Separations, Volume 6, Issue 3 (September 2019) – 12 articles

Mangiferin has been reported to exhibit anti-viral, anti-cancer, anti-diabetic, immunomodulatory and hepatoprotective properties. This study aimed to develop an HPLC method to isolate mangiferin from Salacia chinensis L. root; investigate the impact of solvents on yield; optimise the ultrasound-assisted extraction (UAE) technique; and compare mangiferin yield with continuously shaking extraction (CSE) and decoction techniques. The results showed that mangiferin, with a purity of over 88%, could be achieved by HPLC using a mixture of solvent A (water: acetonitrile: orthophosphoric acid, 96.8:3:0.2 (v/v/v)) and solvent B (acetonitrile). Solvent type significantly affected the extraction yield of mangiferin, and a mixture of acetone and water gave the highest extraction yield, as compared to other solvents or mixtures. UAE conditions, such as ultrasonic power, temperature, time and concentration of acetone significantly affected the extraction of mangiferin. Optimal UAE conditions were at an ultrasonic power of 250 W, temperature of 50 °C, acetone concentration of 40% and extraction time of 60 min. These optimal conditions could extract approximately 92 mg, whereas CSE and decoction only extracted 89.20 mg and 58.71 mg of mangiferin, respectively, from 1 g of S. chinensis root. Therefore, these UAE conditions are recommended for the extraction of mangiferin from S. chinensis root for further utilisation. Full article

In May 2019, the Food and Drug Administration (FDA) proposed a quality range (QR) method for the comparative analytical assessment in biosimilar studies. In this process, several reference product lots are necessary, selected from a wide period of manufacturing dates with different shelf lives, to calculate the total variability expressed as the standard deviation of reference product lots. This one depends on the between-lots variation and analytic method uncertainty (i.e., within-lots variation). During this time, the analytical method must be in control and stable but with an appropriate accuracy and precision. In such a situation, various control charts were used to fix the method requirements and detect small changes in the process. The results indicate that the method is indeed in control and stable, but does not meet the requirements of the Analytical Target Profile (ATP) approach, independently of the established uncertainty range. However, it does satisfy the traditional approach for an uncertainty range of ±2%. The application of this new QR approach shows that the selection of reference lots has an impact on the estimated standard deviation of the reference product, and consequently on the QR, penalizing good test products. The contribution of the analytic method error is known and in-control through the validation process. However, the between-lots variation requires a higher attention and control by the manufacturer. All these aspects were analyzed, using simulation and real-data from various bevacizumab lots. Full article

A large number of polar stationary phases with diverse chemistry have been developed for various applications in hydrophilic interaction chromatography (HILIC). However, column manufacturers employ different testing procedures to evaluate retention of the polar stationary phases. This renders the retention data impossible for comparison and makes it difficult for the users to select the right stationary phase based on retention. We have evaluated 25 polar stationary phases using cytosine and uracil as the model compounds in various mobile phase conditions. These stationary phases show a wide range of retention characteristics for the model compounds. The ranking of the stationary phases does not change drastically with the acetonitrile level in the mobile phase. Full article

Chemometric analysis of comprehensive two-dimensional chromatography coupled to time-of-flight mass spectrometry (GC×GC-TOFMS) data has been reported with various workflows, yet little effort has been devoted to evaluating the impacts of workflow variation on study conclusions. The report presented herein aims to investigate the effects of different pre-processing and variable selection strategies on the scores’ plot outputs from GC×GC-TOFMS data acquired from lavender and tea tree essential oils. Our results suggest that pre-processing, such as applying log transformation to the data set, can result in significant differentiation of sample clustering when compared to only mean centering. Additionally, exploring differences between analysis of variance, Fisher-ratio, and partial least squares-discriminant analysis feature selection resulted in little variation in scores plots. This work highlights the effects different chemometric workflows can have on results to help facilitate harmonization efforts. Full article

