Chromatography doi: 10.3390/chromatography2040691

Authors: Haleigh Boswell Frank Dorman

Analysis of blood alcohol concentration is a routine analysis performed in many forensic laboratories. This analysis commonly utilizes static headspace sampling, followed by gas chromatography combined with flame ionization detection (GC-FID). Studies have shown several “optimal” methods for instrumental operating conditions, which are intended to yield accurate and precise data. Given that different instruments, sampling methods, application specific columns and parameters are often utilized, it is much less common to find information on the robustness of these reported conditions. A major problem can arise when these “optimal” conditions may not also be robust, thus producing data with higher than desired uncertainty or potentially inaccurate results. The goal of this research was to incorporate the principles of quality by design (QBD) in the adjustment and determination of BAC (blood alcohol concentration) instrumental headspace parameters, thereby ensuring that minor instrumental variations, which occur as a matter of normal work, do not appreciably affect the final results of this analysis. This study discusses both the QBD principles as well as the results of the experiments, which allow for determination of more favorable instrumental headspace conditions. Additionally, method detection limits will also be reported in order to determine a reporting threshold and the degree of uncertainty at the common threshold value of 0.08 g/dL. Furthermore, the comparison of two internal standards, n-propanol and t-butanol, will be investigated. The study showed that an altered parameter of 85 °C headspace oven temperature and 15 psi headspace vial pressurization produces the lowest percent relative standard deviation of 1.3% when t-butanol is implemented as an internal standard, at least for one very common platform. The study also showed that an altered parameter of 100 °C headspace oven temperature and 15-psi headspace vial pressurization produces the lowest MDL of 0.00002 g/dL when n-propanol is implemented as an internal standard. These altered headspace parameters have the potential to produce more precise and accurate BAC determination.

Chromatography doi: 10.3390/chromatography2040669

Authors: James Grinias Bernard Bunner Martin Gilar James Jorgenson

As column volumes continue to decrease, extra-column band broadening has become an increasingly important consideration when determining column performance. Combined contributions due to the injector and connecting tubing in a capillary LC system were measured and found to be larger than expected by Taylor-Aris theory. Variance from sigma-type and tau-type broadening was isolated from eluted peaks using the Foley-Dorsey Exponentially Modified Gaussian peak fitting model and confirmed with computational fluid dynamics. It was found that the tau-type contributions were the main cause for the excessive broadening because of poorly-swept volumes at the connection between the injector and tubing. To reduce tau-type contributions (and peak tailing), a timed pinch mode could be used for analyte injection.

Chromatography doi: 10.3390/chromatography2040655

Authors: Norbert Löwa Patricia Radon Dirk Gutkelch Rinaldo August Frank Wiekhorst

Magnetic nanoparticles (MNPs) exhibit unique magnetic properties making them ideally suited for a variety of biomedical applications. Depending on the desired magnetic effect, MNPs must meet special magnetic requirements which are mainly determined by their structural properties (e.g., size distribution). The hyphenation of chromatographic separation techniques with complementary detectors is capable of providing multidimensional information of submicron particles. Although various methods have already been combined for this approach, so far, no detector for the online magnetic analysis was used. Magnetic particle spectroscopy (MPS) has been proven a straightforward technique for specific quantification and characterization of MNPs. It combines high sensitivity with high temporal resolution; both of these are prerequisites for a successful hyphenation with chromatographic separation. We demonstrate the capability of MPS to specifically detect and characterize MNPs under usually applied asymmetric flow field-flow fractionation (A4F) conditions (flow rates, MNP concentration, different MNP types). To this end MPS has been successfully integrated into an A4F multidetector platform including dynamic ligth scattering (DLS), multi-angle light scattering (MALS) and ultraviolet (UV) detection. Our system allows for rapid and comprehensive characterization of typical MNP samples for the systematic investigation of structure-dependent magnetic properties. This has been demonstrated by magnetic analysis of the commercial magnetic resonance imaging (MRI) contrast agent Ferucarbotran (FER) during hydrodynamic A4F fractionation.

Chromatography doi: 10.3390/chromatography2040642

Authors: Troy Purvis

This article details the elements used in the method verification for the simultaneous high performance liquid chromatography (HPLC) assay of Pentoxifylline, Mupirocin, Itraconazole, and Fluticasone Propionate in Humco™ Lavare Wound base. The method was proven to be linear over 50%–150% of the nominal concentration of the standards. The method was proven to be accurate over 50%–150%, with 98%–102% recovery of the actives from spiked placeboes over that range. The method was shown to be specific to the analytes listed and precise, yielding acceptable results for system reproducibility and method repeatability. The method, as written, is considered to have been verified.

Chromatography doi: 10.3390/chromatography2040625

Authors: Juan Baeza-Baeza Casandra Ortiz-Bolsico María García-Alvarez-Coque

Traditionally, column performance in liquid chromatography has been studied using information from the elution of probe compounds at different flow rates through van Deemter plots, which relate the column plate height to the linear mobile phase velocity. A more recent approach to characterize columns is the representation of the peak widths (or the right and left peak half-widths) for a set of compounds versus their retention times, which, for isocratic elution, give rise to almost linear plots. In previous work, these plots have been shown to facilitate the prediction of peak profiles (width and asymmetry) with optimization purposes. In this work, a detailed study on the dependence of the peak widths (or half-widths) on the flow rate is reported. A new approach to quantify the deterioration of column performance for slow and fast flow rates and to characterize chromatographic columns is proposed. The approach makes use of the width (or half-widths) for a set of compounds with similar interaction kinetics and does not require knowledge of the extra-column contributions to the total variance. The chromatographic data of two sets of compounds of different natures (sulfonamides and β-blockers), eluted from Spherisorb and Chromolith columns with acetonitrile-water mixtures, are used to illustrate the approach.

Chromatography doi: 10.3390/chromatography2040611

Authors: Courtney Thomas Danny Reible

Volatile losses were measured from polydimethylsiloxane (PDMS) passive samplers during determination of contaminant porewater concentrations in sediments. Volatile losses could occur between the time of retrieval and processing of the passive sampler or in intertidal environments where the passive sampler could potentially be exposed above the water surface at low tide. A model was developed to predict losses of absorbed compounds as a function of sorbent geometry and the Henry’s Law Coefficient and PDMS-water partition coefficient of the compound of interest. The model suggests that thin layers of PDMS typically used to minimize equilibration times in passive sampling (≤30 µm) may not provide quantitative measurement of naphthalenes or other lighter volatile compounds without special efforts to reduce losses. The results suggest that the samplers should be processed rapidly onsite or kept at low temperatures after retrieval to maximize retention of more volatile compounds or designed with thick PDMS layers. The results also suggest that less volatile compounds, including phenanthrene, and higher molecular weight polynuclear aromatic hydrocarbons (PAHs) exhibit minimal evaporative losses with typical sample processing times.

Chromatography doi: 10.3390/chromatography2040594

Authors: Tonguc Tasci William Johnson Diego Fernandez Eliana Manangon Bruce Gale

Electrical Field Flow Fractionation (ElFFF) is a sub method in the field flow fractionation (FFF) family that relies on an applied voltage on the channel walls to effect a separation. ElFFF has fallen behind some of the other FFF methods because of the optimization complexity of its experimental parameters. To enable better optimization, a particle based model of the ElFFF systems has been developed and is presented in this work that allows the optimization of the main separation parameters, such as electric field magnitude, frequency, duty cycle, offset, flow rate and channel dimensions. The developed code allows visualization of individual particles inside the separation channel, generation of realistic fractograms, and observation of the effects of the various parameters on the behavior of the particle cloud. ElFFF fractograms have been generated via simulations and compared with experiments for both normal and cyclical ElFFF. The particle visualizations have been used to verify that high duty cycle voltages are essential to achieve long retention times and high resolution separations. Furthermore, by simulating the particle motions at the channel outlet, it has been demonstrated that the top channel wall should be selected as the accumulation wall for cyclical ElFFF to reduce band broadening and achieve high efficiency separations. While the generated particle based model is a powerful tool to estimate the outcomes of the ElFFF experiments and visualize particle motions, it can also be used to design systems with new geometries which may lead to the design of higher efficiency ElFFF systems. Furthermore, this model can be extended to other FFF techniques by replacing the electrical field component of the model with the fields used in the other FFF techniques.

