C18-modified Silica Research Articles

A novel 3D-printed sample preparation method for benzodiazepine quantification in human serum.

Benzodiazepine abuse remains a significant public health concern. Current sample preparation methods for benzodiazepine analysis from human serum often involve complex procedures that require large sample volumes and extensive organic solvent use. To address these limitations, this study presents a novel and efficient sample preparation method utilizing 3D-printed sorbent devices. The 3D-printed devices, fabricated from a thermoplastic composite incorporating C18-modified silica, demonstrated exceptional performance in extracting benzodiazepines from human serum. The method was optimized and validated according to ICH guidelines, ensuring its reliability for quantitative benzodiazepine analysis. Notably, the method required minimal sample and solvent volumes, eliminating the need for protein precipitation, evaporation, and reconstitution. This novel sample preparation approach offers significant advantages over traditional methods, providing a more efficient and environmentally friendly solution for benzodiazepine analysis. The versatility of 3D printing allows for the customization of sorbent devices for various analytes and matrices, expanding the potential applications of this method. Coupled with a rapid and robust LC-MS method optimized with DryLab, this approach presents a valuable and sensitive tool for benzodiazepine monitoring in clinical and toxicological settings.

Hybrid Bilayer Interfaces within Reversed-Phase Chromatographic Silica Formed by Self-Assembly of Long-Chain Primary Alcohols.

Modification of silica interfaces by covalent attachment of functional ligands is a primary means of controlling the interfacial chemistry of porous silicas used in separations, environmental cleanup, and biosensing. Recently, modification of hydrophobic, n-alkyl-silane-functionalized interfaces has been achieved through self-assembly of zwitterionic phospholipids or mixed-charged surfactants to form "hybrid bilayers", producing interfaces that mimic lipid-bilayer partitioning and provide shape-selective partitioning of aromatic hydrocarbons. Charged headgroups, however, introduce electrostatic interactions that strongly influence the retention of ionizable solutes and require careful control over pH and ionic strength in the solution phase. In this work, we propose modification of C18-functionalized chromatographic silica surfaces through self-assembly of long-chain primary alcohols to form uncharged hybrid-bilayer surfaces. Hybrid bilayers formed from alcohols ranging from C12OH to C22OH are investigated with in situ confocal-Raman microscopy, and the spectra indicate that they form highly ordered n-alkane structures, with order increasing as a function of alcohol chain length. Temperature-dependent Raman spectra of C12OH-C22OH hybrid bilayers were collected to investigate their melting transitions. Multivariate curve resolution of these spectra show broad, two-component melting transitions, indicating alcohol and C18 alkyl chains melt simultaneously. These results suggest an interdigitated interfacial structure, where the hydrocarbon chains of the adsorbed alcohol extend into the underlying C18 chains, ordering both layers. Interdigitation is confirmed by a temperature-dependent study of a deuterated C16-OH bilayer, where spectrally resolved Raman bands from deuterated and protiated hydrocarbons melt together. Finally, n-alkyl alcohol bilayers were tested for protein repellency, where no protein adsorption was observed when equilibrated with ∼1 mg/mL bovine serum albumin. Bilayers C16OH in chain length are shelf stable at refrigerated temperatures for months. These results demonstrate long-chain alcohol bilayers can be utilized to control the interfacial hydrocarbon structure of C18-modified silica and have potential for use in separations, biosensing, and anti-biofouling applications.

