Reversed-phase porous silica rods, an alternative approach to high-performance liquid chromatographic separation using the sequential injection chromatography technique

A commercially available porous silica rod column was used as a separation tool for the sequential injection analysis (SIA). A porous solid monolithic column showed high performance at a low pressure, allowing sequential injection analysis to be used for the first time for separation in HPLC fashion. In this contribution, we tried to demonstrate a new separation concept with SIA manifold for the simultaneous determination of four different compounds (methylparaben (MP), propylparaben (PP), triamcinolone acetonide (TCA) and internal standard ketoprofen (KP)) in a pharmaceutical triamcinolon cream 0.1% formulation. A Chromolith Flash RP-18e, 25 mm x 4.6 mm column with a 10 mm pre-column (Merck, Germany) and a FIAlab 3000 system (USA) with an 8-port selection valve and 10 ml syringe were used for sequential injection chromatographic separations in our study. The mobile phase used was acetonitrile-methanol-water (35:5:65, v/v/v) + 0.05% nonylamine, pH 2.5, flow rate 0.6 ml min(-1). The analysis time was <6 min. A novel sequential injection chromatography (SIC) technique with UV spectrophotometric detection was optimised and validated.     

Simple determination of betamethasone and chloramphenicol in a pharmaceutical preparation using a short monolithic column coupled to a sequential injection system

This contribution describes the use of a new separation method based on a reversed-phase sequential injection chromatography (SIC) technique for simultaneous determination of chloramphenicol and betamethasone in pharmaceutical eye drops. A short monolithic column coupled with a sequential injection analysis (SIA) system enabled separation of two compounds in one step. A Chromolith Flash RP-18e, 25 x 4.6 mm column with a 5 mm precolumn (Merck, Germany) and a FIA1ab 3000 system (USA) with a 6-port selection valve and 5 mL syringe were used for sequential injection chromatographic separations in this study. The mobile phase used was acetonitrile-water (30:80, v/v), flow rate 0.48 mL/min; UV detection was at two wavelengths, i.e., 241 and 278 nm (absorption maxima of betamethasone and chloramphenicol, respectively). The basic validation parameters showed good results: linearity of determination for both compounds including internal standard (propylparaben) >0.999; repeatability of determination (RSD) in the range 0.8-1.7% at two different concentration levels, and detection limits in the range 0.5-1.0 mg/mL. The chromatographic resolution between compound peaks was greater than 2.1 and the analysis time was less than 8 min under optimal conditions. The developed sequential injection chromatography method was compared with the HPLC method and was found to be applicable for routine analysis of active compounds in pharmaceutical preparations.     

Flow Injection/Sequential Injection Chromatography: A Review of Recent Developments in Low Pressure with High Performance Chemical Separation

This review focuses on the developments of the various parts of instrumentation and the operation of low to medium pressure flow injection and sequential injection chromatography (FIC and SIC) systems. The report and discussion include solution delivery system, separation column, flow cell and detector, mobile phase management, and online sample pretreatment. Applications of FIC and SIC and their differences as compared to HPLC are also presented.     

Sequential injection chromatographic determination of ambroxol hydrochloride and doxycycline in pharmaceutical preparations

A new separation method based on a novel reversed-phase sequential injection chromatography (SIC) technique was used for simultaneous determination of ambroxol hydrochloride and doxycycline in pharmaceutical preparations in this contribution.The coupling of short monolith with SIA system results in an implementation of separation step to until no-separation low-pressure method.A Chromolith^® Flash RP-18e, 25-4.6 mm column (Merck, Germany) and a FIAlab^® 3000 system (USA) with a six-port selection valve and 5 ml syringe were used for sequential injection chromatographic separations in our study. The mobile phase used was acetonitrile-water (20:90, v/v), pH 2.5 adjusted with 98% phosphoric acid, flow rate 0.48 ml min⁻¹, UV detection was at 213 nm.The validation parameters have shown good results: linearity of determination for both compounds including internal standard (ethylparaben) >0.999; repeatability of determination (R.S.D.) in the range 0.5-5.4% at three different concentration levels, detection limits in the range 0.5-2.0 μg ml⁻¹, and recovery from the pharmaceutical preparation in the range 99.3-99.9%. The chromatographic resolution between peak compounds was >5.0 and analysis time was <9 min under the optimal conditions. The method was found to be applicable for routine analysis of the active compounds ambroxol hydrochloride and doxycycline in various pharmaceutical preparations.     

