【作者】 金丽霞；

【导师】 刘维屏； 林春绵；

【作者基本信息】 浙江工业大学 ， 环境化工， 2012， 博士

【摘要】 农药是人类生产和生活中不可或缺的物资。农药中很多化合物是手性的,目前手性农药已占全部农用化学品的40%,而且这一比例还在逐年增加。手性农药对映异构体之间可能存在活性和毒性方面的差异,因此手性农药的对映体分离与制备具有重要意义。手性农药对映体分离分析是研究对映体选择性差异和环境安全的基础和支撑,手性化合物与手性固定相之间的对映体识别作用机理从理论上为手性分离方法的建立提供技术指导,手性化合物在高效液相色谱中的对映体标识(构型判断)为对映体选择性研究提供基础数据。本论文利用高效液相色谱法,在多糖手性固定相上对手性三唑类杀菌剂和芳氧苯氧丙酸类除草剂进行了对映体分离的研究。(1)在多糖手性固定相Chiralpak AD-H和Chiralcel OD-H( 250×4.6 mm i.d.,5μm)柱上对三唑类手性杀菌剂进行了对映体分离研究。在Chiralpak AD-H柱上,大部分的三唑类化合物得到了完全的基线分离。同时考察了二元流动相体系和三元流动相体系中醇极性添加剂对对映体分离的影响,发现二元体系中异丙醇作为改性剂对此类化合物的分离比乙醇更有优势,而三元体系中流动相能结合两种醇或醇和醚的共同作用进行对映体洗脱。根据圆二色(CD)检测器所得CD光谱图来确定对映体的出峰顺序,考察分离过程中不同醇添加剂时的对映体流出顺序,结果表明流动相中醇添加剂的种类不同则手性化合物对映体流出顺序可能也不同。最后探讨了三唑类化合物在手性固定相上的识别机理。利用分子对接手段,考察了己唑醇等6个杀菌剂在直链淀粉-三(3,5-二甲基苯基氨基甲酸酯)手性固定相(ADMPC)和纤维素-三(3,5-二甲基苯基氨基甲酸酯)手性固定相(CDMPC)上发生识别时的氢键作用,发现除烯唑醇外,其他化合物R/S-构型在CSP上的氢键作用的作用位点存在异同,因此可判断氢键作用是此类化合物在多糖手性固定相上发生手性识别的主要作用之一。(2)在直链淀粉手性固定相Chiralpak AD-H (250×4.6 mm i.d.,5μm)上、正相色谱中,禾草灵等八个芳氧苯氧丙酸类除草剂得到了完全的基线分离,除吡氟氯禾灵在正己烷和异丙醇为流动相时获得分离外,其他化合物均在以正己烷/乙醇(酸形式化合物添加三氟乙酸)为流动相时基线分离。进一步讨论了AOPP除草剂在ADMPC上的对映体识别机理,手性识别主要基于π-π作用、偶极-偶极作用和氢键作用。同样利用分子对接建立此类化合物与ADMPC之间识别的氢键模型,识别中氢键作用主要由化合物上的氧、氮原子和羰基与ADMPC上的-NH之间形成。此外,利用HPLC-UV-CD联用技术、stop-flow法实验扫描得此类化合物CD谱图和正负康顿效应的关系;根据化合物的R/S-结构,利用量子化学法、含时密度泛函理论TDDFT-高斯计算从理论上得到八对对映体各自CD谱图和正负康顿效应的关系,发现此类化合物对映体构型与正负康顿效应的关系均为R-(+)-构型和S-(–)-构型。将两种方法得到CD谱图对比后标识了八个化合物各自对映体在相应色谱条件下的出峰,除吡氟禾草灵、吡氟甲禾灵和吡氟乙禾灵出峰为R>S外,其它均为S>R,由此找到了一种在线判断手性农药对映体绝对构型的方法。(3)从手性固定相填料基质色谱硅胶的制备和表征开始,得到粒径为20μm、比表面积为327.3482 m2/g、孔容为0.577156 cm3/g、孔径为60 ?的色谱硅胶,并将其氨丙基化。进一步合成了纤维素-三(4-甲基苯基甲酸酯)(CTMB)手性固定相并涂覆到自制的色谱硅胶上,匀浆法填装得相应的分析型和半制备型色谱柱。在自制分析型色谱柱上考察了禾草灵除草剂的分离,并将分离条件放大至半制备色谱,结果表明直径为10 mm半制备柱上手动制备每小时可达到mg级,基本能满足对映体靶标活性和非靶标毒性实验研究的需求。更多还原