3D-printed extraction devices fabricated from silica particles suspended in acrylate resin

In recent years, there has been an increasing worldwide interest in the use of alternative sample preparation methods. Digital light processing (DLP) is a 3D printing technique based on using UV light to form photo-curable resin layer upon layer, which results in a printed shape. This study explores the application of this technique for the development of novel drug extraction devices in analytical chemistry. A composite material consisting of a photocurable resin and C18-modified silica particles was employed as a sorbent device, demonstrating its effectiveness in pharmaceutical analysis. Apart from estimating optimal printing parameters, microscopic examination of the material surface, and sorbent powder to resin ratio, the extraction procedure was also optimised. Optimisation included the type and amount of sample matrix additives, desorption solvent, sorption and desorption times, and proper number of sorbent devices needed in extraction protocol. To demonstrate this method's applicability for sample analysis, the solid-phase extraction followed by gas chromatography coupled with mass spectrometry (SPE-GC–MS) method was validated for its ability to quantify benzodiazepine-type drugs. This evaluation confirmed good linearity in the concentration range of 50–1000 ng/mL, with R2 values being 0.9932 and 0.9952 for medazepam and diazepam, respectively. Validation parameters proved that the presented method is precise (with values ranging in-between 2.98 %-7.40 %), and accurate (88.81 % to 110.80 %). A negative control was also performed to investigate possible sorption properties of the resin itself, proving that the addition of C18-modified silica particles significantly increases the extraction efficiency and repeatability.The cost-effectiveness of this approach makes it particularly advantageous for single-use scenarios, eliminating the need for time-consuming sorbent-cleaning procedures, common in traditional solid-phase extraction techniques. Future optimisation opportunities include refining sorbent size, shape, and geometry to achieve lower limits of quantification. As a result of these findings, 3D-printed extraction devices can serve as a viable alternative to commercially available SPE or solid-phase microextraction (SPME) protocols for studying new sample preparation approaches.

On the identification and quantification of proton-initiated species in the synthesis of poly(2-alkyl-2-oxazoline)s by high resolution liquid chromatography

Hydrophilic poly(2-oxazoline)s represent a promising alternative to replace poly(ethylene glycol) in the biomedical field. For that purpose, reliable analytical protocols to confirm identity and quantity of impurities are required. In particular, side products deriving from chain transfer reactions occurring during the cationic ring-opening polymerization and incomplete end-capping processes may be present. The analytical approach must hence be capable of separating polymers according to minor changes regarding their end group. We demonstrate that liquid chromatography, relying on a monolithic C18-modified silica column and isocratic as well as gradient elution using water / acetonitrile mixtures and varying detectors, can accomplish such demanding high resolution separations. Poly(2-ethyl-2-oxazoline)s (PEtOx) with acetyl, hydroxyl, and phthalimide ω-end groups were investigated. Identification of side products was achieved through coupling with electrospray ionization mass spectrometry. UV / Vis detection was applied to quantify chain transfer products in PEtOx comprising biphenyl moieties. In addition, gradient elution enabled the separation of PEtOx into macromolecules according to their specific degrees of polymerization in molar mass ranges around 2,000 g mol-1.

Independent mode sorption of perfluoroalkyl acids by single and multiple adsorbents.

Infinite dilution partition coefficients, Kp,0, of a series of unbranched perfluoralkylacids, PFAAs with 3 to 8 CF2 units between water and commercially available weak anion exchange (WAX) and strong anion exchange (MAX) polymers, C18-modified silica, hydrophilic-lipophilic balance polymer (HLB), and Al2O3 sorbents were determined with self-packed columns using an HPLC-MS/MS setup. The anionic WAX sorbent shows a much higher adsorption affinity (about 450 fold) for PFBA than was observed for the applied hydrophobic sorbent HLB. Since the incremental value for each CF2 group is smaller when the electrostatic adsorption process is observed, the hydrophobic partition coefficient of HLB supersedes the electrostatic one of WAX at around PFTeDA. Adsorption of PFAAs to Al2O3 was weak and did not show a clear chain length dependency. A recently developed independent mode (IM) adsorption model is a more accurate model to combine the electrostatic and hydrophobic interaction terms. This model predicts the correct behaviour of especially short chain PFAAs in soil or sediment sorption experiments. Factors increasing sorption efficiency of well- and ill-defined single and multiple adsorbents towards PFAAs are discussed. The IM model provides a method to optimise sorption remediation strategies of PFAAs in contaminated waters and proposes a two-step strategy, a starting hydrophobic step followed by an electrostatic one to remove more efficiently the short chain PFAAs.

