Lab-in-syringe automated protein precipitation and salting-out homogenous liquid-liquid extraction coupled online to UHPLC-MS/MS for the determination of beta-blockers in serum.

A sample preparation method involving tandem implementation of protein precipitation and salting-out homogenous liquid-liquid extraction was developed for the determination of beta-blockers in serum. The entire procedure was automated using a computer-controlled syringe pump following the Lab-In-Syringe approach. It is based on the denaturation of serum proteins with acetonitrile followed by salt-induced phase separation upon which the proteins accumulate as a compact layer at the interphase of the solutions. The extract is then separated and diluted in-syringe before being submitted to online coupled UHPLC-MS/MS. A 1 mL glass syringe containing a small stir bar for solution mixing at up to 3000 rpm, was used to deal with sample volumes as small as 100 µL. A sample throughput of 7 h-1 was achieved by performing the chromatographic run and sample preparation procedure in parallel. Linear working ranges were obtained for all analytes between 5 and 100 ng mL-1, with LOD values ranging from 0.4 to 1.5 ng mL-1. Accuracy values in the range of 88.2-106% and high precision of <11% RSD suggest applicability for routine analysis that can be further improved using deuterated standards.

Automated centrifugation-less milk deproteinization and homogenous liquid-liquid extraction of sulfonamides for online liquid chromatography.

A novel approach to the determination of sulfonamides in milk based on the Lab-In-Syringe technique is presented. The method involves automated salting-out liquid-liquid extraction of the analytes, allowing simultaneous sample deproteination without requiring centrifugation or manual sample handling. The procedural parameters, including salt type, solvent-to-sample ratio, and salt solution volume, were studied. The extracts obtained were diluted in situ and transferred to online coupled liquid chromatography for analyte separation carried out in parallel to the subsequent extraction. In this way, a sample throughput of approximately 6 h-1 was achieved. The detection limits ranged from 25 to 32 µg L-1 using a 500 µL milk sample and spectrophotometric detection. The applicability of the developed method to sample analysis was proven by recovery values ranging from 73.2% to 94.1% (86.3% on average) for milk samples of different fat content spiked at 3 µg mL-1 level. Straightforward automation of one of the most laborious preparation steps in food analysis, i.e., deproteination, was demonstrated.

Recent trends on the implementation of reticular materials in column-centered separations.

Advances in the development of column-based analytical separations are strongly linked to the development of novel materials. Stationary phases for chromatographic separation are usually based on silica and polymer materials. Nevertheless, recent advances have been made using porous crystalline reticular materials, such as metal-organic frameworks and covalent organic frameworks. However, the direct packing of these materials is often limited due to their small crystal size and nonspherical shape. In this review, recent strategies to incorporate porous crystalline materials as stationary phases for liquid-phase separations are covered. Moreover, we discuss the potential future directions in their development and integration into suitable supports for analytical applications. Finally, we discuss the main challenges to be solved to take full advantage of these materials as stationary phases for analytical separations.

Zeolitic imidazolate frameworks in analytical sample preparation.

Zeolitic imidazolate frameworks are a class of metal-organic frameworks that are topologically isomorphic with zeolites. Zeolitic imidazolate frameworks are composed of tetrahedrally coordinated metal ions connected by imidazolate linkers and have a high porosity and chemical stability. Here, we summarize the progress made in the application of zeolitic imidazolate frameworks in sample preparation for analytical purposes. This review is focused on analytical methods based on liquid chromatography, gas chromatography, or capillary electrophoresis, where the use of zeolitic imidazolate frameworks has contributed to increasing the sensitivity and selectivity of the method. While bulk zeolitic imidazolate frameworks have been directly used in analytical sample preparation protocols, a variety of strategies for their magnetization or their incorporation into sorbent particles, monoliths, fibers, stir bars, or thin films, have been developed. These modifications have facilitated the handling and application of zeolitic imidazolate frameworks for a number of analytical sample treatments including magnetic solid-phase extraction, solid-phase microextraction, stir bar sorptive extraction, or thin film microextraction, among other techniques.

Lab-In-Syringe for automated double-stage sample preparation by coupling salting out liquid-liquid extraction with online solid-phase extraction and liquid chromatographic separation for sulfonamide antibiotics from urine.