Chromatography doi: 10.3390/chromatography2030580

Authors: Martha Wells

Fulvic (FAs) and humic acids (HAs) are chemically fascinating. In water, they have a strong propensity to aggregate, but this research reveals that tendency is regulated by ionic strength. In the environment, conductivity extremes occur naturally—freshwater to seawater—warranting consideration at low and high values. The flow field flow fractionation (flow FFF) of FAs and HAs is observed to be concentration dependent in low ionic strength solutions whereas the corresponding flow FFF fractograms in high ionic strength solutions are concentration independent. Dynamic light scattering (DLS) also reveals insight into the conductivity-dependent behavior of humic substances (HSs). Four particle size ranges for FAs and humic acid aggregates are examined: (1) <10 nm; (2) 10 nm–6 µm; (3) 6–100 µm; and (4) >100 µm. Representative components of the different size ranges are observed to dynamically coexist in solution. The character of the various aggregates observed—such as random-extended-coiled macromolecules, hydrogels, supramolecular, and micellar—as influenced by electrolytic conductivity, is discussed. The disaggregation/aggregation of HSs is proposed to be a dynamic equilibrium process for which the rate of aggregate formation is controlled by the electrolytic conductivity of the solution.

Chromatography doi: 10.3390/chromatography2030567

Authors: Anthony Malanoski Brandy Johnson Jeffrey Erickson Carrie Sun Connie Scoggins Mansoor Nasir Jason Bongard Martin Moore

This study sought to demonstrate the impact of surface-target interactions in a device bearing geometrical features through evaluations of chemical modification to the surface and alteration of the charge state of both surface and targets. The movement of commercial fluorescent nanoparticles through a field of geometric features was monitored based on the output of the field measured at different points transverse to the direction of flow. Modification of the device surfaces using carboxylate, amine, phenyl, and fluorinated groups was considered. The functional group on the surface was found to significantly impact biased particle movement in the device with amine groups leading to the greatest differences. Other factors, such as flow rate, pH, and initial particle concentration, were also found to have a significant impact on particle concentrations across the feature field.

Chromatography doi: 10.3390/chromatography2030545

Authors: Muriel Bouby Nicolas Finck Horst Geckeis

The magnesian smectite hectorite is a corrosion product frequently detected in nuclear waste glass alteration experiments. The structural incorporation of a single trivalent lanthanide was previously demonstrated. Hectorite was presently synthesized, for the first time, in the presence of several lanthanides (La, Eu, Yb) following a multi-step synthesis protocol. The smallest-sized particles (nanoparticles, NPs) were isolated by centrifugation and analyzed by asymmetrical flow field-flow fractionation (AsFlFFF) coupled to ICP-MS, in order to obtain information on the elemental composition and distribution as a function of the size. Nanoparticles can be separated from the bulk smectite phase. The particles are able to accommodate even the larger-sized lanthanides such as La, however, with lower efficiency. We, therefore, assume that the incorporation proceeds by substitution for octahedral Mg accompanied by a concomitant lattice strain that increases with the size of the lanthanides. The presence of a mixture does not seem to affect the incorporation extent of any specific element. Furthermore, syntheses were performed where in addition the tetravalent zirconium or thorium elements were admixed, as this oxidation state may prevail for many actinide ions in a nuclear waste repository. The results show that they can be incorporated as well.

Chromatography doi: 10.3390/chromatography2030529

Authors: Murugesapillai Mylvaganam Beth Binnington Monique Budani Anna Soltyk Clifford Lingwood

Although globotetraosylceramide (Gb4) is only recognized by a single member of the verotoxin family namely, the pig edema disease toxin (VT2e), removal of the acetyl group from the terminal N-acetyl hexosamine of Gb4 to generate the free amino sugar containing species (aminoGb4) results in the generation of a glycolipid preferentially recognized by all members of the verotoxin family (i.e., VT1, VT2, VT2c, and VT2e). GT3, a site-specific mutant of VT2e, in which Gb4 recognition is lost but Gb3 binding is retained, also binds aminoGb4. We have now compared the binding of VT1, VT2, VT2e, and GT3 to a series of aminoGb4 derivatives using a TLC overlay technique. DimethylaminoGb4 is bound by VT1 and VT2 but not VT2e or GT3; formylaminoGb4 binds all toxins but poorly to VT2 and preferentially VT2e; trifluoroacetylaminoGb4 binds only VT2e and GT3; isopropylaminoGb4 binds VT1 and poorly to VT2; benzylaminoGb4 binds all four toxins. Thus, there is a marked distinction between the permissible amino substitutions for VT1 and VT2e binding. GT3 is a hybrid between these in that, according to the substitution, it behaves similarly either to VT1 or to VT2e. For each species, GT3 does not however, show a hybrid binding between that of VT1 and VT2e. Analysis of the binding as a function of pH shows opposite effects for VT1 and VT2e: decreased pH increases VT1, but decreases VT2e receptor glycolipid binding.

Chromatography doi: 10.3390/chromatography2030515

Authors: Neus Jornet-Martínez Pascual Serra-Mora Yolanda Moliner-Martínez Rosa Herráez-Hernández Pilar Campíns-Falcó

In the present work, the performance of carbon nanotubes (c-CNTs) functionalized polydimethylsiloxane (PDMS) based coatings as extractive phases for in-tube solid phase microextraction (IT-SPME) coupled to Capillary LC (CapLC) has been evaluated. Carboxylic-single walled carbon nanotubes (c-SWNTs) and carboxylic-multi walled carbon nanotubes (c-MWNTs) have been immobilized on the activated surface of PDMS capillary columns. The effect of different percentages of diphenyl groups in the PDMS extractive phase has also been evaluated. The extraction capability of the capillary columns has been tested for different organic pollutants, nitrogen heterocyclic compounds and polycyclic aromatic compounds (PAHs). The results indicated that the use of the c-CNTs-PDMS capillary columns improve pyriproxyfen and mainly PAH extraction. Triazines were better extracted by unmodified TRB-35 and modified c-CNTs-PDMSTRB-5. The results showed that the extraction capability of the c-CNT capillary columns depends not only on the polarity of the analytes (as it occurs with PDMS columns) but also on the interactions that the analytes can establish with the immobilized c-CNTs on the PDMS columns. The extraction efficiency has been evaluated on the basis of the preconcentration rate that can be achieved, and, in this sense, the best c-CNTs-PDMS capillary column for each group of compounds can be proposed.

Chromatography doi: 10.3390/chromatography2030502

Authors: James Grinias Robert Kennedy

Large-size (4–5 µm) superficially porous particles yield lower plate heights (e.g., the minimal reduced plate height or hmin ≈ 1.5) than fully porous particles of a similar size when packed into large-bore columns. This property allows for better chromatographic performance without the higher pressures required for smaller particles. This study explores the use of such particles in microfluidic LC columns where materials and fitting pressure limits can constrain the size of particle used. The theoretically predicted performance improvements compared to fully porous particles were not demonstrated in capillary columns (with hmin ≈ 2 for both particle types), in agreement with previous studies that examined smaller superficially porous particles. Microfluidic columns were then compared to capillary columns. Capillary columns significantly outperformed microfluidic columns due to imperfections imposed by microfluidic channel asymmetry and world-to-chip connection at the optimal flow rate; however, superficially porous particles packed in microfluidic LC columns had flatter plate height versus flow rate curves indicating potential for better performance at high reduced velocities.