Hybrid-Supported Bilayers Formed with Mixed-Charge Surfactants on C18-Functionalized Silica Surfaces.

Mixtures of cationic-anionic surfactants have been shown to spontaneously form ordered monolayers at hydrophobic-hydrophilic boundaries, including air-water and oil-water interfaces. In this work, confocal Raman microscopy is used to investigate the structure of hybrid-supported surfactant bilayers (HSSBs) formed by deposition of a distal leaflet of mixed cationic-anionic surfactants onto a proximal leaflet of n-alkane (C18) chains on the interior surfaces of chromatographic silica particles. The surface coverage of the two surfactants in a hybrid bilayer was determined from carbon analysis and the relative Raman scattering of their respective head-groups. Within the measurement uncertainty, the stoichiometric ratio of the two surfactants is one-to-one, equivalent to mixed-charge-surfactant monolayers at air-water and oil-water interfaces and consistent with the role of the head-group electrostatic interactions in their formation. When self-assembled on the hydrophobic surface, pairs of oppositely charged n-alkyl chain surfactants resemble a phospholipid (phosphatidylcholine) molecule, with its zwitterionic head-group and two hydrophobic acyl chain tails. Indeed, the structure of these hybrid-supported surfactant bilayers on C18-modified silica surfaces is similar to that of hybrid-supported lipid bilayers (HSLBs) on the same supports, but with denser and more-ordered n-alkyl chains. Hybrid-supported surfactant bilayers exhibit a melting phase transition (gel to liquid-crystalline phase) with structural and energetic characteristics similar to those of hybrid-supported bilayers prepared from a zwitterionic phospholipid of the same alkyl chain length. These mixed-charge surfactants on n-alkane-modified silica are stable in water over time (months), results that suggest the potential use of these hybrid bilayers for generating supported lipid-bilayer-like surfaces or for separation applications.

Hydrophobicity of carbohydrates and related hydroxy compounds

The hydrophobic interaction of carbohydrates and other hydroxy compounds with a C18-modified silica gel column was measured with pure water as eluent, thereby expanding the range of measurements already published. The interaction is augmented by structure strengthening salts and decreasing temperature. Although the interaction of the solute with the hydrophobic interface is expected to only imperfectly reflect its state in aqueous bulk solution, the retention can be correlated to hydration numbers calculated from molecular mechanics studies given in the literature. No correlation can be established towards published hydration numbers obtained by physical methods (isentropic compressibility, O-17 NMR relaxation, terahertz spectroscopy, and viscosity). The hydrophobicity is discussed with respect to the chemical structure. It increases with the fraction and size of hydrophobic molecular surface regions.

Synthesis and modification of monodisperse silica microspheres for UPLC separation of C60 and C70

Using a modified Stöber method, monodisperse silica microspheres with average diameters from 1 μm to 2 μm are synthesized as UPLC column fillers. With a column as short as 50 mm, the 1.5 μm C18-modified silica stationary phase can separate C60 and C70 in 7 minutes with ultra high efficiency.

Confocal Raman Microscopy for in Situ Measurement of Octanol-Water Partitioning within the Pores of Individual C18-Functionalized Chromatographic Particles.

