sp．Hm）在 Rr＝0．11、0．39处有 2个峰，这与 CAMAG紫外灯下观察的斑点数量相符。通过 HPLC－MS分析可以推测出：m)《三－门－氯J，3－苯并嗯陛千－基氧)苯氧基」丙酸乙酯水解是降解菌 Alcallernes p丑与 AlcalfgenesSP．Huv降解精嗯咆禾草灵的公共降解途径之一，其产物为m)六二－历－氯习，卜苯并嗯哇－2－基氧)苯氧基」丙酸和乙醇。更多还原

【Abstract】 Bacterial strains which could decompose fenoxaprop-p-ethyl were isolated from a batch culture with fenoxaprop-p-ethyl as carbon source and the sludge from waste water dealing system of Huaxing chemical engineering Co L.td . After determining the rest quantity of fenoxaprop-p-ethyl with HPLC , H was selected as dominant degradation bacterial strain for its best degradation ability to fenoxaprop-p-ethyl . H8-1 was also selected as another dominant degradation bacterial strain for its higher degradation ability to fenoxaprop-p-ethyl than the other strains after UV mutation . Fenoxaprop-p-ethyl degradation dynamics was studied with HPLC method , and degradation products of fenoxaprop-p-ethyl had been separated with HPTLC . The main study results are as follows :1 .Eight bacterial strains which could decompose fenoxaprop-p-ethyl were isolated from a batch culture with fenoxaprop-p-ethyl as carbon source and the sludge from waste water dealing system of Huaxing chemical engineering Co L.td . After determining the rest quantity of fenoxaprop-p-ethyl with HPLC , H was selected as dominant degradation bacterial strain for its best degradation ability to fenoxaprop-p-ethyl .Based on the morphological appearances and physiological characteristics ,H was identified as Alcaligenes sp and named as Alcaligenes sp.H .2.H8-1 was also selected as another dominant degradation bacterial strain for its higher degradation ability to fenoxaprop-p-ethyl than the other strains after UV induce . H8-1 was marked as Alcaligenes sp. Huv . With the concentration of fenoxaprop-p-ethyl varing from 5mg/L to 100mg/L , Alcaligenes sp. Huvhas better degradation ability to fenoxaprop-p-ethyl than Alcaligenes sp.H .3. fenoxaprop-p-ethyl degradation dynamics was studied . Degradation kinetics follow the first order kinetics .4. It proved that degradation ability of mixed bacterial by Alcaligenes sp.H and Alcaligenes sp. Huv to fenoxaprop-p-ethyl is superior to Alcaligenes sp.H , superior to Alcaligenes sp. Huv after the quantity of Alcaligenes sp. Huv surpasses half , and with the quantity increasing of Alcaligenes sp. Huv , degradation ability of mixed bacterial by and Alcaligenes sp. Huv to fenoxaprop-p-ethyl increases respectively comparing their half-time . Their half-times are 37.66h, 30.39h and 23.90h to mixed bacterial (Alcaligenes sp.H : Alcaligenes sp. Huv 2.5:1,1:1,1:2.5 ) ,64.17h to Alcaligenes sp.H and 35.18h to Alcaligenes sp. Huv.5. Degradation rates of fenoxaprop-p-ethyl were positively correlative to biomass .On thecontrary , the fortified concentration is opposite the degradative rate . It can be seen from their half-time and their half-time are 67.28h, 35.0h and 21.72h to Alcaligenes sp.H , 35.54h ,21.32h and 19.58h to Alcaligenes sp. Huv correlative to their different biomass .Results showed that biomass has a appropriately fortified concentration .When surpassing this biomass , the degradative rate can not grow obviously .6. With the concentration of fenoxaprop-p-ethyl varing from 5mg/L to 100mg/L , the half-time of Alcaligenes sp.H and Alcaligenes sp. Huv increased respectively .The different half-time of Alcaligenes sp.H and Alcaligenes sp. Huv with different concentration are 47.47h,65.38h,68.61h,86.63h,135.88h and 18.24h,35.54h,46.51h,57.27h,72.19h .Effects of different concentration of fenoxaprop-p-ethyl to degradation showed that Alcaligenes sp.H and Alcaligenes sp. Huv had better degradation in some a concentration and the degradation ability to fenoxaprop-p-ethyl of Alcaligenes sp.H and Alcaligenes sp. Huv was obviously decreased in high concentration.7. Chlorothalonil, dodecylbenfenxaprop -p-ethylene sulfonic aid sodium salt and Cr+6can inhibite the degradation ability of Alcaligenes sp.H , Alcaligenes sp. Huv and the mixed bacterial by Alcaligenes sp.H and Alcaligenes sp. Huv to fenoxaprop-p-ethyl. The degradation ability of Alcaligenes sp.H to fenoxaprop-p-ethyl was severely inhibited by Chlorothalonil. In early period , Alcaligenes sp.H , Alcaligenes sp. an更多还原