Chromatography doi: 10.3390/chromatography2030488

Authors: Jackson Frantz Fadi Alkhateeb Kevin Thurbide

The use of a novel micro pressurized liquid extraction (µPLE) method for the isolation of 16 priority pollutant polycyclic aromatic hydrocarbons (PAHs) from various solid samples is explored. The technique employs rapid heating in a single static extraction mode to remove analytes in a matter of seconds from 5–10 mg samples using only 125 µL of solvent. For example, results show that 30 s extractions with toluene at 200 °C produce respective PAH recovery ranges of 90%–130% and 88%–114% from samples of soil and smoked chicken. Comparatively, solids containing significant amounts of biochar were more challenging to extract from. For instance, when using a pure biochar sample matrix, recoveries for the 16 PAHs range from only 33%–66% after 60 s of extraction with toluene at 200 °C. Overall, these extraction results agree very well with those reported when using conventional methods on similar samples. Therefore, the findings indicate that µPLE can potentially provide an alternative sample preparation method for PAHs that is both very rapid and requires little solvent.

Chromatography doi: 10.3390/chromatography2030472

Authors: Joontaek Park Anand Mittal

We developed an improved model for predicting the steric-entropic effect on the separation behaviors of rod-like particles in flow field-flow fractionation. Our new model incorporates the “pole-vault” rotation of a rod-like particle near a wall under shear flow into the original model developed by Beckett and Giddings which considered only Brownian rotation. We investigated the effect of the aspect ratio on the retention ratios and the cross-sectional concentration distribution in the separation of rods in field-flow fractionation (FFF). Our analyses involved comparing the results predicted using the original model and those from the new model under various rod geometries and flow conditions. We found that the new model can show the aspect ratio-enhanced elution trend in certain flow conditions for the assumption of non-constant cloud thickness (ratio between the cross flow rate and the rod diffusivity). We also deducted that the flow conditions allowing for the aspect ratio-enhanced elution are related to the interplay among the axial flow rate, cloud thickness, and rod geometry. The new model can be viewed as a prototype to qualitatively show the aspect ratio-enhanced trend since its quantitative agreement with the experimental data must be improved for our future work.

Chromatography doi: 10.3390/chromatography2030452

Authors: Cristina Félez Anna Molet Oscar Núñez

A field-amplified sample injection-micellar electrokinetic chromatography (FASI-MEKC) method for the determination of 14 benzophenones (BPs) in a food simulant used in migration studies of food packaging materials was developed, allowing almost baseline separation in less than 21 min. The use of a 10 mM sodium dodecyl sulfate (SDS) solution as sample matrix was mandatory to achieve FASI enhancement of the analyzed BPs. A 21- to 784-fold sensitivity enhancement was achieved with FASI-MEKC, obtaining limits of detection down to 5.1–68.4 µg/L, with acceptable run-to-run precisions (RSD values lower than 22.3%) and accuracy (relative errors lower than 21.0%). Method performance was evaluated by quantifying BPs in the food simulant spiked at 500 µg/L (bellow the established specific migration limit for BP (600 µg/L) by EU legislation). For a 95% confidence level, no statistical differences were observed between found and spiked concentrations (probability at the confidence level, p value, of 0.55), showing that the proposed FASI-MEKC method is suitable for the analysis of BPs in food packaging migration studies at the levels established by EU legislation.

Chromatography doi: 10.3390/chromatography2030436

Authors: Haydn Mitchell Spencer Schultz Philip Costanzo Andres Martinez

The thermally responsive hydrogel N,N'-methylenebisacrylamide-cross-linked poly(N-isopropylacrylamide) (PNIPAM) was developed and evaluated as a reagent storage and delivery system for microfluidic paper-based analytical devices (microPADs). PNIPAM was shown to successfully deliver multiple solutions to microPADs in specific sequences or simultaneously in laminar-flow configuration and was found to be suitable for delivering four classes of reagents to the devices: Small molecules, enzymes, antibodies and DNA. PNIPAM was also able to successfully deliver a series of standard glucose solutions to microPADs equipped to perform a colorimetric glucose assay. The results of these tests were used to produce an external calibration curve, which in turn was used to determine the concentration of glucose in sample solutions. Finally, PNIPAM was used to store the enzyme horseradish peroxidase for 35 days under ambient conditions with no significant loss of activity. The combination of PNIPAM and microPADs may allow for more complex assays to be performed on paper-based devices, facilitate the preparation of external calibration curves in the field, and extend the shelf life of microPADs by stabilizing reagents in an easy-to-use format.

Chromatography doi: 10.3390/chromatography2030422

Authors: Thao Nguyen Jingyu Liu Vincent Hackley

Gold nanorods (GNRs) are of particular interest for biomedical applications due to their unique size-dependent longitudinal surface plasmon resonance band in the visible to near-infrared. Purified GNRs are essential for the advancement of technologies based on these materials. Used in concert, asymmetric-flow field flow fractionation (A4F) and single particle inductively coupled mass spectrometry (spICP-MS) provide unique advantages for fractionating and analyzing the typically complex mixtures produced by common synthetic procedures. A4F fractions collected at specific elution times were analyzed off-line by spICP-MS. The individual particle masses were obtained by conversion of the ICP-MS pulse intensity for each detected particle event, using a defined calibration procedure. Size distributions were then derived by transforming particle mass to length assuming a fixed diameter. The resulting particle lengths correlated closely with ex situ transmission electron microscopy. In contrast to our previously reported observations on the fractionation of low-aspect ratio (AR) GNRs (AR < 4), under optimal A4F separation conditions the results for high-AR GNRs of fixed diameter (≈20 nm) suggest normal, rather than steric, mode elution (i.e., shorter rods with lower AR generally elute first). The relatively narrow populations in late eluting fractions suggest the method can be used to collect and analyze specific length fractions; it is feasible that A4F could be appropriately modified for industrial scale purification of GNRs.

Chromatography doi: 10.3390/chromatography2030410

Authors: Fabrizio Cincotta Antonella Verzera Gianluca Tripodi Concetta Condurso

The sesquiterpene compounds present in red wines were characterized and quantified by Headspace Solid-Phase Microextraction in combination with Gas Chromatography–Mass Spectrometry (HS-SPME-GC-MS). Sixteen sesquiterpenes were identified, mainly hydrocarbons but also derived oxygenated compounds. Sesquiterpenes were acyclic, monocyclic, byciclic and tryciclic. Sesquiterpenes were detected in SIM (selected ion monitoring) mode using their characteristics ions. All the sesquiterpenes were identified by mass spectral data, linear retention indices (LRI), literature data and injection of standards where available. Quantitative results were obtained using the method of standard additions. The method showed an average LOD = 0.05 µg L−1 and LOQ = 0.15 µg L−1. The monocyclic sesquiterpene with the germacrene skeleton, Germacrene D and the bicyclic sesquiterpene with the muurolane skeleton, α-muurolene were present in all the wine samples analysed. Syrah wines were the samples richest in sesquiterpenes in agreement with their typical spicy and woody notes. The results evidenced the possibility to use sesquiterpenes for wine authenticity and traceability.

Chromatography doi: 10.3390/chromatography2030392

Authors: Tyler Shendruk Gary Slater

A novel field-flow fractionation (FFF) technique, in which two opposing external forces act on the solute particles, is proposed. When the two external forces are sufficiently strong and scale differently as a function of the solutes’ property of interest (such as the solute particle size), a sharp peak in the retention ratio (dramatic drop in elution time) is predicted to exist. Because the external forces oppose one another, we refer to this novel technique as adverse-mode FFF. The location of this peak is theoretically predicted and its ideal width estimated. The peak can become quite sharp by simultaneously increasing the strength of both fields, suggesting that adverse-mode FFF could be a useful technique for accurately measuring single species solute size.

