Ammonium is the most energetically favorable form of inorganic nitrogen that phytoplankton can take up, and its availability often limits phytoplankton biomass. Recently, the majority of methods for quantification of ammonium at nanomolar concentrations have used the reaction between ammonium and o-phthaldialdehyde (OPA). Two different approaches have been employed. One involves gas diffusion of ammonia across a membrane. The other entails a more direct approach and utilizes sulfite as a reducing agent to form a fluorescent product. A benefit of the separation technique is that natural fluorescence of the samples does not yield a false positive signal, however membrane failure and clogging are challenging analytical problems. The direct reaction between the analyte and OPA, although devoid of problems associated with membranes, requires a correction for background fluorescence. This work presents the development of a reverse flow injection analysis (rFIA) method with automatic background fluorescence correction and a low detection limit, along with its application to measure underway ammonium concentrations in Tampa Bay during a bloom of the toxic dinoflagellate Karenia brevis with concurrent fish kills in June of 2021. During field testing elevated concentrations of ammonium were found to coincide with the largest number of observed floating dead fish and lethal K. brevis concentrations. The method employs a sulfite-formaldehyde reagent mixed with the sample into which an OPA reagent is injected and heated to activate fluorescence. Fluorescence of the sample is measured before the injection of OPA and at the peak of OPA injection, allowing for differentiation of the background fluorescence from the signal proportional to analyte. The detection limit and the limit of quantitation for this technique are 2.3 nM and 7.5 nM, respectively, and the coefficient of variation is 0.6% for replicate deionized water blanks (N = 10). Advantages of this method compared to techniques capable of both nanomolar detection of ammonium and improved accuracy via background fluorescence correction include a greater sample throughput, an increase in reported useful reagent lifetime, and the ability to quantify ammonium in coastal waters.

铵是浮游植物可以吸收的最有利的无机氮形式，其可用性通常会限制浮游植物的生物量。最近，大多数纳摩尔浓度铵的定量方法都使用了铵和邻苯二甲醛 (OPA) 之间的反应。已经采用了两种不同的方法。一种涉及气体扩散氨穿过膜。另一种需要更直接的方法，并利用亚硫酸盐作为还原剂来形成荧光产物。分离技术的一个好处是样品的自然荧光不会产生假阳性信号，但是膜失效和堵塞是具有挑战性的分析问题。分析物和 OPA 之间的直接反应虽然没有与膜相关的问题，但需要对背景荧光进行校正。这项工作介绍了一种反向流动注射分析(rFIA) 方法的开发，该方法具有自动背景荧光校正和低检测限，以及其在有毒甲藻Karenia brevis2021 年 6 月同时发生鱼类死亡。在现场测试期间，发现升高的铵浓度与观察到的最大数量的漂浮死鱼和致命短杆菌浓度相吻合。该方法使用与样品混合的亚硫酸盐-甲醛试剂，其中注入 OPA 试剂并加热以激活荧光。在注入 OPA 之前和 OPA 注入的峰值处测量样品的荧光，以便将背景荧光与与分析物成比例的信号区分开来。该技术的检测限和定量限分别为 2.3 nM 和 7.5 nM，重复去离子水空白的变异系数为 0.6% ( N = 10)。与能够以纳摩尔检测铵并通过背景荧光校正提高准确性的技术相比，该方法的优势包括更大的样品通量、延长报告的有用试剂寿命以及量化沿海水域中铵的能力。