Sequential Injection Analysis System Research Articles

Automatic flow methodology for kinetic and inhibition studies of reactions with poorly water-soluble substrates in ionic liquid systems

In the present work an automatic generic tool, based on sequential injection analysis (SIA) for kinetic and inhibition studies of reactions with poorly water-soluble compounds in ionic liquid (IL)-containing systems, is described. The oxidation of the poorly water-soluble phenolic compound, caffeic acid, catalyzed by the mushroom tyrosinase, in different 1-butyl-3-methylimidazolium tetrafluoroborate ([bmim][BF 4])/buffer mixtures as reaction media, was investigated. This determination was based on measuring depletion rate of the substrate caffeic acid at its maximum wavelength ( λ max 311 nm). The influence of several parameters such as substrate and enzyme concentration, temperature, pH, delay times and measurement periods on the sensitivity and performance of the SIA system were studied and the optimum reaction conditions subsequently selected. The obtained results showed that tyrosinase was active in oxidising caffeic acid in this water-miscible IL and the presence of an impaired tyrosinase activity with increase in [bmim][BF 4] concentration as an increase in the apparent Michaelis–Menten constant ( K M app ) was observed while the maximum reaction rate ( V max app ) remained fairly constant. The results were compared to those obtained when the assay was performed in water/methanol mixtures under the same conditions to substantiate [bmim][BF 4] as an alternative to conventional organic solvents. Additionally, it was shown that tyrosinase is effectively inhibited by the substrate analogues tested ( trans-cinnamic acid and 3,4-dihydroxybenzoic acid) in the IL-containing aqueous system used.

Enzyme based assays in a sequential injection format: A review

Sequential injection analysis systems have been extensively exploited in the last decades for the implementation of enzyme based assays aiming the evaluation of enzyme activity or the determination of specific analytes. The most prominent aspects of the automation of enzymatic assays in these systems are discussed in this review. Special attention is devoted to the mode of enzyme manipulation in homogeneous or heterogeneous media and to the comparison with batch and flow injection enzyme methodologies. The possibility of implementing strategies for the enhancement of selectivity in specific determinations is also addressed. The more recent trends in this field are discussed focusing mainly on the miniaturization resorting to the lab-on valve platform as well as on the bead injection concept.

Sequential injection analysis system with spectrophotometric detection for determination of norfloxacin and ciprofloxacin in pharmaceutical formulations

This work proposes a sequential injection analysis (SIA) system for the spectrophotometric determination of norfloxacin (NOR) and ciprofloxacin (CIP) in pharmaceutical formulations. The methodology was based on the reaction of these drugs with p-(dimethylamino)cinnamaldehyde in micellar medium, producing orange colored products (λmax = 495 nm). Beer´s law was obeyed in the concentration range from 2.75x10-5 to 3.44x10-4 mol L-1 and 3.26x10-5 to 3.54x10-4 mol L-1 for NOR and CIP, respectively and sampling rate was 25 h-1. Commercial samples were analyzed and results obtained through the proposed method were in good agreement with those obtained using the reference procedure for a 95% confidence level.

A simple automated method for the determination of nitrate and nitrite in infant formula and milk powder using sequential injection analysis.

A fast and efficient automated method using a sequential injection analysis (SIA) system, based on the Griess, reaction was developed for the determination of nitrate and nitrite in infant formulas and milk powder. The system enables to mix a measured amount of sample (previously constituted in the liquid form and deproteinized) with the chromogenic reagent to produce a colored substance whose absorbance was recorded. For nitrate determination, an on-line prereduction step was added by passing the sample through a Cd minicolumn. The system was controlled from a PC by means of a user-friendly program. Figures of merit include linearity (r2 > 0.999 for both analytes), limits of detection (0.32 mg kg−1 NO3-N, and 0.05 mg kg−1 NO2-N), and precision (sr%) 0.8–3.0. Results were statistically in good agreement with those obtained with the reference ISO-IDF method. The sampling frequency was 30 hour−1 (nitrate) and 80 hour−1 (nitrite) when performed separately.