Chromatography doi: 10.3390/chromatography2030382

Authors: Eri Moriyama Hiroyuki Kataoka

A simple and sensitive method for the analysis of oxytocin was developed using automated on-line in-tube solid-phase microextraction (SPME) coupled with liquid chromatography-tandem mass spectrometry (LC–MS/MS). Oxytocin was separated within 3 min on a Zorbax Eclipse XDB-C8 column, with water/methanol (10/90, v/v) as the mobile phase at a flow rate of 0.2 mL min−1. Electrospray ionization conditions in the positive ion mode were optimized for MS/MS detection by multiple reaction monitoring. The optimum in-tube SPME conditions were 20 draw/eject cycles of 40 µL sample at a flow rate of 250 µL min−1 using a Supel-Q PLOT capillary column as an extraction device. The extracted oxytocin was easily desorbed from the capillary by passage of the mobile phase, and no carryover was observed. The calibration curves for oxytocin were linear (r = 0.9981) in the range of 0−5.0 ng mL−1, and the relative standard deviations at each point were below 14.7% (n = 3). The limit of detection of this method was 4.0 pg mL−1, and its sensitivity was 58-fold higher than that of the direct injection method. This method was applied successfully to the analysis of oxytocin in saliva samples without any other interference peaks.

Chromatography doi: 10.3390/chromatography2030293

Authors: Sybille Merkle Kim Kleeberg Jan Fritsche

Solid-phase microextraction (SPME) is a simple, sensitive, rapid and solvent-free technique for the extraction of analytes from gaseous, liquid and solid samples and takes a leading position among microextraction methods. Application of SPME in sample preparation has been increasing continuously over the last decade. It is most often used as an automatized fiber injection system coupled to chromatographic separation modules for the extraction of volatile and semivolatile organic compounds and also allows for the trace analysis of compounds in complex matrices. Since SPME was first introduced in the early 1990s, several modifications have been made to adapt the procedure to specific application requirements. More robust fiber assemblies and coatings with higher extraction efficiencies, selectivity and stability have been commercialized. Automation and on-line coupling to analytical instruments have been achieved in many applications and new derivatization strategies as well as improved calibration procedures have been developed to overcome existing limitations regarding quantitation. Furthermore, devices using tubes, needles or tips for extraction instead of a fiber have been designed. In the field of food analysis, SPME has been most often applied to fruit/vegetables, fats/oils, wine, meat products, dairy and beverages whereas environmental applications focus on the analysis of air, water, soil and sediment samples.

Chromatography doi: 10.3390/chromatography2020277

Authors: Trine Andersen Federico Cozzi Henrik Simonsen

HS-SPME-GC-MS has been suggested as a fast and robust analytical platform for the product characterization of sesquiterpene synthases. The choice of fiber and injection temperature can have a significant effect on the observed product profile, due to the chemical rearrangements that can occur on the fiber material. Here we present a systematic study on the effects of fiber choice and injection port temperature on the observed sesquiterpenoid profile of four sesquiterpene synthases expressed in Nicotiana benthamiana. We found that the absorbent material PDMS was much less likely to support acid-induced rearrangement of sesquiterpenoids when compared to the adsorbent materials PDMS/DVB, PDMS/CAR, and PDMS/CAR/DVB. Furthermore, utilizing an injection port temperature at 160 °C almost eliminated the inherent thermal instability of germacrene sesquiterpenoids. Thus, for fast screening of sesquiterpene synthases, the results suggest that PDMS fibers and an injection temperature of 160 °C provide a fast and reproducible HS-SPME GC-MS method when using H2 as carrier gas.

Chromatography doi: 10.3390/chromatography2020265

Authors: Lingshuang Cai Jacek Koziel Matthew O'Neal

Insects trigger plants to release volatile compounds that mediate the interaction with both pest and beneficial insects. Soybean aphids (Aphis glycines) induces soybean (Glycine max) leaves to produce volatiles that attract predators of the aphid. In this research, we describe the use of solid-phase microextraction (SPME) for extraction of volatiles from A. glycines-infested plant. Objectives were to (1) determine if SPME can be used to collect soybean plant volatiles and to (2) use headspace SPME-GC-MS approach to screen compounds associated with A. glycines-infested soybeans, grown in the laboratory and in the field, to identify previously known and potentially novel chemical markers of infestation. A total of 62 plant volatiles were identified, representing 10 chemical classes. 39 compounds had not been found in previous studies of soybean volatile emissions. 3-hexen-1-ol, dimethyl nonatriene, indole, caryophyllene, benzaldehyde, linalool, methyl salicylate (MeSA), benzene ethanol, and farnesene were considered herbivore-induced plant volatiles (HIPVs). For reproductive field-grown soybeans, three compounds were emitted in greater abundance from leaves infested with A. glycines, cis-3-hexen-1-ol acetate, MeSA and farnesene. In summary, SPME can detect the emission of HIPVs from plants infested with insect herbivores.

Chromatography doi: 10.3390/chromatography2020253

Authors: Hanzhuo Fu Frank Cid Nat Dworkin James Cocores Gloria Shore

Kratom is a tree planted in Southeast Asia, including Thailand, Malaysia, Myanmar (Burma) and elsewhere in the region. A long history of usage and abuse of kratom has led to the classification of kratom as a controlled substance in its native Thailand and other Southeast Asian countries. However, kratom is not controlled in the United States, and the wide availability of kratom on the Internet and in the streets has led to its emergence as an herbal drug of misuse. With the increasing popularity of kratom, efficient protocols are needed to detect kratom use. In this study, a rapid method for the analysis of kratom compounds, mitragynine and 7-hydroxymitragynine, in human urine has been developed and validated using high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS). The chromatographic system employed a 2.6-μm 100 mm × 2.1 mm phenyl-hexyl analytical column and gradient elution with a 0.4-mL/min flow rate of water and acetonitrile as mobile phases. A triple quadrupole mass spectrometer was used as the detector for data acquisition. The analyst was the quantification software. The established method demonstrated linearity of >0.99 for both analytes, and low detection limits were obtained down to 0.002581 ng/mL for mitragynine and 0.06910 ng/mL for 7-hydroxymitragynine. The validated method has been utilized for clinical analysis of urine for the purpose of mitragynine and 7-hydroxymitragynine detection.

Chromatography doi: 10.3390/chromatography2020239

Authors: Jacek Giersz Krzysztof Jankowski Monika Truskolaska

A method for conducting fast and efficient gas chromatography based on short multicapillaries in straight alignment combined with atomic emission detection was developed for field analysis. The strategy enables for speciation analysis of organometallic compounds. The analytes are simultaneously ethylated and preconcentrated on a solid phase microextraction (SPME) fiber placed in the headspace over the sample for 25 min. The ethylated species are then completely separated and selectively quantified within 25 s under isothermal conditions. A new miniaturized speciation analyzer has been constructed and evaluated. The system consists of a GC injection port and a lab-made miniaturized GC unit directly coupled with miniaturized plasma excitation source. The emitted light is transferred via optical fiber and registered with a miniaturized charged coupled device (CCD) based spectrometer. Working parameters for multicapillary column gas chromatography with atomic emission detector, including carrier gas flow rate, desorption temperature, and GC column temperature, were optimized to achieve good separation of analytes. Basic investigations of the fundamental properties of 5 cm-long multicapillary column, to evaluate its potential and limitations as a rapid separation unit, are presented. The adaptation of the technique for use with a SPME system and with a multichannel element-selective plasma-emission detector is highlighted.

Chromatography doi: 10.3390/chromatography2020225

Authors: Irena Choma Wioleta Jesionek

The richness of bioactive compounds in plant materials encourages continuous development of separation methods and bioassays for their isolation and identification. Thin-layer chromatography-direct bioautography links separation on the adsorbent layer with biological tests performed directly on it. Therefore, the method is very convenient for searching plant constituents with biological activity, such as antibiotics. Test bacteria grow directly on a plate surface excluding places where antibacterials are located. They can be detected with reagents converted by living bacteria. TLC-DB is a high throughput method enabling analyses of many samples in parallel and the comparison of their activity. Both screening and semi-quantitative analysis is possible. The targeted compounds can be identified using spectroscopic methods, mostly mass spectrometry, that can be performed directly on a TLC plate. This paper discusses all above mentioned aspects of TLC-DB, illustrating them with literature, schemes and our own results.