The presence of free silanols on alkyl-bonded reversed-phase stationary phases is responsible for broad and asymmetrical peaks when basic drugs are chromatographed with conventional octadecylsilane (C18) columns due to ionic interactions. In the last few years, ionic liquids (ILs) have attracted attention to reduce this undesirable silanol activity. ILs should be considered as dual modifiers (with a cationic and anionic character), which means that both cations and anions are able to adsorb on the stationary phase, creating a positively or negatively charged layer, depending on the relative adsorption. The accessibility of basic compounds to the silanols is prevented by both the IL cation and anion, improving the peak profiles. A comparative study of the performance of six imidazolium-based ILs, differing in their cation/anions, as modifiers of the chromatographic behavior of a group of ten β-adrenoceptor antagonists, is addressed. Mobile phases containing cationic amines (triethylamine and dimethyloctylamine) were used as a reference for the interpretation of the results. Using a mathematical model based on two chemical equilibria, the association constants between the solutes and modified stationary phase as well as those between solutes and the additive in the mobile phase were estimated. These values, together with the changes in retention and peak shape, were used to obtain conclusions about the retention mechanism, changes in the nature of the chromatographic system, and silanol suppression effect. Full article

Through the development of solid phase microextraction (SPME) technologies, thin film solid phase microextraction (TF-SPME) has been repeatedly validated as a novel sampling device well suited for various applications. These applications, encompassing a wide range of sampling methods such as onsite, in vivo and routine analysis, benefit greatly from the convenience and sensitivity TF-SPME offers. TF-SPME, having both an increased extraction phase volume and surface area to volume ratio compared to conventional microextraction techniques, allows high extraction rates and enhanced capacity, making it a convenient and ideal sampling tool for ultra-trace level analysis. This review provides a comprehensive discussion on the development of TF-SPME and the applications it has provided thus far. Emphasis is given on its application to thermal desorption, with method development and optimization for this desorption method discussed in detail. Moreover, a detailed outlook on the current progress of TF-SPME development and its future is also discussed with emphasis on its applications to environmental, food and fragrance analysis. Full article

This case study describes data analysis of a chromatogram distributed for the 2019 GC×GC Data Challenge for the Tenth Multidimensional Chromatography Workshop (Liege, Belgium). The chromatogram resulted from chemical analysis of a terpene-standards sample by comprehensive two-dimensional chromatography with mass spectrometry (GC×GC-MS). First, several aspects of the data quality are assessed, including detector saturation and oscillation, and operations to prepare the data for analyte detection and identification are described, including phase roll for modulation-cycle alignment and baseline correction to account for the non-zero detector baseline. Then, the case study presents operations for analyte detection with filtering, a new method to flag false detections, interactive review to confirm detected peaks, and ion-peaks detection to reveal peaks that are obscured by noise or coelution. Finally, the case study describes analyte identification including mass-spectral library search with a new method for optimizing spectra extraction, retention-index calibration from preliminary identifications, and expression-based identification checks. Processing of the first 40 min of data detected 144 analytes, 21 of which have at least one percent response, plus an additional 20 trace and/or coeluted analytes. Full article

The development of antibiotic resistance by microbials has long been acknowledged. The major challenge worldwide is to develop novel, natural, and potent antibiotics against the multidrug resistant bacteria. In this study, our aim was to develop the method for a highly sensitive instrument, ultra-high performance liquid chromatograph-high resolution mass spectrometer (UHPLC-HRMS), to evaluate the antibacterial property of a natural product. Aechmea magdalenae (Andre) Andre ex Baker, a plant belonging to the family Bromeliaceae, a native of Central America was used in this study. Based on the available literature, it was hypothesized that Aechmea magdalenae has antibacterial activity. In addition, the profiling done on A. magdalenae using gas chromatography-mass spectrometry (GC-MS) also revealed the presence of medicinally important chemical compounds, such as acetic acid. Minimum inhibitory concentration (MIC) of dried Aechmea plant extract was determined for the first time using 96-well plate assay, followed by determination of antibacterial potential using LC-MS. The reason being that other dried methanolic plant extracts, such as Vismia macrophylla, lined up for antibacterial testing have dark extracts, for which determining the antibacterial potential and reading the results with the naked eye would be challenging. To overcome the situation of dark plant extracts, a generalized novel LC-MS method was developed that was used for the plant A. magdalenae, and would be used further for other plants. A blue indicator called resazurin was added to the wells; resazurin, upon incubation with the living cells, got reduced to resorufin (which was pink), while it remained blue with bacterial growth inhibition. The mass difference created due to reduction of resazurin to resorufin was detected by using LTQ Orbitrap Discovery in positive ion mode to determine the antibacterial activity of the plant extract. The sample preparation for LC-MS assay included centrifugation of the samples taken from 96-well plate, followed by filtration of the supernatant, before exposing them to C-18 column. The results obtained from full scan LC-MS spectrum consistently demonstrated the presence of resorufin from wells with bacterial growth, and resazurin from wells with inhibition through peaks of relevant masses. Full article