Development of an online citrate/Ca2+sensing system for dialysis

An online citrate and Ca(2+) sensing system based on sequential injection analysis (SIA) is developed as a safety module for hemodialysis. Host 1 displays high affinity towards citrate, and was selected for this study owing to its unique structural features. The o-aminomethylphenylboronic moiety can effectively interact with the α-hydroxycarboxylate moiety of citrate and the remaining two guanidiniums may further stabilize the complex via hydrogen bonds. Fura-2 chelates to Ca(2+) with a high selectivity and affinity and was utilized in this study for Ca(2+) measurements. The citrate sensing chemistry via an indicator displacement assay is orthogonal to the Ca(2+) sensing chemistry, and the use of sophisticated chemometrics is not required for data analysis. The citrate and Ca(2+) concentrations in dialysate samples are measured with the developed SIA system. The obtained citrate concentrations were verified via a commercially available enzymatic assay and an NMR method, respectively, while the Ca(2+) concentrations were verified via atomic absorption.

Automated evaluation of the effect of ionic liquids on catalase activity

An automated assay for the evaluation of the influence of ionic liquids on the activity of catalase was developed. The activity and inhibition assays were implemented in a sequential injection analysis (SIA) system and intended to contribute for the estimation of the toxicity of the tested compounds. The fast developed methodology was based on the oxidation of the non-fluorescent probe amplex red, in the presence of H2O2, to produce resorufin, a strong fluorescent compound. Catalase activity was monitored by the decreased of the fluorescence intensity due to the consumption of H2O2 by the enzyme. The activity assays were performed in strictly aqueous media and in the presence of increasing concentrations of seven commercially available ionic liquids and sodium azide, a strong inhibitor of catalase. IC50 values between 0.15 and 2.77M were obtained for the tested compounds, revealing distinct inhibitory effects. This allowed us to perform some considerations about the toxicity of the tested cations and anions. The developed SIA methodology showed to be robust and exhibited good repeatability in all the assay conditions. On the other hand, it proved to be in good agreement with the actual concerns of “Green Chemistry” since it involved the consumption of less than 200 μL of reagents and the production of only 1.7mL of effluent (per cycle) and at the same time reduced the operator exposure resulting in increased environmental and human safety.

Simultaneous Determination of L- and D-T4 Using a Sequential Injection Analysis/Sensors System

A sequential injection analysis system was selected for the simultaneous determination of L- and D-T4 in raw material and pharmaceutical products, because of the highest precision and accuracy, the lower consumption of sample and buffer, and the flexibility of selecting different buffers accordingly with the type of sensors utilized as detectors. An amperometric biosensor based on D-amino acid oxidase was used as detector for D-T4 and an immunosensor based on anti-L-T4 was used as detector for L-T4. The SIA/biosensors system can be used reliably on-line in synthesis process control, for the simultaneous assay of L- and D- T4 with a frequency of more than 30 samples per hour.

SIA system employing the transient response from a potentiometric sensor array—Correction of a saline matrix effect

A Sequential Injection Analysis (SIA) system and an 8-potentiometric all-solid-state sensor array were coupled in a simple and automated electronic tongue device. The potentiometric sensors used were planar microfabricated structures with standard PVC membranes deposited onto a gold contact. The SIA system permitted the automated operation and generation of the calibration data, needed to build an Artificial Neural Network model, thanks to the precise dosing and mixing of volumes of stock solutions. The resolution of a four-ion mixture, i.e. ammonium, sodium, nitrate and chloride was the study case used for characterization of the system. Two different variants for signal acquisition, steady-state and transient recording, were arranged and compared. The dynamic treatment is shown to offer improved performance thanks to the benefits of the kinetic resolution. For this, it first extracts meaningful data from a FFT transform of each sensor's transient, which is then fed to an ANN model for estimation of each concentration in the four-ion mixture. While in a standard laboratory situation there was no difference between the two approaches, the dynamic treatment allowed the correction of a matrix effect in the case study, where an uncontrolled saline effect could be counterbalanced.

Multicommutation in Flow Systems: A Useful Tool for Pharmaceutical and Clinical Analysis

Multicommutation is a methodology of increasing use for the development of chemical analysis systems. Spe- cifically, it refers to continuous-flow configurations designed with computer-controlled commutators. The flow system, based on the implementation of solenoid valves in Flow Injection Analysis (FIA), is called Multicommutated Flow Sys- tem (MCFIA). These flow systems can be easily re-configured by just changing the procedure with the software, resulting in increased versatility, potential for automation and minimization of both reagent consumption and waste generation. The use of solenoid pumps for propelling the solutions has been recently introduced and is known as Multipumping Flow Sys- tems (MPFS). Finally, Multisyringe Flow Injection Analysis (MSFIA) has been developed implementing multicommuta- tion principles in a modified Sequential Injection Analysis (SIA) system. This review covers a brief description about the fundamentals of the above mentioned methodologies as well as their ap- plication to pharmaceutical and clinical analysis. Several examples are given and some particular aspects are highlighted, such as the differences between measuring the analytical signal in solution or on a solid support (flow-through optosen- sor), a detailed classification of the reported methods in terms of the flow technique employed, or the novelty of each pub- lished method.