Octanol-water partitioning is one of the most widely used predictors of hydrophobicity and lipophilicity. Traditional methods for measuring octanol-water partition coefficients (K(ow)), including shake-flasks and generator columns, require hours for equilibration and milliliter quantities of sample solution. These challenges have led to development of smaller-scale methods for measuring K(ow). Recent advances in microfluidics have produced faster and smaller-volume approaches to measuring K(ow). As flowing volumes are reduced, however, separation of water and octanol prior to measurement and detection in small volumes of octanol phase are especially challenging. In this work, we reduce the receiver volume of octanol-water partitioning measurements from current practice by six-orders-of-magnitude, to the femtoliter scale, by using a single octanol-filled reversed-phase, octadecylsilane-modified (C18-silica) chromatographic particle as a collector. The fluid-handling challenges of working in such small volumes are circumvented by eliminating postequilibration phase separation. Partitioning is measured in situ within the pore-confined octanol phase using confocal Raman microscopy, which is capable of detecting and quantifying a wide variety of molecular structures. Equilibration times are fast (less than a minute) because molecular diffusion is efficient over distance scales of micrometers. The demonstrated amount of analyte needed to carry out a measurement is very small, less than 50 fmol, which would be a useful attribute for drug screening applications or testing of small quantities of environmentally sensitive compounds. The method is tested for measurements of pH-dependent octanol-water partitioning of naphthoic acid, and the results are compared to both traditional shake-flask measurements and sorption onto C18-modified silica without octanol present within the pores.

Systematic investigation of factors controlling supercritical fluid extraction (SFE) of spiked and aged PCBs from edible tissues of the blue crab (Callinectes sapidus).

Systematic investigation of factors controlling supercritical fluid extraction (SFE) of spiked and naturally incurred (aged) PCBs from edible tissues of the Blue Crab (Callinectes sapidus) was undertaken. Effects of extraction pressure, temperature and time, CO2 flow rate and total volume, and collection temperature were assessed. Temperature dramatically impacted extraction efficiency, especially at lower pressures. Surprisingly, extraction of both spiked and aged PCBs was flow rate dependent, counter to prevailing views regarding the relative ease of SFE of spiked versus aged contaminants from environmental matrices. PCBs were optimally trapped on a 1:1 mixture of C18-modified and porous silica at 0°C and eluted with <2 mL isooctane at 90°C. A combined 10 min static/30 min dynamic extraction at 35.5 MPa and 150°C with a CO2 flow rate of 3 mL min(-1) yielded maximum (quantitative) recoveries of spiked and aged PCBs. Resulting solvent extracts required no cleanup and could be analyzed directly by halogen-selective GC with MS confirmation.

Direct observation of the orientation dynamics of single protein-coated nanoparticles at liquid/solid interfaces.

We have observed the rotational dynamics of single protein-coated gold nanorods (AuNRs) on C18-modified silica surfaces in real time by dual-channel polarization dark-field microscopy. Four different rotational states were identified, depending on the apparent strength of interactions between the AuNRs and the surface. The distributions of the states could be regulated by adjusting the salt concentration, and the state transitions were verified by monitoring the entire desorption process of a single AuNR. Our study provides insight into the interfacial orientation and dynamics of nanoparticles and could be useful for in vitro biophysics and the separation of proteins.

Hydrophilic Interaction Chromatography Using a Meter-Scale Monolithic Silica Capillary Column for Proteomics LC-MS

A meter-scale monolithic silica capillary column modified with urea-functional groups for hydrophilic interaction liquid chromatography (HILIC) was developed for highly efficient separation of biological compounds. We prepared a ureidopropylsilylated monolithic silica capillary column with a minimum plate height of 12 μm for nucleosides and a permeability of 2.1 × 10(-13) m(2), which is comparable with the parameters of monolithic silica-C18 capillary columns. Over 300,000 theoretical plates were experimentally obtained in HILIC with a 4 m long column at 8 MPa; this is the best result yet reported for HILIC. A 2 m long ureidopropylsilylated monolithic silica capillary column was utilized to develop a HILIC mode LC-MS system for proteomics applications. Using tryptic peptides from human HeLa cell lysate proteins, we identified the comparable numbers of peptides and proteins in HILIC with those in reversed-phase liquid chromatography (RPLC) using a C18-modified monolithic silica column when shallow gradients were applied. In addition, approximately 5-fold increase in the peak response on average was observed in HILIC for commonly identified tryptic peptides due to the high acetonitrile concentration in the HILIC mobile phase. Since HILIC mode LC-MS shows orthogonal selectivity to RPLC mode LC-MS, it is useful as a complementary tool to increase proteome coverage in proteomics studies.