Chromatography doi: 10.3390/chromatography2020213

Authors: Pierre-Hugues Stefanuto Katelynn Perrault Jean-François Focant Shari Forbes

Security control is becoming a major global issue in strategic locations, such as airports, official buildings, and transit stations. The agencies responsible for public security need powerful and sensitive tools to detect warfare agents and explosives. Volatile signature detection is one of the fastest and easiest ways to achieve this task. However, explosive chemicals have low volatility making their detection challenging. In this research, we developed and evaluated fast chromatographic methods to improve the characterization of volatile signatures from explosives samples. The headspace of explosives was sampled with solid phase micro-extraction fiber (SPME). Following this step, classical gas chromatography (GC) and comprehensive two-dimensional GC (GC×GC) were used for analysis. A fast GC approach allows the elution temperature of each analyte to be decreased, resulting in decreased thermal degradation of sensitive compounds (e.g., nitro explosives). Using fast GC×GC, the limit of detection is further decreased based on the cryo-focusing effect of the modulator. Sampling of explosives and chromatographic separation were optimized, and the methods then applied to commercial explosives samples. Implementation of fast GC methods will be valuable in the future for defense and security forensics applications.

Chromatography doi: 10.3390/chromatography2020195

Authors: Vignesh Rajamanickam Christoph Herwig Oliver Spadiut

Monolithic columns are a special type of chromatography column, which can be used for the purification of different biomolecules. They have become popular due to their high mass transfer properties and short purification times. Several articles have already discussed monolith manufacturing, as well as monolith characteristics. In contrast, this review focuses on the applied aspect of monoliths and discusses the most relevant biomolecules that can be successfully purified by them. We describe success stories for viruses, nucleic acids and proteins and compare them to conventional purification methods. Furthermore, the advantages of monolithic columns over particle-based resins, as well as the limitations of monoliths are discussed. With a compilation of commercially available monolithic columns, this review aims at serving as a ‘yellow pages’ for bioprocess engineers who face the challenge of purifying a certain biomolecule using monoliths.

Chromatography doi: 10.3390/chromatography2020188

Authors: Dolcie DeGrandchamp Sage Hartlaub Bernard Fried Joseph Sherma

The effects of 5, 25, and 40 Echinostoma caproni miracidia on the sugar content of young adult and mature adult Biomphalaria glabrata were studied using high performance thin layer chromatography (HPTLC)-densitometry. Analysis was done on the snail’s digestive gland gonad complex (DGG) at two and four weeks postmiracidial exposure. The sugars were extracted from the DGG using 70% ethanol and analyzed on silica gel HPTLC plates with a preadsorbent zone using 1-butanol-glacial acetic acid-diethyl ether-deionized water (27:18:5:3) mobile phase. The separated bands were then detected using alpha-naphthol-sulfuric reagent and quantified by densitometry at 515 nm. Significant differences were found in the maltose content between two and four weeks post exposure for both age groups. Additionally, significantly lower maltose and glucose levels were observed in the high exposure groups of both ages.

Chromatography doi: 10.3390/chromatography2020167

Authors: Andrés Domínguez Carmen Jarne Vicente Cebolla Javier Galbán María Savirón Jesús Orduna Luis Membrado María-Pilar Lapieza Elena Romero Isabel Sanz Vicente Susana De Marcos Rosa Garriga

Hyphenated HPTLC has been used to analyze several neutral sphingolipids acting as lysosomal storage disease (LSD) biomarkers. Automated multiple development (AMD) provides separation of lipid peaks, which are detected and quantified using fluorescence detection by intensity changes (FDIC) after primuline post-impregnation. A final online transfer to a mass spectrometer by means of an elution-based interface allows their identification using electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI).Given that the increases in fluorescent emission detected by FDIC are produced by non-specific, electrostatic interactions between the primuline and hydrocarbon chains in the ceramide backbones of sphingolipids, it is a non-destructive detection technique, allowing the precise location and transfer of biomarker peaks to a mass spectrometer using an elution interface. By using primuline as a fluorophore, the technique is also compatible with ESI-APCI and does not interfere with the MS of sphingolipids. APCI provides useful and complementary structural information to the ESI for sphingolipid identification. Moreover, FDIC emission can be used for quantitative purposes. Results include the determination of sphingomyelin (SM) in human-plasma samples (RSD < 6%) by means of a standard addition method with non-linear calibration, and the identification of globotriaosylceramide (Gb3) in the plasma of a Fabry patient. Only one HPTLC plate is needed to perform the analysis.

Chromatography doi: 10.3390/chromatography2020156

Authors: Štěpánka Trachtová Hana Zapletalová Alena Španová Daniel Horák Hana Kolářová Bohuslav Rittich

Using magnetic particles as a solid-phase extraction system is the most frequently used micro-technique for DNA isolation. Particles with a complete covering of magnetic cores by a polymer are hence preferred. Quantitative polymerase chain reaction (qPCR) was used for the evaluation of the polymer coating efficiency of hydrophilic magnetic poly(2-hydroxyethyl methacrylate-co-glycidyl methacrylate) (P(HEMA-co-GMA)) and poly(glycidyl methacrylate) (PGMA) microspheres with/without carboxyl groups. The inhibition effect of magnetic microspheres was identified by the shift in Cq values (ΔCq) after the addition of different amounts of microspheres to PCR mixtures. With the increase of microsphere concentrations, the shift in Cq values to higher values was usually observed. P(HEMA-co-GMA) microspheres containing carboxyl groups extinguished the fluorescence at concentrations over 2 mg mL−1 in a PCR mixture without any influence on the synthesis of PCR products. No PCR products (inhibition of DNA amplification) were detected in the presence of more than 0.8 mg mL−1 in the PCR mixture of PGMA microspheres. Atomic force microscopy (AFM) was used for the determination of the surface morphology of the microspheres. The microspheres were spherical, and their surface was non-porous.

Chromatography doi: 10.3390/chromatography2020141

Authors: Jude Abia

Information on the surface properties of three novel chemically bonded phase packing materials for High performance liquid chromatography (HPLC) were obtained using spectra obtained by solid state cross-polarization (CP) magic-angle spinning (MAS) nuclear magnetic resonance (NMR) spectroscopic experiments for the 29Si, and 13C nuclei. These packing materials were: Cogent bidentate C18 bonded to type-C silica, hybrid packing materials XTerra MS C18, and XBridge Prep. C18. The spectra obtained using cross-polarization magic angle spinning (CP-MAS) on the Cogent bidentate C18 bonded to type-C silica show the surface to be densely populated with hydride groups (Si-H), with a relative surface coverage exceeding 80%. The hybrid packing materials XTerra and XBridge gave spectra that reveal the silicon atoms to be bonded to organic moieties embedded in the molecular structure of these materials with over 90% of the alkyl silicon atoms found within the completely condensed silicon environments. The hydrolytic stability of these materials were investigated in acidic aqueous solutions at pHs of 7.0 and 3.0, and it was found that while the samples of XTerra and XBridge were not affected by hydrolysis at this pH range, the sample of Cogent lost a significant proportion of its Si-H groups after five days of treatment in acidic aqueous solution.

Chromatography doi: 10.3390/chromatography2010125

Authors: Sue-Siang Teh Gertrud Morlock

Hemp, flax and canola seed cakes are byproducts of the plant oil extraction industry that have not received much attention in terms of their potential use for human food instead of animal feed. Thus, the bioactivity profiling of these oilseed cakes is of interest. For their effect-directed analysis, planar chromatography was combined with several (bio)assays, namely 2,2-diphenyl-1-picrylhydrazyl scavenging, acetylcholine esterase inhibition, planar yeast estrogen screen, antimicrobial Bacillus subtilis and Aliivibrio fischeri assays. The streamlined high-performance thin-layer chromatography (HPTLC)-bioassay method allowed the discovery of previously unknown bioactive compounds present in these oilseed cake extracts. In contrast to target analysis, the direct link to the effective compounds allowed comprehensive information with regard to selected effects. HPTLC-electrospray ionization-mass spectrometry via the elution-head based TLC-MS Interface was used for a first characterization of the unknown effective compounds. The demonstrated bioactivity profiling on the feed/food intake side may guide the isolation of active compounds for production of functional food or for justified motivation of functional feed/food supplements.