Sequential injection trace determination of iron in natural waters using a long‐pathlength liquid core waveguide and different spectrophotometric chemistries

We developed a sequential injection analysis (SIA) method for the determination of iron in coastal, ground, and surface fresh waters using two different reagents (ferrozine and 1,10‐phenathroline). The methodology uses a double‐line SIA system to improve mixing conditions between sample and reagent solutions. A liquid wave‐guide capillary cell (LWCC) with 1.0 m pathlength, 550 µm i.d., and 250 µL internal volume was used to enhance the sensitivity of the determination. The detection limits for the ferrozine and 1,10‐phenanthroline reagent were 0.15 and 0.35 µg L−1, respectively. The system provides a linear response up to 20 µg L−1 and has a high throughput rate (41 h−1), low reagent consumption, and also low effluent production for both reagents. The developed method was applied to natural waters (river, well, ground, potable, and sea waters) and one reference water sample.

11-Molybdobismuthophosphate—A new reagent for the determination of ascorbic acid in batch and sequential injection systems

The guanidinium salt of the new heteropolymolybdate 11-molybdobismuthophosphate Gua(6)PBiMo(11)O(40) (11-MBP) was synthesized, characterized and used as a reagent for batch spectrophotometric (SP) and sequential injection determination of ascorbic acid (AsA). When compared to other Keggin's heteropolyanions, the reduction of 11-MBP with AsA is both fast and maximal within a pH range of 1.6-2.0. The stoichiometry of the reaction was determined using molar ratio and continuous variation methods and was shown to be 1:1. The molar absorptivity of the reduced form of 11-MBP was 6.0 x 10(3)L mol(-1)cm(-1) at 720 nm. The reaction is also specific for AsA. Only cysteine, hydroquinone and hydroxyacids were found to interfere with the reaction, while no interference was observed with the common reducing agents, including reducing sugars, catecholamines, nitrite, sulfite and iron(II) ions. Batch SP and sequential injection analysis (SIA) systems were developed for the determination of AsA, with calibration ranges of the SP methods at 2x10(-6)-8 x 10(-5)M for a 10mm cell and 5 x 10(-7)-3 x 10(-5)M for a 50mm cell and a limit of detection at 3 x 10(-7)M. The linear range of the SIA method was 6 x 10(-6)-5 x 10(-4)M, with a detection limit of 2 x 10(-6)M and a sample throughput of 15 h(-1). The proposed methods were successfully used for the determination of AsA in both pharmaceuticals and fruit juices, and the results were consistent with those provided by the 2,6-dichlorophenolindophenol method.

Two analyte calibrations from the transient response of a single potentiometric sensor employed with the SIA technique

Simultaneous quantification of Cd 2+ and Pb 2+ in solution has been correctly targeted using the kinetic information from a single non-specific potentiometric sensor. Dual quantification was accomplished from the complex information in the transient response of an electrode used in a Sequential Injection Analysis (SIA) system and recorded after step injection of sample. Data was firstly preprocessed with the Discrete Wavelet Transform (DWT) to extract significant features and then fed into an Artificial Neural Network (ANN) for building the calibration model. DWT stage was optimized regarding the wavelet function and decomposition level, while the ANN stage was optimized on its structure. To simultaneously corroborate the effectiveness of the approach, two different potentiometric sensors were used as study case, one using a glass selective to Cd 2+ and another a PVC membrane selective to Pb 2+.