A double-stage Lab-In-Syringe automated extraction procedure coupled online to HPLC for the determination of four sulfonamides in urine has been developed. Our method is based on homogeneous liquid-liquid extraction at pH 3 using water-miscible acetonitrile with induction of phase separation by the addition of a saturated solution of kosmotropic salts MgSO4 and NaCl. The procedure allowed extraction of the moderately polar model analytes and the use of a solvent that is compatible with the used separation technique. The automated sample preparation system based on the stirring-assisted Lab-In-Syringe approach was coupled on-line with HPLC-UV for the subsequent separation of the sulfonamide antibiotics. To improve both preconcentration factor and extract cleanup, the analytes were trapped at pH 10 in an anion-exchange resin cartridge integrated into the HPLC injection loop thus achieving a double-stage sample clean-up. Analytes were eluted using an acidic HPLC mobile phase in gradient elution mode. Running the analytes separation and the two-step preparation of the following sample in parallel reduced the total time of analysis to mere 13.5 min. Limits of detection ranged from 5.0 to 7.5 µg/L with linear working ranges of 50-5000 µg/L (r2 > 0.9997) and RSD ≤ 5% (n = 6) at a concentration level of 50 µg/L. Average recovery values were 102.7 ± 7.4% after spiking of urine with sulfonamides at concentrations of 2.5 and 5 mg/L followed by 5 times dilution. To the best of our knowledge, the use of Lab-In-Syringe for the automation of coupled homogeneous liquid-liquid extraction and SPE for preparation of the complex matrices suitable for separation techniques is here presented for the first time.

A flow-based platform hyphenated to on-line liquid chromatography for automatic leaching tests of chemical additives from microplastics into seawater.

An automatic flow-based system as a front end to liquid chromatography (LC) for on-line dynamic leaching of microplastic materials (polyethylene of medium density and poly(vinyl chloride)) with incurred phthalates and bisphenol A is herein presented. The microplastic particles were packed in a metal column holder, through which seawater was pumped continuously by resorting to advanced flow methodology. Each milliliter of the leachable (bioaccessible) fraction of chemical additives was preconcentrated on-line using a 10 mm-long octadecyl monolithic silica column placed in the sampling loop of the injection valve of a HPLC system that served concomitantly for analyte uptake and removal of the seawater matrix. After loading of the leachate fraction, the LC valve was switched to the inject position and the analytes were eluted and separated by a monolithic column (Onyx C18HD 100 × 4.6 mm) using an optimized acetonitrile/water gradient with UV detection at 240 nm. The automatic flow method including dynamic flow-through extraction, on-line sorptive preconcentration, and matrix clean-up was synchronized with the HPLC separation, which lasted ca. 9 min. The only two currently available multi-component certified reference materials (CRM) of microplastics (CRM-PE002 and CRM-PVC001) were used for method development and validation. Out of the eight regulated phthalates contained in the two CRMs, only the 2 most polar species, namely, dimethyl phthalate and diethyl phthalate as well as bisphenol A, were leached significantly by the seawater in less than 2 h, with bioaccessibility percentages of 51-100%. The leaching profiles were monitored and modeled with a first-order kinetic equation so as to determine the rate constants for desorption in a risk assessment scenario. Intermediate precision values of bioaccessibility data for three batches of CRMs were for the suite of targeted compounds ≤22%. This work for the first time reports a fully automatic flow method with infinite sink capacity (i.e., using a surplus of extracting solution) for the target species able to mimic the leaching of additives from plastic debris across the water body in marine settings under worst-case extraction conditions.

Direct-immersion single-drop microextraction and in-drop stirring microextraction for the determination of nanomolar concentrations of lead using automated Lab-In-Syringe technique.