Chromatography doi: 10.3390/chromatography2010118

Authors: Bernard Fried Aditya Reddy

This review examines the recent high performance thin layer chromatography (HPTLC) literature on the effects of biotic and abiotic factors on certain analytes in the medically important freshwater snail, Biomphalaria glabrata. The analytes studied were lipids, lipophilic pigments, amino acids, and carbohydrates. As determined by HPTLC, various factors, such as larval parasitism, estivation, temperature changes, and others, alter the metabolism of the snail and cause significant changes in the chemical contents of the analytes under study.

Chromatography doi: 10.3390/chromatography2010096

Authors: Zsuzsanna Kuklenyik Michael Gardner Bryan Parks David Schieltz Jon Rees Lisa McWilliams Yulanda Williamson James Pirkle John Barr

In this report we demonstrate a practical multivariate design of experiment (DoE) approach for asymmetric flow field-flow fractionation (AF4) method optimization using separation of lipoprotein subclasses as an example. First, with the aid of commercially available software, we built a full factorial screening design where the theoretical outcomes were calculated by applying established formulas that govern AF4 channel performance for a 5–35 nm particle size range of interest for lipid particles. Second, using the desirable ranges of instrumental parameters established from theoretical optimization, we performed fractional factorial DoE for AF4 separation of pure albumin and ferritin with UV detection to narrow the range of instrumental parameters and allow optimum size resolution while minimizing losses from membrane immobilization. Third, the optimal range of conditions were tested using response surface DoE for sub-fractionation of high and low density lipoproteins (HDL and LDL) in human serum, where the recovery of the analytes were monitored by fraction collection and isotope-dilution LC-MS/MS analysis of each individual fraction for cholesterol and apolipoproteins (ApoA-1 and ApoB-100). Our results show that DoE is an effective tool in combining AF4 theoretical knowledge and experimental data in finding the most optimal set of AF4 instrumental parameters for quantitative coupling with LC-MS/MS measurements.

Chromatography doi: 10.3390/chromatography2010079

Authors: Mónica Díaz-Bao Rocío Barreiro José Miranda Alberto Cepeda Patricia Regal

Monolithic columns are gaining interest as excellent substitutes to conventional particle-packed columns. These columns show higher permeability and lower flow resistance than conventional liquid chromatography columns, providing high-throughput performance, resolution and separation in short run times. Monoliths possess also great potential for the clean-up and preparation of complex mixtures. In situ polymerization inside appropriate supports allows the development of several microextraction formats, such as in-tube solid-phase and pipette tip-based extractions. These techniques using porous monoliths offer several advantages, including miniaturization and on-line coupling with analytical instruments. Additionally, monoliths are ideal support media for imprinting template-specific sites, resulting in the so-called molecularly-imprinted monoliths, with ultra-high selectivity. In this review, time-saving LC columns and preparative applications applied to the analysis of residues and contaminants in food in 2010–2014 are described, focusing on recent improvements in design and with emphasis in automated on-line systems and innovative materials and formats.

Chromatography doi: 10.3390/chromatography2010066

Authors: Mengliang Zhang Peter Harrington

A method for the determination of trichloroethylene (TCE) in water using portable gas chromatography/mass spectrometry (GC/MS) was developed. A novel sample preparation method, liquid–liquid microextraction assisted solid phase microextraction (LLME–SPME), is introduced. In this method, 20 µL of hexane was added to 10 mL of TCE contaminated aqueous samples to assist headspace SPME. The extraction efficiency of SPME was significantly improved with the addition of minute amounts of organic solvents (i.e., 20 µL hexane). The absolute recoveries of TCE at different concentrations were increased from 11%–17% for the samples extracted by SPME to 29%–41% for the samples extracted by LLME–SPME. The method was demonstrated to be linear from 10 to 1000 ng mL−1 for TCE in water. The improvements on extraction efficiencies were also observed for toluene and 1, 2, 4-trichlorobenzene in water by using LLME–SPME method. The LLME–SPME method was optimized by using response surface modeling (RSM).

Chromatography doi: 10.3390/chromatography2010020

Authors: Allan Alla Keith Stine

Monolithic column materials offer great advantages as chromatographic media in bioseparations and as solid-supports in biocatalysis. These single-piece porous materials have an interconnected ligament structure that limits the void volume inside the column, thus increasing the efficiency without sacrificing the permeability. The preparation of monolithic materials is easy, reproducible and has available a wide range of chemistries to utilize. Complex, heterogeneous and isobaric glycan structures require preparation methods that may include glycan release, separation and enrichment prior to a comprehensive and site-specific glycosylation analysis. Monolithic column materials aid that demand, as shown by the results reported by the research works presented in this review. These works include selective capture of glycans and glycoproteins via their interactions with lectins, boronic acids, hydrophobic, and hydrophilic/polar functional groups on monolith surfaces. It also includes immobilization of enzymes trypsin and PNGase F on monoliths to digest and deglycosylate glycoproteins and glycopeptides, respectively. The use of monolithic capillary columns for glycan separations through nano-liquid chromatography (nano-LC) and capillary electrochromatography (CEC) and coupling these columns to MS instruments to create multidimensional systems show the potential in the development of miniaturized, high-throughput and automated systems of glycan separation and analysis.

Chromatography doi: 10.3390/chromatography2010019

Authors: Chromatography Editorial Office

The editors of Chromatography would like to express their sincere gratitude to the following reviewers for assessing manuscripts in 2014:[...]

Chromatography doi: 10.3390/chromatography2010001

Authors: Wenda Zhu Jacek Koziel Lingshuang Cai H. Duygu Özsoy J. Van Leeuwen

Fruit leathers (FLs) production produces some not-to-specification material, which contains valuable ingredients like fruit pulp, sugars and acidulates. Recovery of FL for product recycling requires decolorization. In earlier research, we proved the efficiency of an ozone-based decolorization process; however, it produces carbonyls as major byproducts, which could be of concern. A headspace solid-phase microextraction with on-fiber derivatization followed by gas chromatography-mass spectrometry was developed for 10 carbonyls analysis in ozonated FL solution/suspension. Effects of dopant concentration, derivatization temperature and time were studied. The adapted method was used to analyze ozonated FL solution/suspension samples. Dopant concentration and derivatization temperature were optimized to 17 mg/mL and 60 °C, respectively. Competitive extraction was studied, and 5 s extraction time was used to avoid non-linear derivatization of 2-furfural. The detection limits (LODs) for target carbonyls ranged from 0.016 and 0.030 µg/L. A much lower LOD (0.016 ppb) for 2-furfural was achieved compared with 6 and 35 ppb in previous studies. Analysis results confirmed the robustness of the adapted method for quantification of carbonyls in recycled process water treated with ozone-based decolorization. Ethanal, hexanal, 2-furfural, and benzaldehyde were identified as byproducts of known toxicity but all found below levels for concern.

Chromatography doi: 10.3390/chromatography1040211

Authors: Swee Tan Jean Yong Liya Ge

Capillary electrophoresis (CE) coupled with mass spectrometry (MS) or tandem mass spectrometry (MS/MS) is reported as an alternative and potentially useful method for the simultaneous analysis of various classes of phytohormones with diversified structures, including indole-3-acetic acid (IAA), indole-3-butyric acid (IBA), abscisic acid (ABA), gibberellic acid (GA), zeatin (Z), N6-benzyladenine (BA), α-naphthaleneacetic acid (NAA) and 2,4-dichlorophenoxyacetic acid (2,4-D). The key to the CE-MS/MS analysis was based on electroosmotic flow reversal using a cationic polymer-coated capillary. Under optimum conditions, a baseline separation of eight phytohormones was accomplished within 30 min using 60 mM ammonium formate/formic acid buffer of pH 3.8 with −20 kV as the separation voltage. The accessibility of MS/MS together with the characterization by migration properties obtained by CE allows for the development of CE-MS/MS as an emerging potential method for the analysis of different classes of phytohormones in a single run. The utility of the CE-MS/MS method was demonstrated by the comprehensive screening of phytohormones in coconut (Cocos nucifera L.) water after pre-concentration and purification through solid-phase extraction (SPE) cartridge. IAA, ABA, GA and Z were detected and quantified in the purified coconut water extract sample.