Sequential injection analysis implementing multiple standard additions for As speciation by liquid chromatography and atomic fluorescence spectrometry (SIA-HPLC-AFS)

An analytical procedure for multiple standard additions of arsenic species using sequential injection analysis (SIA) is proposed for their quantification in seafood extracts. SIA presented flexibility for generating multiple specie standards at the ng mL⁻¹ concentration level by adding different volumes of As(III), As(V), monomethylarsonic (MMA) and dimethylarsinic (DMA) to the sample. The mixed sample plus standard solutions were delivered from SIA to fill the HPLC injection loop. Subsequently, As species were separated by HPLC and analyzed by atomic fluorescence spectrometry (AFS). The proposed system comprised two independently controlled modules, with the HPLC loop acting as the intermediary device. The analytical frequency was enhanced by combining the actions of both modules. While the added sample was flowing through the chromatographic column towards the detection system, the SIA program started performing the standard additions to another sample. The proposed method was applied to spoiled seafood extracts. Detection limits based on 3σ for As(III), As(V), MMA and DMA were 0.023, 0.39, 0.45 and 1.0 ng mL⁻¹, respectively.     

Fast simultaneous spectrophotometric determination of naphazoline nitrate and methylparaben by sequential injection chromatography

Fast simultaneous determination of naphazoline nitrate and methylparaben in pharmaceuticals using separation method based on a novel reversed-phase sequential injection chromatography (SIC) is described in this contribution as an alternative to classical HPLC. A Chromolith™ Flash RP-18e (25 mm × 4.6 mm) column (Merck^®, Germany) and a FIAlab^® 3000 system (USA) with a six-port selection valve and 5.0 ml syringe pump were used for sequential injection chromatographic separations in our study. The mobile phase used was methanol/water (40:65, v/v), pH 5.2 adjusted with triethylamine 0.8 μl ml⁻¹ and acetic acid, at flow rate 0.9 ml min⁻¹. UV detection provided by DAD detector and two wavelengths were simultaneously monitored for increasing sensitivity of determination. Detector was set up at 220 nm for naphazoline nitrate and 256 nm for methylparaben and ethylparaben (IS). There is no necessity to use pre-adjustment of sample of nasal drops (only dilution with mobile phase) so the time of the whole analysis is very short. The validation parameters have shown good results: linearity of determination for both components (naphazoline nitrate and methylparaben), correlation coefficient >0.999; repeatability of determination (R.S.D.) in the range 0.5-1.6% at three different concentration levels, detection limits 0.02 μg ml⁻¹ (naphazoline nitrate) and 0.20 μg ml⁻¹ (methylparaben and ethylparaben), and recovery from the pharmaceutical preparations in the range 100.06-102.55%. The chromatographic resolution between peaks of compounds was more than 4.0 and analysis time was less than 4 min under the optimal conditions. The advantages and drawbacks of SIC against classical HPLC are discussed showing that SIC can be an advantageous alternative in many cases.     

Microfluidic sequential injection analysis system based on polydimethylsiloxane (PDMS) chip with integrated pneumatic-actuated valves

A sequential injection analysis (SIA) system based on polydimethylsiloxane (PDMS) chip with integrated pneumatic-actuated valves was developed. A novel SIA operation mode using multiphase laminar flow effect and pneumatic microvalve control was proposed. The sample and reagent solutions were synchronously loaded and injected in the chip-based sample injection module instead of multi-step sequential injection by a multiposition valve and a reciprocating pump as in conventional SIA system. The sample and reagent injection volumes were reduced to ca. 1.1 nL. The present system has the advantages of simple structure, fast and convenient operation, low sample and reagent consumption, and high degree of integration and automation. The system operation conditions were optimized using fluorescein as model sample. Its feasibility in biological analysis was preliminarily demonstrated in enzyme inhibition assay.     