Two operational modes for Lab-In-Syringe automation of direct-immersion single-drop microextraction have been developed and critically compared using lead in drinking water as the model analyte. Dithizone was used in the presence of masking additives as a sensitive chromogenic complexing reagent. The analytical procedure was carried out inside the void of an automatic syringe pump. Normal pump orientation was used to study extraction in a floating drop of a toluene-hexanol mixture. Placing the syringe upside-down allowed the use of a denser-than-water drop of chloroform for the extraction. A magnetic stirring bar was placed inside the syringe for homogenous mixing of the aqueous phase and enabled in-drop stirring in the second configuration while resulting in enhanced extraction efficiency. The use of a syringe as the extraction chamber allowed drop confinement and support by gravitational differences in the syringe inlet. Keeping the stirring rates low, problems related to solvent dispersion such as droplet collection were avoided. With a drop volume of 60 µL, limits of detection of 75 nmol L-1 and 23 nmol L-1 were achieved for the floating drop extraction and the in-drop stirring approaches, respectively. Both methods were characterized by repeatability with RSD typically below 5%, quantitative analyte recoveries, and analyte selectivity achieved by interference masking. Operational differences were critically compared. The proposed methods permitted the routine determination of lead in drinking water to be achieved in less than 6 min.

Online coupling of fully automatic in-syringe dispersive liquid-liquid microextraction with oxidative back-extraction to inductively coupled plasma spectrometry for sample clean-up in elemental analysis: A proof of concept.

A proof of concept of a novel automatic sample cleanup approach for metal assays in troublesome matrixes as a front-end sample pre-treatment to inductively coupled plasma optical emission spectroscopy - ICP-OES - is herein presented. Target metals, namely, copper, lead, and cadmium were complexed in-system quantitatively using ammonium pyrrolidine dithiocarbamate (APDC) and transferred into a minute volume of toluene as extractant employing lab-in-syringe magnetic stirring-assisted dispersive liquid-liquid microextraction (LIS-MSA-DLLME). After discharge of the sample, the analytes were back-extracted into nitric acid and injected on-line into ICP-OES. To promote and expedite this process in-syringe, advantage was taken from oxidative decomposition of the chelate by potassium iodate, reported in this article for the first time. Experimental conditions for LIS-MSA-DLLME were optimized by Box-Benkhen multivariate analysis using the geometric mean of analyte recoveries as the desirability function. Times of extraction and back-extraction of 300s and 100s, respectively, pH 5.5 at 30mmol/L acetate, 300µL of extraction solvent, and 600µmol/L of APDC were finally applied. Online interfacing to ICP-OES for back-extract analysis yielded average repeatabilities for Cd, Cu, and Pb of 2.9%, 3.5%, and 3.5% with limits of detections (3s) of 1.9, 1.4, and 5.6ng/mL, respectively. Oxidative back-extraction was proven reliable for the determination of metal species in coastal seawater, surrogate digestive fluids and soil leachates with recovery values for Cd, Cu, and Pb ranging from 90% to 118%, 68% to 104%, and 86% to 112%, respectively.

Fully Automatic In-Syringe Magnetic Stirring-Assisted Dispersive Liquid-Liquid Microextraction Hyphenated to High-Temperature Torch Integrated Sample Introduction System-Inductively Coupled Plasma Spectrometer with Direct Injection of the Organic Phase.

A proof of concept study involving the online coupling of automatic dispersive liquid-liquid microextraction (DLLME) to inductively coupled plasma optical emission spectrometry (ICP OES) with direct introduction and analysis of the organic extract is herein reported for the first time. The flow-based analyzer features a lab-in-syringe (LIS) setup with an integrated stirring system, a Meinhard nebulizer in combination with a heated single-pass spray chamber, and a rotary injection valve, used as an online interface between the microextraction system and the detection instrument. Air-segmented flow was used for delivery of a fraction of the nonwater miscible extraction phase, 12 µL of xylene, to the nebulizer. All sample preparative steps including magnetic stirring assisted DLLME were carried out inside the syringe void volume as a size-adaptable yet sealed mixing and extraction chamber. Determination of trace level concentrations of cadmium, copper, lead, and silver as model analytes has been demonstrated by microextraction as diethyldithiophosphate (DDTP) complexes. The automatic LIS-DLLME method features quantitative metal extraction, even in troublesome sample matrixes, such as seawater, salt, and fruit juices, with relative recoveries within the range of 94-103%, 93-100%, and 92-99%, respectively. Furthermore, no statistically significant differences at the 0.05 significance level were found between concentration values experimentally obtained and the certified values of two serum standard reference materials.