Chromatography doi: 10.3390/chromatography1040194

Authors: Elsa Cabo-Calvet Casandra Ortiz-Bolsico Juan Baeza-Baeza María García-Alvarez-Coque

The effect of the modifier concentration and flow rate on the chromatographic performance of a second generation Chromolith® RP-18e column, under isocratic and gradient elution with acetonitrile-water mixtures, was examined using four sulphonamides as probe compounds. The acetonitrile concentration was varied between 5 and 55% (v/v), and the flow rate between 0.1 and 5.0 mL/min, keeping the other factors constant. The changes in both retention and peak profile were modelled, and used to build simple plots, where the logarithm of the retention factor was represented against the modifier concentration (in gradient elution, against the initial modifier concentration), and the half-widths or widths against the retention time (in gradient elution, against the time at the column outlet). A particular plot was needed for describing the retention of each sulphonamide, but due to the similar interaction kinetics, a unique plot described the changes in the half-widths for all four sulphonamides. The changes in retention with the flow showed that allegedly in the second generation Chromolith, the column deformation observed for the first generation Chromolith, with the applied pressure at increasing flow, is decreased.

Chromatography doi: 10.3390/chromatography1040176

Authors: Eli Moore H. Harvey Jessica Faux David Goodlett Brook Nunn

Proteins are the largest defined molecular component of marine organic nitrogen, and hydrolysable amino acids, the building blocks of proteins, are important components of particulate nitrogen in marine sediments. In oceanic systems, the largest contributors are phytoplankton proteins, which have been tracked from newly produced bloom material through the water column to surface sediments in the Bering Sea, but it is not known if proteins buried deeper in sediment systems can be identified with confidence. Electrophoretic gel protein extraction methods followed by proteomic mass spectrometry and database searching were used as the methodology to identify buried phytoplankton proteins in sediments from the 8–10 cm section of a Bering Sea sediment core. More peptides and proteins were identified using an SDS-PAGE tube gel than a standard 1D flat gel or digesting the sediment directly with trypsin. The majority of proteins identified correlated to the marine diatom, Thalassiosira pseudonana, rather than bacterial protein sequences, indicating an algal source not only dominates the input, but also the preserved protein fraction. Abundant RuBisCO and fucoxanthin chlorophyll a/c binding proteins were identified, supporting algal sources of these proteins and reinforcing the proposed mechanisms that might protect proteins for long time periods. Some preserved peptides were identified in unexpected gel molecular weight ranges, indicating that some structural changes or charge alteration influenced the mobility of these products during electrophoresis isolation. Identifying buried photosystem proteins suggests that algal particulate matter is a significant fraction of the preserved organic carbon and nitrogen pools in marine sediments.

Chromatography doi: 10.3390/chromatography1040159

Authors: Rod Chalk Georgina Berridge Leela Shrestha Claire Strain-Damerell Pravin Mahajan Wyatt Yue Opher Gileadi Nicola Burgess-Brown

Mass spectrometry (MS) remains under-utilized for the analysis of expressed proteins because it is inaccessible to the non-specialist, and sample-turnaround from service labs is slow. Here, we describe 3.5 min Liquid-Chromatography (LC)-MS and 16 min LC-MSMS methods which are tailored to validation and characterization of recombinant proteins in a high throughput structural biology pipeline. We illustrate the type and scope of MS data typically obtained from a 96-well expression and purification test for both soluble and integral membrane proteins (IMPs), and describe their utility in the selection of constructs for scale-up structural work, leading to cost and efficiency savings. We propose that value of MS data lies in how quickly it becomes available and that this can fundamentally change the way in which it is used.

Chromatography doi: 10.3390/chromatography1030141

Authors: Luís Dias Cédric Sequeira Ana Veloso Jorge Morais Mara Sousa António Peres

In this work, the main organic acids (citric, malic and ascorbic acids) and sugars (glucose, fructose and sucrose) present in commercial fruit beverages (fruit carbonated soft-drinks, fruit nectars and fruit juices) were determined. A novel size exclusion high performance liquid chromatography isocratic green method, with ultraviolet and refractive index detectors coupled in series, was developed. This methodology enabled the simultaneous quantification of sugars and organic acids without any sample pre-treatment, even when peak interferences occurred. The method was in-house validated, showing a good linearity (R > 0.999), adequate detection and quantification limits (20 and 280 mg L−1, respectively), satisfactory instrumental and method precisions (relative standard deviations lower than 6%) and acceptable method accuracy (relative error lower than 5%). Sugars and organic acids profiles were used to calculate dose-over-threshold values, aiming to evaluate their individual sensory impact on beverage global taste perception. The results demonstrated that sucrose, fructose, ascorbic acid, citric acid and malic acid have the greater individual sensory impact in the overall taste of a specific beverage. Furthermore, although organic acids were present in lower concentrations than sugars, their taste influence was significant and, in some cases, higher than the sugars’ contribution towards the global sensory perception.

Chromatography doi: 10.3390/chromatography1030120

Authors: Katelynn Perrault Barbara Stuart Shari Forbes

Odour profiling of decomposed remains is important for understanding the mechanisms that cadaver dogs and forensically-relevant insects use to locate decomposed remains. The decomposition odour profile is complex and has been documented in outdoor terrestrial environments. The purpose of this study was to perform longitudinal analysis of the volatile organic compound (VOC) profile in soils associated with decomposed remains across all stages of decomposition. Two VOC collection techniques (sorbent tubes and solid phase microextraction) were used to collect a wider analyte range and to investigate differences in collection techniques. Pig carcasses were placed in an outdoor research facility in Australia to model the decomposition process and VOCs were collected intermittently over two months. VOCs of interest were identified over the duration of the trial, showing distinct trends in compound evolution and disappearance. The collection techniques were complementary, representing different subsets of VOCs from the overall profile. Sorbent tubes collected more decomposition-specific VOCs and these compounds were more effective at characterising the matrix over an extended period. Using both collection techniques improves the likelihood of identifying the complete VOC profile of decomposition odour. Such information is important for the search and recovery of victim remains in various stages of decomposition.

Chromatography doi: 10.3390/chromatography1030108

Authors: Sean Harshman Claude Grigsby Darrin Ott

Exhaled breath condensate (EBC) has been established as a potential source of respiratory biomarkers. Compared to the numerous small molecules identified, the protein content of EBC has remained relatively unstudied due to the methodological and technical difficulties surrounding EBC analysis. In this review, we discuss the proteins identified in EBC, by mass spectrometry, focusing on the significance of those proteins identified. We will also review the limitations surrounding mass spectral EBC protein analysis emphasizing recommendations to enhance EBC protein identifications by mass spectrometry. Finally, we will provide insight into the future directions of the EBC proteomics field.

Chromatography doi: 10.3390/chromatography1030096

Authors: Akio Makishima

A new simple and quick method has been established for separation of Cu from solutions using an extraction chromatographic resin utilizing Aliquat® 336 (commercially available as TEVA™ resin) and Cu(I). This method involves the use of a one milliliter column containing 0.33 mL TEVA™ resin on 0.67 mL Amberchrom® CG-71C acrylic resin. Copper was adsorbed on the column by forming Cu(I) with 0.15% ascorbic acid in 0.05 mol·L−1 HBr, while other major elements except Zn showed no adsorption. After removal of the major elements (Na, Mg, Al, P, K, Ca, Cr, Mn, Fe, Co and Ni), Cu was recovered using 2 mol·L−1 HNO3. The recovery yield and total blank were 102% ± 2% and 0.25 ng, respectively. To evaluate the separation method, Cu isotope ratios were determined by a standard-sample-standard bracketing method using multicollector inductively coupled plasma-mass spectrometry (ICP-MS), with a repeatability of 0.04‰ and 0.25‰ (SD), for the standard solution and the solutions from low S (<0.1% S) silicate standards, respectively.