Sequential injection technique for determination of phenoxybenzamine hydrochloride and metoclopramide in pharmaceutical formulations

A sequential injection analysis (SIA) system is described for the determination of phenoxybenzamine hydrochloride and metoclopramide using spectrophotometer as detector. The method is based on the detection of an unstable red intermediate compound resulting from the reaction of phenoxybenzamine hydrochloride or metoclopramide with the diazotizating product of p-phenylenediamine with sodium nitrite in hydrochloric acid medium. The sampling frequency is 69 h−1 and 75 h−1 for phenoxybenzamine hydrochloride and metoclopramide, respectively. The linear range is 10–400 μg/mL for phenoxybenzamine hydrochloride with a detection limit of 0.081 μg/mL and 20–250 μg/mL for metoclopramide with a detection limit of 0.034 μg/mL. The RSD is 1.01 and 0.45% for phenoxybenzamine hydrochloride and metoclopramide, respectively. The proposed methods were used to determine phenoxybenzamine hydrochloride and metoclopramide in pharmaceuticals. The results are compared with those obtained by pharmacopoeia method.

Sequential injection analysis (SIA) and response surface methodology: A versatile small volume approach for optimization of photo-Fenton processes

Optimization of photo-Fenton degradation of copper phthalocyanine blue was achieved by response surface methodology (RSM) constructed with the aid of a sequential injection analysis (SIA) system coupled to a homemade photo-reactor. Highest degradation percentage was obtained at the following conditions [H 2O 2]/[phthalocyanine] = 7, [H 2O 2]/[FeSO 4] = 10, pH = 2.5, and stopped flow time in the photo reactor = 30 s. The SIA system was designed to prepare a monosegment containing the reagents and sample, to pump it toward the photo-reactor for the specified time and send the products to a flow-through spectrophotometer for monitoring the color reduction of the dye. Changes in parameters such as reagent molar ratios, residence time and pH were made by modifications in the software commanding the SI system, without the need for physical reconfiguration of reagents around the selection valve. The proposed procedure and system fed the statistical program with degradation data for fast construction of response surface plots. After optimization, 97% of the dye was degraded.

Sequential injection system for phospholipase A 2 activity evaluation: Studies on liposomes using an environment-sensitive fluorescent probe

This work reports the development of an automatic methodology based on the use of 1-anilinonaphthalene-8-sulfonate (ANS) as an interfacial fluorescent probe for detecting the hydrophobic environment shift around the probe, caused by the hydrolytic action of PLA 2 on the liposomes. The implementation of this reaction in a sequential injection analysis (SIA) system along with the use of the mixing chambers permitted the evaluation of PLA 2 activity and assessment of the inhibitory effect of the non-steroidal anti-inflammatory drugs (NSAIDs) on PLA 2 activity. Several studies were performed with the aim of establishing the appropriate flow system configuration: the liposome substrate; PLA 2 and ANS optimum concentrations and incubation times before and after the enzyme addition. Based on these studies, the optimum reaction conditions were selected. It was shown that PLA 2 is effectively inhibited by the NSAIDs tested (meloxicam, tolmetin and ibuprofen) and by the α-lipoic acid, used as a positive control. Results obtained from the flow system are in agreement with those provided by the comparison batch procedures. The proposed methodology is in fact more efficient and rapid than the comparison batch experiments, enabling the exact timing of fluidic manipulations and precise control of the reaction conditions.

Pre-evaluation of metal ions as a catalyst on chemiluminometric sequential injection analysis with luminol–H 2O 2 system

Catalytic effect of metal ions on luminol chemiluminescence (CL) was investigated by sequential injection analysis (SIA). The SIA system was set up with two solenoid micropumps, an eight-port selection valve, and a photosensor module with a fountain-type chemiluminescence cell. The SIA system was controlled and the CL signals were collected by a LabVIEW program. Aqueous solutions of luminol, H 2O 2, and a sample solution containing metal ion were sequentially aspirated to the holding coil, and the zones were immediately propelled to the detection cell. After optimizing the parameters using 1 × 10 −5 M Fe 3+ solution, catalytic effect of some metal species was compared. Among 16 metal species examined, relatively strong CL responses were obtained with Fe 3+, Fe 2+, VO 2+, VO 3 −, MnO 4 −, Co 2+, and Cu 2+. The limits of detection by the present SIA system were comparable to FIA systems. Permanganate ion showed the highest CL sensitivity among the metal species examined; the calibration graph for MnO 4 − was linear at the concentration level of 10 −8 M and the limit of detection for MnO 4 − was 4.0 × 10 −10 M (S/N = 3).