【Abstract】 Pesticides play a very important role in human life. About 40% of current-used agrochemicals are chiral, and the proportion is increasing for that more and more complex chemicals are introduced. Enantiomers of chiral pesticides always show drastically stereoselective activity and toxicity. Hence, the separation and the purification of single-enantiomer are meaningful. Enantiomeric separations and analysis are the foundation for the study of enantioselectivity and environmental fate. Chiral recognition mechanism between enantiomers and chiral stationary phase (CSP) provides a theoretical guidance for separation. Determinating the elution order by high-performance liquid chromatography (HPLC) with R/S-configurations is basic data for enantioselective studies. All these data are not easy to obtain.Typical chiral triazole fungicides and aryloxyphenoxypropanoic acid (AOPP) herbicides were separated on several polysaccharide CSPs by normal phase HPLC. (1) Enantiomeric separations of 23 triazole fungicides were conducted on two polysaccharide CSPs (Chiralpak AD-H and Chiralcel OD-H columns, 250×4.6 mm i.d., 5μm). For most of the compounds, baseline separations were obtained on Chiralpak AD-H (resolution RS>1.5). Effect of mobile phase composition were evaluated on a binary and a ternary mobile phase systems, respectively. On the binary mobile phase system, isopropanol as the organic modifier showed better performance than ethanol. On the ternary system, the combined action of two modifiers could be adopted to the elution. The resolved enantiomers were distinguished by the signals of circular dichroism (CD) detector. The elution reversal was obtained when different alcohols (ethanol, isopropanol and/or methanol) were used in the mobile phase. Finally, the hydrogen-bonding interactions on chiral discriminations between triazole fungicides and the polysaccharide CSPs (ADMPC and CDMPC) were discussed with a molecular docking treatment. Except diniconazole, the hydrogen-bonding interactions of R/S-enantiomers occurred in different sites between the enantiomers and the CSPs. It was deduced that the hydrogen-bonding interaction is one of the main interactions for the discrimination of these fungicides.(2) Eight AOPP herbicides were well resolved on a Chiralpak AD-H column (250×4.6 mm i.d., 5μm) by a normal phase HPLC. The resolution was occurred with an n-hexane/ethanol mixture as the mobile phase (trifluoroacetic acid as the additive for acidic compounds) except haloxyfop with an n-hexane/isopropanol/TFA mixture. Chiral discriminations were mainly occurred with theπ-πinteractions, the dipole-dipole interactions and the hydrogen-bonding interactions. Besides, with the molecular docking treatment, it was deduced that the hydrogen-bonding interactions referred to the O, N atoms and carbonyl groups of enantiomers interacted with–NH group of ADMPC CSP. Furthermore, a method using HPLC tandem ultraviolet (UV) detector and CD detector was used to determine the elution order of R/S-enantiomers. Experimental CD-Cotton effect spectra of R/S-configurations were scanned in a stop-flow experiment. According to a quantum chemistry method, theoretical CD-Cotton effect spectra of R/S-configurations were calculated and simulated, and it was found that the R-configurations of these herbicides were corresponding to a positive-Cotton effect as R-(+)-enantiomer, and S-configurations were corresponding to a negative-Cotton effect as S-(–)-enantiomer, respectively. Comparison of the experimental CD and computed CD spectra indicated that the elution order of these herbicides were R>S for fluazifop-butyl, haloxyfop-methyl and haloxyfop-2-ethoxyethyl, and S>R for other AOPP herbicides under the specified separation conditions.(3) Silica gel with appropriated particle size (20μm), appropriated BET surface area (327.3482 m~2/g), pore volume (0.577156 cm~3/g) and pore size (60 ?) were synthesized as the carrier of chromatographic stuffing after reacted withγ-aminopropyltriethoxysilane. Moreover, Cellulose tris(4-methylbenzoate) (coated-CTMB) CSP was synthesized and then coated onto silica gel. Both an analytical column (250×4.6 mm i.d., 20μm) and a semi-prepared one (250×10 mm i.d., 20μm) were packed. Enantiomeric separation of diclofop-methyl was investigated on the analytical column, and the sufficient separation was obtained using n-hexane/isopropanol 85/15 as the mobile phase (RS 2.95). The elution order was R-(+)-diclofop-methyl>S-(–)-diclofop-methyl. By magnifying the column specification to a semi-one, the pure-enantiomer was purified with a handling collection treatment. The purification could meet a production with mg/h level.更多还原