Flow-Batch Sample Preparation for Fractionation of the Stress Signaling Phytohormone Salicylic Acid in Fresh Leaves.

Salicylic acid (SA) is an important stress signaling phytohormone and plays an essential role in physiological processes in plants. SA fractionation has been carried out batchwise, which is not compatible with the high analytical demand in agronomical studies and increases susceptibility to analytical errors. In this context, a novel flow-batch sample preparation system for SA fractionation on fresh plant leaves was developed. It was based on microwave-assisted extraction with water and conversion of the conjugated species to free SA by alkaline hydrolysis. Free and total SA were quantified by fluorimetry after separation by sequential injection chromatography in a C18 monolithic column. The proposed procedure is directly applicable to plant leaves containing up 16 mg kg-1 SA, with a limit of detection of 0.1 mg kg-1 of SA, coefficient of variation of 3.0% (n = 10), and sampling rate of 4 samples h-1. The flow-batch sample preparation system was successfully applied to SA fractionation in sugarcane, corn, and soybean leaves without clogging or increasing in backpressure. The proposed approach is simple, less time-consuming, and more environmentally friendly in comparison to batchwise procedures.

Exploitation of a short monolithic column for in-line separation and preconcentration: Environmental friendly determination of the emerging pollutant salicylic acid in natural waters.

Salicylic acid is an emerging environmental contaminant, usually found at ng/L concentrations in natural waters. Its quantification usually involves liquid chromatography-tandem mass spectrometry, which requires complex and costly instrumentation as well as time-consuming sample pretreatment, typically involving large solvent volumes. In this work, sequential injection chromatography was exploited to develop a fast, green, cost-effective, and highly sensitive procedure for fluorimetric determination of salicylic acid in natural waters. Analyte preconcentration directly on the chromatographic column (on-column preconcentration) was exploited to improve detectability, yielding an enrichment factor of 122 (1.75 mL of sample) and takes only 8.5 min per determination. A detection limit of 20 ng/L, a linear response range from 0.06 to 5.00 µg/L, coefficients of variation lower than 3.0% (n = 10), and recoveries within 86 and 114% were estimated. The procedure was applied for the analyses of freshwater samples and results agreed with those obtained by liquid chromatography-tandem mass spectrometry at the 95% confidence level. The proposed procedure encompasses in-line concentration, isolation/separation, and detection, without the need for sample clean-up, thus minimizing the consumption of organic solvents and risk of analyte losse.

Dual thermostating in flow analysis.

An advanced strategy involving concentric tubes is proposed for fast and controlled heating (or cooling) of the reaction medium in flow analysis. Different temperatures are set by sequentially circulating two thermostated water streams through the outer larged bore (2.0mm i.d.) silicone tube, which acted as a water-jacket of the inner (0.8mm i.d.) PTFE tube, and directing the sample zone to flow through it. Each end of the outer tube is connected to a three-way valve that selects the stream to flow inside it. For 25-85cm tube lengths and a 12.0mLmin-1 flow rate, the time interval required for temperature attainment, and the uniformity of temperature along the tube were evaluated. For the 85-cm tube, low differences in temperatures along the coil (1.1-8.7°C) and between programmed and attained values (2.3-13.4°C) were noted within a wide range of pre-set temperatures (15-75°C). The feasibility of the innovation in flow analysis was demonstrated in a model system relying on the iodide-nitrite reaction. The strategy allows fast (15-120s) thermostating of the reaction medium in a versatile and simple way, and is especially attractive when two controlled temperatures are set during the analytical course. Potentialities and limitations of the innovation are discussed.

Tracer-monitored flow titrations.

The feasibility of implementing tracer-monitored titrations in a flow system is demonstrated. A dye tracer is used to estimate the instant sample and titrant volumetric fractions without the need for volume, mass or peak width measurements. The approach was applied to spectrophotometric flow titrations involving variations of sample and titrant flow-rates (i.e. triangle programmed technique) or concentration gradients established along the sample zone (i.e. flow injection system). Both strategies required simultaneous monitoring of two absorbing species, namely the titration indicator and the dye tracer. Mixing conditions were improved by placing a chamber with mechanical stirring in the analytical path aiming at to minimize diffusional effects. Unlike most of flow-based titrations, the innovation is considered as a true titration, as it does not require a calibration curve thus complying with IUPAC definition. As an application, acidity evaluation in vinegars involving titration with sodium hydroxide was selected. Phenolphthalein and brilliant blue FCF were used as indicator and dye tracer, respectively. Effects of sample volume, titrand/titrant concentrations and flow rates were investigated aiming at improved accuracy and precision. Results were reliable and in agreement with those obtained by a reference titration procedure.