Sequential injection chromatography against HPLC and CE: Application to separation and quantification of amoxicillin and clavulanic acid

Sequential injection chromatography (SIC) has been recently proposed as an alternative separation technique. SIC has the advantages of inexpensiveness, rapid operation procedure, small dimension, short stabilization time, friendly maintenance process and less reagent consumption. In contrast, SIC has had suffered from some limitations. In the current study, a higher pressure resistant selection valve with additional ports than that available in commercially SIC systems was installed. This development allows achieving solution propulsion without showing leakage and linking more standard/sample solutions. In addition, a new method for the separation and quantification of amoxicillin and clavulanic acid in pharmaceutical formulations has been optimized and validated. A comparative study on the efficiency of the SIC method with previous HPLC and CE methods was conducted as well. Besides the benefits of the instrumentation of SIC, the proposed method has shown some advantages over previous HPLC and CE methods with respect to reagent consumption and sample frequency. Other such analytical characters of the SIC method as resolution, peak symmetry, number of theoretical plates, linearity range, accuracy, precision and limits of detection and quantification, recorded comparable results with those obtained from previous HPLC and CE methods.     

A novel application of Onyx™ monolithic column for simultaneous determination of salicylic acid and triamcinolone acetonide by sequential injection chromatography

A novel and fast simultaneous determination of triamcinolone acetonide (TCA) and salicylic acid (SA) in topical pharmaceutical formulations by sequential injection chromatography (SIC) as an alternative to classical high performance liquid chromatography (HPLC) has been developed. A recently introduced Onyx™ monolithic C18 (50 mm × 4.6 mm, Phenomenex^®) with 5 mm monolithic precolumn were used for the first time for creating sequential injection chromatography system based on a FIAlab^® 3000 with a six-port selection valve and 5.0 mL syringe pump in study. The mobile phase used was acetonitrile/water (35:65, v/v), pH 3.3 adjusted with acetic acid at flow rate 0.9 mL min⁻¹. UV detection provided by fibre-optic DAD detector was set up at 240 nm. Propylparaben was chosen as suitable internal standard (IS). There is only simple pre-adjustment of the sample of topical solution (dilution with mobile phase) so the analysis is not uselessly elongated. Parameters of the method showed good linearity in wide range, correlation coefficient >0.999; system precision (relative standard deviation, R.S.D.) in the range 0.45-1.95% at three different concentration levels, detection limits (3σ) 1.00 μg mL⁻¹ (salicylic acid), 0.66 μg mL⁻¹ (triamcinolone acetonide) and 0.33 μg mL⁻¹ (propylparaben) and recovery from the pharmaceutical preparations in the range 97.50-98.94%. The chromatographic resolution between peaks of compounds was more than 4.5 and analysis time was 5.1 min under the optimal conditions. The advantages of sequential injection chromatography against classical HPLC are discussed and showing that SIC can be a method of option in many cases.     

Potential of multisyringe flow-based multicommutated systems

This work is aimed at emphasizing the potential of the multicommutated systems based on the multisyringe flow injection analysis (MSFIA) modality. First, the characteristics, advantages and withdraws offered by flow analysis systems based on the different non-segmented modalities are briefly described. In these systems, multicommutation and computer control of the analytical process occupy a predominant place, as in the case of sequential injection analysis (SIA), multicommutated flow injection analysis (MCFIA), MSFIA and multipumping flow systems (MPFS). Next, several examples are given and different aspects of the implementation of analysers based on MSFIA designs for the construction of different analysis systems, including intelligent (smart) systems, use of sample pre-treatment automatic systems, for chromatographic and non-chromatographic determinations as well as use of monolithic or capillary electrophoresis columns are considered.     

Interactions between selected bile salts and Triton X-100 or sodium lauryl ether sulfate

Background

Sequential injection chromatography (SIC) is a new alternative separation technology. It has some advantages over high performance liquid chromatography (HPLC) regarding simplicity, inexpensiveness, portability, ease of use, maintenance requirement and operation time. In contrast, SIC has had suffered from some limitations.

Results

The current work involves four achievements. (a) One of the limitations of SIC has been overcome. A higher pressure resistant selection valve with additional ports was installed in an SIC system. This development allows propelling solution without showing solution leakage. (b) A new inexpensive rapid and green method for the separation and quantification of propranolol (PRP) and hydrochlorothiazide (HTZ) in their formulations was optimized and validated. (c) A miniaturized multi-channel fiber optic detector was coupled with the newly developed SIC system to detect PRP and HTZ at 270 and 290 nm, respectively. This issue enhanced the sensitivity rather than using a single-channel detector. (d) A comparative study on the efficiency of the SIC method with that of previous HPLC methods was conducted.