Ionic liquids-assisted fabrication of silica-based monolithic columns

The mesopores of a monolithic silica column are very important and useful for chromatographic separation since they can offer sufficiently large surface area. In this paper, a novel method with the assistance of an ionic liquid (1-butyl-3-methylimidazolium tetrafluoroborate ([bmin]BF 4)) was developed for the preparation of a C18-modified monolithic silica column for the first time, in which, the through pores and mesopores were formed simultaneously during the sol–gel reaction. The method is effective to simplify the preparation process of the silica-based monolithic columns. The factors influencing the sol–gel process, including the content of methanol and pH, were studied. The chromatographic performance of the prepared monolithic column was evaluated by the separation of alkylbenzenes.

Microfluidic “thin chips” for chemical separations

This paper describes the design, development and application of microfluidic "thin chips" fabricated from PDMS. Thin chips consist of multiple layers of PDMS chemically bonded onto each other. Unlike thicker PDMS chips that suffer from lack of sensitivity due to PDMS absorption in the VIS and UV range, the thinness of these chips allows for the detection of chromophoric species within the microchannel via an external fiber optics detection system. C18-modified reversed-phase silica particles are packed into the microchannel using a temporary taper created by a magnetic valve and separations using both pressure- and electrochromatographic-driven methods are detailed.

Methacrylate monolithic capillary columns for gradient peptide separations

For the separation of peptides with gradient-elution liquid chromatography a poly(butyl methacrylate- co-ethylene dimethacrylate) (BMA) monolithic capillary column was prepared and tested. The conditional peak capacity was used as a metric for the performance of this column, which was compared with a capillary column packed with C18-modified silica particles. The retention of the peptides was found to be smaller on the BMA column than on the particulate C18 column. To obtain the same retention in isocratic elution an approximately 15% (v/v) lower acetonitrile concentration had to be used in the mobile phase. The retention window in gradient elution was correspondingly smaller with the BMA column. The relation between peak width and retention under gradient conditions was studied in detail. It was found that in shallow gradients, with gradient times of 30 min and more, the peak widths of the least retained compounds are strongly increased with the BMA column. This was attributed to the fact that these compounds migrate and elute with an unfavorable high retention factor. More retained compounds are eluted later in the gradient, but with a lower effective retention factor. With shallow gradients the peak capacity of the BMA column (≈90) was clearly lower than that of a conventional packed column (≈150). On the other hand, with steep gradients, when components elute with a low effective retention factor, the performance of the BMA column is relatively good. With a gradient time of 15 min similar peak widths and thus similar peak capacities (≈75) were found for the packed and the monolithic column. Two strategies were investigated to obtain higher peak capacities with methacrylate monolithic columns. The use of lauryl methacrylate (LMA) instead of butyl methacrylate (BMA) gave an increase in retention and narrower peaks for early eluting peptides. The peak capacity of the LMA column was ≈125 in a 60 min gradient. Another approach was to use a longer BMA column which resulted in a peak capacity of ≈135 could be obtained in 60 min.

Large‐pore mesoporous SBA‐15 silica particles with submicrometer size as stationary phases for high‐speed CEC separation

A new mesoporous sphere-like SBA-15 silica was synthesized and evaluated in terms of its suitability as stationary phases for CEC. The unique and attractive properties of the silica particle are its submicrometer particle size of 400 nm and highly ordered cylindrical mesopores with uniform pore size of 12 nm running along the same direction. The bare silica particles with submicrometer size have been successfully employed for the normal-phase electrochromatographic separation of polar compounds with high efficiency (e.g., 210,000 for thiourea), which is matched well with its submicrometer particle size. The Van Deemeter plot showed the hindrance to mass transfer because of the existence of pore structure. The lowest plate height of 2.0 microm was obtained at the linear velocity of 1.1 mm/s. On the other hand, because of the relatively high linear velocity (e.g., 4.0 mm/s) can be generated, high-speed separation of neutral compounds, anilines, and basic pharmaceuticals in CEC with C18-modified SBA-15 silica as stationary phases was achieved within 36, 60, and 34 s, respectively.

