A micro-sequential injection spectrophotometric procedure for DNA assay was developed based on the employment of a lab-on-valve (LOV) meso-fluidic analytical system. A small amount of crystal violet solution (10μl) was de-colored inside the flow cell of the LOV at the presence of 5μl λ-DNA/HindIII within a certain pH range, and the absorbance decrease of crystal violet solution at 591 nm was measured via optical fibers and was employed as the basis of quantification. A uni-variant approach was adopted for the optimization of experimental parameters, including buffer pH, concentration and volume of crystal violet solution, reaction time and sample/reagent loading flow rates. A linear calibration graph was obtained within 0.2-6.0μgml−1, along with a detection limit of 0.07μgml−1. The procedure was applied for the determination of λ-DNA/HindIII in synthetic samples in comparison with a documented procedure.

Termed the third generation of flow-injection analysis, sequential injection (SI)-lab-on-valve (LOV) has specific advantages and allows novel, unique applications-not least as a versatile front end to a variety of detection techniques. This review presents and discusses progress to date of the SI-LOV approach as well as its applications in the automation and the micro-miniaturization of on-line sample pre-treatment. Special emphasis is placed on using SI-LOV in conjunction with bead injection (BI) for on-line separation and pre-concentration of ultra-trace levels of metals by exploiting the renewable micro-column approach. With detection by ETAAS and ICP-MS, it is shown, as illustrated by recent results in the authors' laboratory, that this methodology eliminates the problems encountered in conventional on-line column pre-concentration systems, improves the overall operational efficiency and yields the robustness necessary for routine assays. Also discussed is the future potential of the SI-LOV approach as a front end to various analytical protocols.

A sequential injection (SI) on-line matrix removal and trace metal preconcentration procedure by using a novel microcolumn packed with PTFE beads is described, and demonstrated for trace cadmium analysis with detection by electrothermal atomic absorption spectrometry (ETAAS). The analyte is initially complexed with diethyldithiophosphate (DDPA) and adsorbed onto the column, which is afterwards eluted with 50ml of ethanol and subsequently introduced via air segmentation into the graphite tube for quantification. The ETAAS determination is synchronized with sample pre-treatment in the SI system. No flow resistance is encountered at a flow rate of 9.0ml min21 through the column. The preconcentration efficiency was improved significantly by using the packed column as compared with a knotted reactor. With a 40 s sample loading time at 4.5ml min21 along with a sampling frequency of 16 h21, quantitative adsorption of cadmium (99% retention efficiency) and an enrichment factor of 59.4 were obtained, as compared with only 46.7% and 28.0 by using a knotted reactor of similar internal surface area as the packed column. The detection limits and precision (RSD, 0.1mg l21 Cd) are at the same levels, i.e., 1.3ng l21 (LOD), 1.3% (RSD) for the packed column, and 1.2ng l21 (LOD), 1.5% (RSD) for the knotted reactor. The practical applicability of the procedure is demonstrated by the determination of trace levels of cadmium in three certified reference materials.

An automatic protocol for in-situ assay of dsDNA is presented by employing a micro-sequential injection lab-on-valve meso-fluidic system, which facilitates precise fluidic handling at the 0.1-10ml level. Sub-nano-liter to a few micro-liters of DNA sample and ethidium bromide (EB) solutions were introduced into the meso-fluidic system, where EB binding onto DNA takes place and an intercalated DNA-EB adduct was formed, which was afterwards excited in the flow cell of the LOV by a 473nm laser beam, and the emitted fluorescence was monitored in-situ via optical fibers. The experimental variables, i.e., pH of the buffer solution, the concentration and volume of EB solution, the reaction time and the fluid flow rates, were investigated. By loading 600nl sample and 1.0ml EB solution, a linear calibration graph was obtained within 0.03-3.0mg ml21 (dsDNA), and a detection limit (3s) of 0.009mg ml21 was achieved, along with a sampling frequency of 60h21 and a precision of 1.9% at the 1.0mg ml21 level. The detection limit was further improved to 0.006mg ml21 by increasing the sample volume to 2.0ml. Plasmid DNA in E. Coli extraction and l-DNA/Hind III in four synthetic samples were assayed by using this procedure. For the plasmid DNA, a good agreement with the documented UV method was obtained, while spiking recoveries for the synthetic samples were 95.6-103.4%.

An automated sequential injection (SI) on-line solvent extraction-back extraction eparation/preconcentration procedure is described. Demonstrated for the assay of cadmium by electrothermal atomic absorption spectrometry (ETAAS), the analyte is initially complexed with ammonium pyrrolidinedithiocarbamate (APDC) in citrate buffer and the chelate is extracted into isobutyl methyl ketone (IBMK), which is separated from the aqueous phase by means of a newly designed dual-conical gravitational phase separator. A metered amount of the organic eluate is aspirated and stored in the PTFE holding coil (HC) of the SI-system. Afterwards, it is dispensed and mixed with an aqueous back extractant of dilute nitric acid containing Hg(II) ions as stripping agent, thereby facilitating a rapid metal-exchange reaction with the APDC ligand and transfer of the Cd into the aqueous phase. The aqueous phase is separated in a second dual-conical gravitational phase separator, and 30 l of it is entrapped and metered in a sample loop (SL) and subsequently introduced via air segmentation into the graphite tube for analyte quantification. The ETAAS determination is performed in parallel with the separation/preconcentration process of the ensuing sample. An enrichment factor of 21.4, a detection limit of 2.7ng l−1, along with a sampling frequency of 13h−1 were obtained at a sample flow rate of 6.0ml min−1. The precision (R. S. D.) at the 0.4g l−1 level was 1.8% as compared to 3.2% when quantifying the organic extractant directly. The applicability of the procedure is demonstrated for the determination of trace levels of cadmium in three certified reference materials.