Conclusions

Besides the benefits of the instrumentation of SIC, the proposed method is rapider and more reagent-saving than previous HPLC methods. The total volume of consumed reagents and sample was 4.04 mL. The sample frequency was 22 samples/h. Other such analytical characters of the SIC method as resolution, peak symmetry, numbers of theoretical plates, linearity range, accuracy, precision and limits of detection and quantification recorded comparable results.     

Highly sensitive gas-diffusion sequential injection analysis based on flow manipulation

The present paper describes approaches utilizing the powerful flow manipulation capabilities of sequential injection analysis (SIA) to substantially improve the efficiency of gas-diffusion separation compared to its traditional implementation in flow injection analysis (FIA). Ammonia, ethylamine, diethylamine and triethylamine were used as model analytes in this study. Eleven flow manipulation approaches involving continuous flow, stop-flow, oscillating flow, and the introduction of air bubbles to separate the sample zone from the donor solution were tested. Improvement in sensitivity compared to traditional gas-diffusion FIA exceeding one order of magnitude was achieved. It was observed that this improvement increased with the molecular size of the analyte.     

Sequential injection chromatographic determination of paracetamol, caffeine, and acetylsalicylic acid in pharmaceutical tablets

In this contribution, a new separation method for simultaneous determination of paracetamol, caffeine, acetylsalicylic acid, and internal standard benzoic acid was developed based on a novel reversed-phase sequential injection chromatography (SIC) technique with UV detection. A Chromolith Flash RP-18e, 25-4.6mm column (Merck, Germany) and a FIAlab 3000 system (USA) with an 8-port selection valve and a 5 mL syringe were used for sequential injection chromatographic separations in our study. The mobile phase used was acetonitrile-(0.01 M) phosphate buffer (10:90, v/v) pH 4.05, flow rate 0.6 mL min(-1). UV detection was at 210 and 230 nm. The validation parameters showed good results: linearity (r >0.999) for all compounds, detection limits in the range 0.3-0.8 microg mL(-1), repeatability (RSD) of peak heights between runs in the range 1.10-4.30% at three concentration levels and intra-day repeatability of the retention times in the range 0.28-0.43%. The analysis time was <6 min. The method was found to be applicable for the routine analysis of the active compounds paracetamol, caffeine, and acetylsalicylic acid in pharmaceutical tablets.     

Multisyringe Chromatography (MSC): An Effective and Low Cost Tool for Water-Soluble Vitamin Separation

The advent of monolithic columns has facilitated the combination of chromatographic techniques with nonseparation flow techniques by virtue of the low overpressure introduced by them. Thus far, these combinations have provided excellent results derived from the high selectivity of liquid chromatography and ease of sample handling in nonseparation flow techniques. The joint use of multisyringe flow injection analysis and monolithic columns has provided multisyringe chromatography (MSC).

Simultaneous spectrophotometric determination of ascorbic acid, nicotinic acid, nicotinamide, pyridoxine hydrochloride, thiamine hydrochloride, and riboflavin in commercial drinks using a MSC method, is described in this contribution as an inexpensive alternative to classical HPLC. The MSC method provides baseline separation of all these compounds with resolutions values greater than 2, except for B1 and B6 with resolution of 1,2.

A ChromolithTM Flash RP-18e (50 mm × 4.6 mm) column, a multisyringe burette provided with two 10 mL high-precision bidirectional syringes and an 8-port multiposition valve were used for a dual isocratic chromatographic separation in our study.     

Flow techniques in water analysis

In the present work the main flow techniques for the analysis and monitoring of several parameters of interest in the quality control of different types of waters are reviewed. Firstly, a review involving the advantages and disadvantages of flow techniques, from those currently out-dated, such as segmented flow analysis (SFA), to the most modern techniques, such as flow injection analysis (FIA), sequential injection analysis (SIA) and multi-commutation techniques (MCFA), is carried out. On the other hand, a new technique, the multi-syringe flow analysis (MSFA) is hereby described for the first time as both a fast and robust alternative. Its possibilities, limitations and potential advantages when using this technique either on its own or coupled to SIA, which carries out a previous sample handling, are outlined.     