Pulsed flows in flow analysis: Potentialities, limitations and applications.

In flow analysis, use of a steady and pulseless flow was considered essential for ensuring a reproducible handling of the flowing sample. To this end, peristaltic and syringe pumps have been the propelling device in the vast majority of the flow analysers. Recently, the number of applications involving pulsed flow has been increasing. Most of them refer to use of solenoid pumps, the essence of the so-called multi-pumping flow systems. This review critically discusses the characteristics, potentialities and limitations of the pulsed flow systems, emphasizing the main advantageous characteristics of the streams involved, such as high radial mass transference and good mixing of the fluids. Diverse contributions ranging from instrumentation development to analytical applications are presented.

Flow analysis in Brazil: contributions over the last four decades.

The main contributions of Brazilian researchers to the field of flow analysis are reviewed, with an emphasis on historical developments, conceptual aspects, system design, and analytical applications. Contributions after the advent of flow injection analysis are highlighted. Novel approaches (e.g. zone merging, zone sampling, zone trapping, multi-site detection, and multi-commutation), flow modalities (e.g. monosegmented flow analysis, flow-batch analysis, multi-pumping flow analysis), as well as the pioneering implementation of different detection techniques (e.g. potentiometry, turbidimetry, flame atomic absorption spectrometry, inductively coupled plasma-optical emission spectrometry, and gravimetry) and analytical steps (e.g. titrations, membrane-less gas diffusion, and electrolytic dissolution) are highlighted. Strategies to improve analytical figures of merit and the use of the flow analyser as a tool for teaching purposes are also discussed. Contributions from Brazilian workers in the context of system miniaturization, "green" chemistry, analysis of complex samples, novel strategies and materials for in-line analyte separation/concentration, and proposals for expert systems are also highlighted. The large-scale analysis of samples of agronomical, environmental, industrial, and clinical relevance is emphasized.

A flow system with zone merging and zone trapping in the main reactor applied to spectrophotometric catalytic determination of cobalt in grasses.

A flow system with zone merging and zone trapping in the main reactor was proposed. The sample and reagent inserted aliquots merge together and the resulting zone is directed towards a displaceable reactor inside which its most concentrated portion is trapped. After the pre-set TRAP period, the handled sample is released towards detection. A comparison with an analogous flow system exploiting zone stopping revealed the superior characteristics of sampling rate and system operation; moreover, the sample inserted volume plays little influence on sampling rate. The system was applied to the spectrophotometric determination of cobalt in pastures, and enhanced figures of merit (sampling rate=18 h(-1); peak height r.s.d.=0.7%, detection limit=0.046 µg L(-1) Co; reagent consumption=330 µg of Tiron per measurement; 98%

Zone trapping/merging zones in flow analysis: a novel approach for rapid assays involving relatively slow chemical reactions.

A novel strategy for accomplishing zone trapping in flow analysis is proposed. The sample and the reagent solutions are simultaneously inserted into convergent carrier streams and the established zones merge together before reaching the detector, where the most concentrated portion of the entire sample zone is trapped. The main characteristics, potentialities and limitations of the strategy were critically evaluated in relation to an analogous flow system with zone stopping. When applied to the spectrophotometric determination of nitrite in river waters, the main figures of merit were maintained, exception made for the sampling frequency which was calculated as 189 h(-1), about 32% higher relatively to the analogous system with zone stopping. The sample inserted volume can be increased up to 1.0 mL without affecting sampling frequency and no problems with pump heating or malfunctions were noted after 8-h operation of the system. In contrast to zone stopping, only a small portion of the sample zone is halted with zone trapping, leading to these beneficial effects.

Evidences of turbulent mixing in multi-pumping flow systems.

Multi-pumping flow systems exploit pulsed flows delivered by solenoid pumps. Their improved performance rely on the enhanced radial mass transport inherent to the pulsed flow, which is a consequence of the establishment of vortices thus a tendency towards turbulent mixing. This paper presents several evidences of turbulent mixing in relation to pulsed flows, such as recorded peak shape, establishment of fluidized beds, exploitation of flow reversal, implementation of relatively slow chemical reactions and/or heating of the reaction medium. In addition, Reynolds number associated with the GO period of a pulsed flow is estimated and photographic images of dispersing samples flowing under laminar regime and pulsed flow conditions are presented.