In the countercurrent chromatography and centrifugal partition chromatography, separation method transfer and scale-up is often described as an easy and straightforward procedure. Separation methods are usually developed on lab scale columns and subsequently transferred using linear scale-up factors to semi-preparative or preparative columns of the same column design. However, the separation methods described in the literature have been developed on various columns of different design and size. This is accompanied by differences in the separation behavior of the columns and therefore makes separation method transfer difficult. In the current study, the separation performances of different columns were evaluated and compared. Linear correlations of stationary phase retention and column efficiency as a function of flow rate were found to be applicable for the calculation of separation resolution in the typical operating range of each column. In this context, a two-point short-cut approach for a fast column characterization is recommended. This allows a quick prediction of the separation method transferability between columns, which saves experimental time and effort. In the current study, the transferability between five different columns from lab scale countercurrent chromatography (CCC) (18 mL) to semi-preparative centrifugal partition chromatography (CPCs) (250 mL) with different cell numbers and design is investigated. Full article

The use of green extraction phases has gained much attention in different fields of study, including in sample preparation for the determination of organic compounds by chromatography techniques. Green extraction phases are considered as an alternative to conventional phases due to several advantages such as non-toxicity, biodegradability, low cost and ease of preparation. In addition, the use of greener extraction phases reinforces the environmentally-friendly features of microextraction techniques. Thus, this work presents a review about new materials that have been used in extraction phases applied to liquid and sorbent-based microextractions of organic compounds in different matrices. Full article

The study explores the complex volatile fraction of extra-virgin olive oil by combining high concentration-capacity headspace approaches with comprehensive two-dimensional gas chromatography, which is coupled with time of flight mass spectrometry. The static headspace techniques in this study are: (a) Solid-phase microextraction, with multi-polymer coating (SPME- Divinylbenzene/Carboxen/Polydimethylsiloxane), which is taken as the reference technique; (b) headspace sorptive extraction (HSSE) with either a single-material coating (polydimethylsiloxane—PDMS) or a dual-phase coating that combines PDMS/Carbopack and PDMS/EG (ethyleneglycol); (c) monolithic material sorptive extraction (MMSE), using octa-decyl silica combined with graphite carbon (ODS/CB); and dynamic headspace (d) with either PDMS foam, operating in partition mode, or Tenax TA™, operating in adsorption mode. The coverage of both targeted and untargeted 2D-peak-region features, which corresponds to detectable analytes, was examined, while concentration factors (CF) for a selection of informative analytes, including key-odorants and off-odors, and homolog-series relative ratios were calculated and the information capacity was discussed. The results highlighted the differences in concentration capacities, which were mainly caused by polymer-accumulation characteristics (sorptive/adsorptive materials) and its amount. The relative concentration capacity for homologues and potent odorants was also discussed, while headspace linearity and the relative distribution of analytes, as a function of different sampling amounts, was examined. This last point is of particular interest in quantitative studies where accurate data is needed to derive consistent conclusions. Full article

Metal-organic frameworks (MOFs) are porous hybrid materials composed of metal ions and organic linkers, characterized by their crystallinity and by the highest known surface areas. MOFs structures present accessible cages, tunnels and modifiable pores, together with adequate mechanical and thermal stability. Their outstanding properties have led to their recognition as revolutionary materials in recent years. Analytical chemistry has also benefited from the potential of MOF applications. MOFs succeed as sorbent materials in extraction and microextraction procedures, as sensors, and as stationary or pseudo-stationary phases in chromatographic systems. To date, around 100 different MOFs form part of those analytical applications. This review intends to give an overview on the use of MOFs in analytical chemistry in recent years (2017–2019) within the framework of green analytical chemistry requirements, with a particular emphasis on possible toxicity issues of neat MOFs and trends to ensure green approaches in their preparation. Full article