Longevity and other practical benefits of monolithic silica columns in the analysis of samples with complex matrices

At the turn of the millennium, the monolithic columns invoked new chances in HPLC. Even more than their organic polymer-based siblings, the inorganic silica-based monoliths targeted the territory of classical fully porous particle-packed columns, promising many benefits. Based on the number of published articles, the monoliths attracted academics just in the first few years after their introduction to the market. Lately, as superficially porous particles and sub-2-micron fully porous particles dominated the market, they stayed in the focus of routine laboratories and those who really appreciated the high porosity of the monolithic bed. The monoliths' practical benefits cannot be easily traced in the literature when they gradually lose academics' interest. Nevertheless, after more than 20 years of our experience, we still favor silica monoliths for their low back pressure and longevity when analyzing samples of clinical, pharmaceutical, and environmental origin. At the same time, the high permeability of monoliths enabled the birth of sequential injection chromatography, the medium-pressure separation technique based on the flexible flow manifold. This minireview aims to check, discuss, and summarize the practical aspects of monolithic silica columns in HPLC and medium-pressure sequential injection chromatography (SIC) that may not be visible at first sight but are evident retrospectively.     

Sequential Injection Analysis Hyphenated with Other Flow Techniques: A Review

In the last two decades, sequential injection analysis (SIA) became an important automation tool in several analytical fields as it provided advantages in terms of versatility, robustness, and consumption of samples and reagents.

The noteworthy versatility of an SIA selection valve allowed its association with several units, including devices typically associated with other flow techniques, resulting in generally better analytical performance.

This review discusses the hyphenization of SIA with other flow approaches outlining its advantages and motivations, which are related mainly with sample manipulation and solutions management. The future trends and perspectives in this field are also addressed.     

Development of a sequential injection chromatography (SIC) method for determination of simazine,atrazine, and propazine

This paper describes the development of a sequential injection chromatography (SIC) procedure for separation and quantification of the herbicides simazine, atrazine, and propazine exploring the low backpressure of a 2.5 cm long monolithic C₁₈ column. The separation of the three compounds was achieved in less than 90 s with resolution >1.5 using a mobile phase composed by ACN/1.25 mmol/L acetate buffer (pH 4.5) at the volumetric ratio of 35:65 and flow rate of 40 μL/s. Detection was made at 223 nm using a flow cell with 40 mm of optical path length. The LOD was 10 μg/L for the three triazines and the quantification limits were of 30 μg/L for simazine and propazine and 40 μg/L for atrazine. The sampling frequency is 27 samples per hour, consuming 1.1 mL of ACN per analysis. The proposed methodology was applied to spiked water samples and no statistically significant differences were observed in comparison to a conventional HPLC–UV method. The major metabolites of atrazine and other herbicides did not interfere in the analysis, being eluted from the column either together with the unretained peak, or at retention times well‐resolved from the studied compounds.     

Rapid narrow band elution for on-line SPE using a novel solvent plug injection technique

Determination of trace constituents in biological and environmental samples usually requires a pre-concentration step. While solid-phase extraction (SPE) has been widely used, it is slow, labor intensive and adversely affected by analytical errors from handling. On-line SPE eliminates some of the flaws but often suffers from solvent compatibility problems with the subsequent chromatography separation. In this study, we are presenting a technical solution for overcoming some of these compatibility issues, by utilizing a fully automated, focused SPE sample transfer technique utilizing narrow-band solvent plugs, for seamless hyphenation with high-performance liquid chromatography (HPLC) or flow injection mass spectrometry (MS). A wide range of pharmaceutical compounds was studied in different sample matrices. Short plugs of high elution strength solvent were generated by means of an electrically actuated sample loop and enrichment and transfer steps monitored using on-line SPE-MS. The impact of the solvent plugs on chromatographic separation was studied using hyphenated SPE-LC-MS. By carefully examining elution profiles of solvent plugs of different compositions, optimum conditions for quantitative elution within well-defined volumes were found for all substances. In addition, the highly focused elution bands resulted in excellent retention time and peak area reproducibilities when injected on-line onto HPLC columns. Finally, to demonstrate proof-of-principle, the fully integrated on-line SPE-LC-MS system was applied to the analysis of spiked urine and river water samples.