Automated and ultrasensitive detection of methyl-3-quinoxaline-2-carboxylic acid by using gold nanoparticles probes SIA-rt-PCR

An ultrasensitive and rapid sequential injection analysis (SIA) based on real-time PCR (SIA-rt-PCR) assay was developed by using different nanoparticles for the detection of small molecule chemicals residues. Gold (Au) nanoparticle, conjugated with goat anti-rabbit IgG and duplex strand DNA (dsDNA), was used as a substitute for chemiluminescent probes in an SIA system. By indirect competitive immunoreactions in the SIA system, the gold nanoparticles were attached to antigens which were immobilized by superparamagnetic nanoparticles (SMNPs). The dsDNA on the gold nanoparticles was dehybridized and then the single-stranded DNA (ssDNA) was collected and quantified with rt-PCR, which showed a rather low linearity range from 2.5 atto mol L −1 (aM) to 250 femto mol L −1 (fM) and the LOD was 1.4 aM. This method, which is rapid, automated and capable of high-throughput, was used to detect methyl-3-quinoxaline-2-carboxylic acid (MQCA) residues in real samples. The analytical results had a coefficient of variation of less than 15% and the recovery was 89–108%.

Simple sequential injection analysis system for rapid determination of microalbuminuria

A simple, specific and sensitive sequential injection analysis (SIA) system based on non-immunoassay fluorescent detection has been developed for the determination of urinary albumin. The specific binding of the dye Albumin Blue 580 (AB 580) to albumin in urine generated high emission fluorescent signals. The excitation and emission wavelengths were set at 590 and 610 nm, respectively. The analytical range was obtained from 1 to 100 mg L −1, with a detection limit of 0.3 mg L −1 (S/N = 3). The SIA system gave high precision with relative standard deviations (R.S.D.s) of 0.9% and 1.4% when evaluated with 15 and 100 mg L −1 albumin ( n = 15), respectively. The method exhibited good reproducibility, as assessed by performing four analytical curves on different days, and intra-run CVs (2.3–3.3%) and inter-run CVs (3.8%) were obtained. Rapid operation was achieved with a sample throughput of 37 h −1. This method was successfully applied to the determination of urinary albumin, and the method was highly correlated with the immunoturbidimetric method ( r 2 = 0.965; n = 72).

Development of a flow through photo-reactor to study degradation of organic compounds by Sequential Injection Analysis (SIA)

This work describes a photo-reactor to perform in line degradation of organic compounds by photo-Fenton reaction using Sequential Injection Analysis (SIA) system. A copper phthalocyanine-3,4',4<FONT FACE=Symbol><FONT FACE=Symbol>²</FONT></FONT>,4<FONT FACE=Symbol><FONT FACE=Symbol>²¢</FONT></FONT>-tetrasulfonic acid tetrasodium salt dye solution was used as a model compound for the phthalocyanine family, whose pigments have a large use in automotive coatings industry. Based on preliminary tests, 97% of color removal was obtained from a solution containing 20 µmol L-1 of this dye.

Immobilization of glucose oxidase and lactate dehydrogenase onto magnetic nanoparticles for bioprocess monitoring system

Glucose oxidase (GOD) and lactate dehydrogenase (LDH) were immobilized onto magnetic nanoparticles, viz. Fe3O4, via carbodiimide and glutaraldehyde. The immobilization efficiency was largely dependent upon the immobilization time and concentration of glutaraldehyde. The magnetic nanoparticles had a mean diameter of 9.3 nm and were superparamagnetic. The immobilization of GOD and LDH on the nanoparticles slightly decreased their saturation magnetization. However, the FT-IR spectra showed that GOD and LDH were immobilized onto the nanoparticles by different binding mechanisms, the reason for which was not well explained. The optimum pH values of the immobilized GOD and LDH were changed to 8 and 10, respectively. The free and immobilized enzyme kinetic parameters (Km and Vmax) were determined by Michaelis-Menten enzyme kinetics. The Km values for free and immobilized GOD were 0.168 and 0.324 mM, respectively, while those for free and immobilized LDH were 0.19 and 0.163 mM for NAD, and 2.976 and 4.785 mM for lactate, respectively. High operational stability was observed, with more than 80% of the initial enzyme activity being retained for the immobilized GOD up to 12 h and for the immobilized LDH up to 24 h. The immobilized GOD was applied to a sequential injection analysis system for the application of bioprocess monitoring.