Effect of surface coverage on the conformation and mobility of C18-modified silica gels

C18-modified silica gels with surface coverages of 2 to 8.2 micromol m(-2), were prepared by different synthetic pathways and characterized by Fourier Transform infrared spectroscopy (FTIR), solid-state nuclear magnetic resonance (NMR) spectroscopy, and chromatographic measurements. The effects of temperature and bonding density on the conformational order of C18-modified silica gels were studied in detail by FTIR spectroscopy. The silane functionality and degree of cross-linking of silane ligands on the silica surface were evaluated by 29Si cross-polarization magic-angle spinning (CP/MAS) NMR and the structural order and mobility of the alkyl chains were investigated by 13C CP/MAS NMR spectroscopy. CH2 symmetric and anti-symmetric stretching bands and CH2 wagging bands were used as IR probes to monitor the conformational order and flexibility of the alkyl chains in the C18 phases. Qualitative information about the conformational order was obtained from frequency shifts of the CH2 symmetric and anti-symmetric stretching bands. The relative amounts of kink/gauche-trans-gauche, double-gauche, and end-gauche conformers in the alkyl chains were determined by analysis of CH2 wagging bands. These results indicate that surface coverage plays a dominant role in the conformational order of C18-modified silica gels. The FTIR and NMR data are discussed in the context of the chromatographic shape-selectivity differences.

Improved liquid chromatographic determination of nine currently used (fluoro)quinolones with fluorescence and mass spectrometric detection for environmental samples

An HPLC method using C18-modified silica as stationary phase has been developed for environmental trace analysis of nine (fluoro)quinolones. Detection is done by fluorescence measurement or MS using the modes of SIM and selected reaction monitoring (SRM). Best separation is achieved with a gradient consisting of 50 mM formic acid and methanol, which is fully compatible with MS coupling. LOQs (S/N of 10) for fluorescence detection are between 10 and 60 microg/L, depending on the analyte. MS detection (SIM and SRM) yields LOQs that are better by a factor of at least an order of magnitude. Sample preconcentration and sample clean-up is accomplished by SPE (preconcentration factor of 1000), leading to LOQs in the low ng/L range. Recoveries of the preconcentration procedure are better than 80% for all analytes. The suitability for real samples has been demonstrated by analyzing surface waters, municipal waste waters, sewage treatment plant effluents, sewage sludge, and sediment taken from rivers and fish ponds. The method should also be useful for determination of residues of (fluoro)quinolones in food or other matrices. The degradation of the (fluoro)quinolones has been examined over 5 days in order to get information about the decomposition rate and the degradation products eventually occurring in the environment.

Dansylation of Endocrine-disrupting Alkylphenolic Compounds on the Solid Phase Used for Extraction from Environmental Water

Abstract The dansylation reaction of endocrine-disruptive alkylphenols on a C18-modified silica gel (ODS) instead of organic solvents was investigated with the aim of developing an environment-friendly and efficient derivatization method for the HPLC analyses. The solid-phase aqueous dansylation system has been incorporated into a preconcentration step performed by solid-phase extraction (SPE) from tap water.

Spectrofluorimetric determination of surface-bound thiol groups and its application to the analysis of thiol-modified silicas.

A sensitive spectofluorimetric method has been developed for the quantitative measurement of surface-bound thiol groups. The procedure is based on the quantitative esterification of the thiol group with Rhodamine B and its subsequent release from the solid by base hydrolysis for spectrofluorimetric determination. Application of the method to the analysis of thiol-modified nanoparticulate silicas yielded results that compared favourably against alternative approaches based on measurements of mercury capacity and iodometric titration of the thiol groups. Non-specific Rhodamine uptake, assessed using unmodified silica and C18-modified silica, did not significantly influence the analytical results. When applied to a typical 50 mg sample, the detection limit of the procedure was 1 nmole SH g(-1) silica.