Chromatography doi: 10.3390/chromatography1020082

Authors: Jeffrey Roth Cody Peer Baskar Mannargudi Helen Swaisland Jung-Min Lee Elise Kohn William Figg

Olaparib (AZD2281) is an orally active PARP-1 inhibitor, primarily effective against cancers with BRCA1/2 mutations. It is currently in Phase III development and has previously been investigated in numerous clinical trials, both as a single agent and in combination with chemotherapy. Despite this widespread testing, there is only one published method that provides assay details and stability studies for olaparib alone. A more sensitive uHPLC-MS/MS method for the quantification of olaparib in human plasma was developed, increasing the range of quantification at both ends (0.5–50,000 ng/mL) compared to previously published methods (10–5,000 ng/mL). The wider range encompasses CMAX levels produced by typical olaparib doses and permits better pharmacokinetic modeling of olaparib elimination. This assay also utilizes a shorter analytical runtime, allowing for more rapid quantification and reduced use of reagents. A liquid-liquid extraction was followed by chromatographic separation on a Waters UPLC® BEH C18 column (2.1 × 50 mm, 1.7 µm) and mass spectrometric detection. The mass transitions m/z 435.4→281.1 and m/z 443.2→281.1 were used for olaparib and the internal standard [2H8]-olaparib, respectively. The assay proved to be accurate (<9% deviation) and precise (CV < 11%). Stability studies showed that olaparib is stable at room temperature for 24 h. in whole blood, at 4 °C for 24 h post-extraction, at −80 °C in plasma for at least 19 months, and through three freeze-thaw cycles. This method proved to be robust for measuring olaparib levels in clinical samples from a Phase I trial.

Chromatography doi: 10.3390/chromatography1020075

Authors: Naoto Furusawa

This paper describes a reserved-phase high-performance liquid chromatographic (HPLC) method for detecting malachite green (MG) and leuco-malachite green (LMG) using an isocratic toxic organic solvent/reagent-free mobile phase. Chromatographic separations were performed an Inertsil® WP300 C4 with 0.02 mol/L octane sulfonic acid–ethanol mobile phase and a photodiode-array detector. The total run time was <5 min. The system suitability was well within the international acceptance criteria. A harmless method for simultaneously detecting MG and LMG was developed and may be further applied to the quantification in foods.

Chromatography doi: 10.3390/chromatography1020065

Authors: Dana Horgen Charles Garner

The enantioseparation of a series of six azulene-centered 1,5-diol enantiomers was studied employing two cellulose-based chiral stationary phases under normal phase conditions (isopropanol/hexanes). The separations were generally quite good on Chiralcel-OD-H, with α values ranging from 1.2 to 8.4 (average 4.0) and resolution values of 0.4–8.3 (average 4.7). Only one of the six enantiomer pairs was not well resolved, but was well separated on Lux cellulose 2 (α 1.4, Rs 8.7). It was observed that the enantioseparations of the RS/SR diastereomers (ave α = 7.8, Rs = 8.2) were dramatically better than that of the corresponding RR/SS diastereomers (ave α = 2.1, Rs = 3.0) on Chiralcel-OD-H. The better-resolved diastereomer pairs correspond to the more strongly retained diastereomers on silica gel. The enantiomers of two benzene 1,5-diols were much more poorly separated on both stationary phases, suggesting that the unusual polarity of the azulene ring enhances critical interactions with these phases.

Chromatography doi: 10.3390/chromatography1020055

Authors: Sonia Gregory Henryk Mach

The state-of-the-art instruments for the determination of viscosity of liquids typically require a significant amount of sample, and have relatively low throughput due to manual and sequential measurements. In this study, it was demonstrated that the pressure generated by the flow of viscous fluids through a capillary could be precisely measured employing high-pressure liquid chromatography systems (HPLC) using glycerol solutions of moderate viscosity as a mobile phase, and correlated to the dynamic (absolute) viscosity. The parameters allowing calculation of the viscosity of glycerol calibration standards as a function of temperature were established. The measurements were made with volumes as small as 10 μL, and the use of an autosampler permitted unattended analysis of a large number samples. The method appears to be particularly well suited for the development of viscous formulations of therapeutic, protein-based macromolecules, where the amount sample is typically limited and relatively wide ranges of conditions are considered in the optimization process. The utility of the methods was illustrated by application to the development of concentrated inactivated virus vaccines.

Chromatography doi: 10.3390/chromatography1010054

Authors: Frank Dorman

It is my great pleasure to serve as the first editor-in-chief for Chromatography, and welcome you all to the readership. This journal will enable us to communicate our findings in “open access” while maintaining the very high scientific requirements of the few other journals that are focused on the type of science that we all find so very interesting. It is safe to say that chromatographic separations are the single most heavily used techniques in the larger field of analytical chemistry, whether applied or fundamental. Without chromatography, we would struggle to answer many of the pressing scientific questions of our time, and without improvements and advancements we will still struggle to probe more deeply into the challenges of the future. Whether it is for better understanding and characterization of starting materials and products, qualitative determination of unknowns in important matrices, quantification of the materials that impact our lives, or the discovery of the emerging compounds of the future, chromatography is the science that ties our various research fields together. [...]

Chromatography doi: 10.3390/chromatography1010024

Authors: Sinéad Currivan Pavel Jandera

The vast cache of methods used in polymeric monolithic column modification is presented herein, with specific attention to post-polymerization modification reactions. The modification of polymeric monolithic columns is defined and can include the modification of pre-existing surface groups, the addition of polymeric chains or indeed the addition of structures such as nano-particles and nano-structures. The use of these modifications can result in the specific patterning of monoliths, useful in microfluidic device design or in the investigation of modification optimization.

Chromatography doi: 10.3390/chromatography1010009

Authors: Shokoufeh Ahmadi Douglas Craig Douglas Goltz

In this study, the separation parameters in micellar electrokinetic chromatography (MEKC)-laser-induced fluorescence (LIF) were optimized for the separation of red and yellow historical dyes in 20 mM borate buffer with 20 mM sodium dodecyl sulfate (SDS). Separation conditions were optimized by changing pH, organic modifier (methanol and acetonitrile) concentrations and applied voltage. The mixtures of dyes used in this study included four anthraquinone dyes (alizarin, purpurin, emodin and carmine) and six flavonoid dyes (luteolin, apigenin, kaempferol, quercetin, morin and myricetin). For this work, dyes were introduced electro-kinetically (10 kV for 5 s) into a 50 cm capillary (10 µm id) and separated using a running potential of 18, 20, 22 and 25 kV. Absolute limits of detection for most of these dyes was less than 1 pg. For dyes such as alizarin, improved detection limits were achieved at pH = 9.24; however dyes such as purpurin had significantly improved detection limits at pH = 8.0. The successful extraction and identification of a number of dyes in plants and textiles samples is also described.

Chromatography doi: 10.3390/chromatography1010001

Authors: Anja Krüger Markus Ganzera

Flavonoids are bioactive constituents in Oroxylum indicum seeds, an Asian traditional remedy used for the treatment of respiratory infections. In this study the first capillary electrophoretic method for their determination is presented. By using a 25 mM borax buffer at pH 9.2 containing 10 mM SDS as detergent, the determination of seven flavonoids was feasible in only 13 min. Method validation confirmed that the assay is in accordance with ICH requirements in respect to linearity, selectivity, sensitivity, accuracy and precision. Quantitative results revealed that baicalein-7-O-gentiobioside is the most abundant flavonoid in the drug (1.19 to 5.33%), followed by other baicalein derivatives (7-O-glucoside, 7-O-glucuronide). These observations were in good qualitative and quantitative agreement with LC-MS results.