利用激光诱导纳秒时间分辨荧光(laser-induced nanosecond time-resolved fluorescence, LITRF)猝灭法原位研究腐植酸(humic acid, HA)分别与母环多环芳烃(polycyclic aromatic hydrocarbons, PAHs)菲(phenanthrene, Phe)及烷基PAHs 9-乙基菲(9-Ethylphenanthrene, 9-EP)和惹稀(retene, Ret)相互作用, 考察HA对母环及烷基PAHs结合特性差异与机制, 对了解PAHs环境行为及生物有效性有重要意义。 结果表明, 通过改变延迟时间(50 ns)可有效消除HA荧光干扰, 实现游离Phe, 9-EP及Ret浓度直接测定。 利用Freundlich非线性等温吸附模型描述Phe, 9-EP和Ret与HA结合特性, LITRF猝灭法与传统荧光法获得的模型拟合参数及单点结合系数结果一致。 其中, 参数n小于1, 表明Phe, 9-EP及Ret与HA均以非线性形式结合, 且9-EP和Ret非线性程度高于Phe; 相同给定平衡浓度下, HA与9-EP和Ret单点结合系数KOC大于Phe, 而9-EP和Ret结合能力相近, 且PAHs与HA结合系数均随给定浓度增加而降低。 疏水性、 取代基及与HA疏水空腔适应能力决定特定PAHs与HA结合特性。 通过荧光寿命分析, HA存在下Phe, 9-EP和Ret寿命分别为36.90, 35.34和35.13 ns, 与未加入HA时的36.36, 35.34和35.84 ns无明显差异, 表明Phe, 9-EP和Ret与HA间的荧光猝灭以静态过程为主。 LITRF猝灭法可快速有效原位研究PAHs与HA相互作用, 有助于实现PAHs生态风险原位评估。

Investigations both on the binding properties and mechanisms of parent polycyclic aromatic hydrocarbons (PAHs) and alkyl PAHs with humic acid (HA) are crucial to understand the environment behavior and bioavailability of PAHs in aquatic ecosystems. Thus, a novel approach for in situ investigation of the interactions between parent PAHs and alkyl PAHs, exemplified by Phenanthrene (Phe), 9-Ethylphenanthrene (9-EP) and Retene (Ret), and HA was established by using fluorescence quenching method with laser-induced nanosecond time-resolved fluorescence (LITRF). By changing the delay time of 50 ns, the fluorescent interference of HA can be eliminated effectively and the concentrations of free Phe, 9-EP and Ret can be determined directly. The binding characteristics of the dissolved HA and Phe, 9-EP and Ret were described by Freundlich nonlinear isothermal model. The results of the model parameters and the single point binding coefficients KOC of Phe, 9-EP and Ret with HA by LITRF quenching method were consistent with those of the conventional fluorescence quenching method. Nonlinear bindings of Phe, 9-EP and Ret to HA were also observed because of the parameter n values below 1, and the degree of nonlinearity of Phe was lower than 9-EP and Ret duo to the n value of Phe larger than 9-EP and Ret. At a same given equilibrium concentration, the KOC of Phe was lower than 9-EP and Ret, and that of 9-EP was near to Ret. The binding affinity of the three PAHs increased with decreasing the equilibrium concentration. The binding characteristics of PAHs with HA largely depends on their hydrophobicity, substituent groups and its ability to fit into hydrophobic cavities in HA. The fluorescence lifetimes of Phe, 9-EP and Ret in presence of HA were 36.90, 35.34 and 35.13 ns, meanwhile the fluorescence lifetimes of Phe, 9-EP and Ret in absence of HA were 36.36, 35.34 and 35.84 ns. There was no significant difference of three PAHs fluorescence lifetime with or without HA, indicating the quenching mechanism for Phe, 9-EP and Ret with HA were primarily static quenching. The LITRF quenching method could be used to in situ explore the interactions between PAHs and HA, which could be contribute to realize the PAHs risk assessment in real time.