Rapid determination of parabens in seafood sauces by high‐performance liquid chromatography: A practical comparison of core–shell particles and sub‐2 μm fully porous particles

In this work, the chromatographic performance of superficially porous particles (Halo core–shell C₁₈ column, 50 mm × 2.1 mm, 2.7 μm) was compared with that of sub‐2 μm fully porous particles (Acquity BEH C₁₈, 50 mm × 2.1 mm, 1.7 μm). Four parabens, methylparaben, ethylparaben, propylparaben, and butylparaben, were used as representative compounds for calculating the plate heights in a wide flow rate range and analyzed on the basis of the Van Deemter and Knox equations. Theoretical Poppe plots were constructed for each column to compare their kinetic performance. Both phases gave similar minimum plate heights when using nonreduced coordinates. Meanwhile, the flat C‐term of the core–shell column provided the possibilities for applying high flow rates without significant loss in efficiency. The low backpressure of core–shell particles allowed this kind of column, especially compatible with conventional high‐performance liquid chromatography systems. Based on these factors, a simple high‐performance liquid chromatography method was established and validated for the determination of parabens in various seafood sauces using the Halo core–shell C₁₈ column for separation.     

On-line HPLC-HRGC coupling and simultaneous transfer of two different LC fractions: Determination of aliphatic alcohols and sterols in olive oil

A modified loop-type interface is decribed which uses two 6-way valves and concurrent eluent evaporation to perform an on-line transfer and simultaneous gas chromatographic analysis of two different fractions pre-separated by liquid chromatography. The interface is used to simultaneously analyze aliphatic alcohols and sterols present in olive oil. LC pre-separation is carried out using a normal phase column (20 cm × 0.21 cm i.d.) and hexane-isopropanol (99:1) as a mobile phase at a flow of 0.2 ml/min; for the GC analysis a 5 % phenyl, 95 % dimethyl siloxane (25 m × 0.32 mm i.d., 0.4 μm film thickness) column is used.     

Core–shell column Tanaka characterization and additional tests using active pharmaceutical ingredients

In the last decade, core–shell particles have gained more and more attention in fast liquid chromatography separations due to their comparable performance with fully porous sub‐2 μm particles and their significantly lower back pressure. Core–shell particles are made of a solid core surrounded by a shell of classic fully porous material. To embrace the developed core–shell column market and use these columns in pharmaceutical analytical applications, 17 core–shell C₁₈ columns purchased from various vendors with various dimensions (50 mm × 2.1 mm to 100 mm × 3 mm) and particle sizes (1.6–2.7 μm) were characterized using Tanaka test protocols. Furthermore, four selected active pharmaceutical ingredients were chosen as test probes to investigate the batch to batch reproducibility for core–shell columns of particle size 2.6–2.7 μm, with dimension of 100 × 3 mm and columns of particle size 1.6 μm, with dimension 100 × 2.1 mm under isocratic elution. Columns of particle size 2.6–2.7 μm were also tested under gradient elution conditions. To confirm the claimed comparable efficiency of 2.6 μm core–shell particles as sub‐2 μm fully porous particles, column performances of the selected core–shell columns were compared with BEH C₁₈, 1.7 μm, a fully porous column material as well.     

Separation of cannabinoids on three different mixed‐mode columns containing carbon/nanodiamond/amine‐polymer superficially porous particles

Three mixed‐mode high‐performance liquid chromatography columns packed with superficially porous carbon/nanodiamond/amine‐polymer particles were used to separate mixtures of cannabinoids. Columns evaluated included: (i) reversed phase (C₁₈), weak anion exchange, 4.6 × 33 mm, 3.6 μm, and 4.6 × 100 mm, 3.6 μm, (ii) reversed phase, strong anion exchange (quaternary amine), 4.6×33 mm, 3.6 μm, and (iii) hydrophilic interaction liquid chromatography, 4.6 × 150 mm, 3.6 μm. Different selectivities were achieved under various mobile phase and stationary phase conditions. Efficiencies and peak capacities were as high as 54 000 N/m and 56, respectively. The reversed phase mixed‐mode column (C₁₈) retained tetrahydrocannabinolic acid strongly under acidic conditions and weakly under basic conditions. Tetrahydrocannabinolic acid was retained strongly on the reversed phase, strong anion exchange mixed‐mode column under basic polar organic mobile phase conditions. The hydrophilic interaction liquid chromatography column retained polar cannabinoids better than the (more) neutral ones under basic conditions. A longer reversed phase (C₁₈) mixed‐mode column (4.6 × 100 mm) showed better resolution for analytes (and a contaminant) than a shorter column. Fast separations were achieved in less than 5 min and sometimes 2 min. A real world sample (bubble hash extract) was also analyzed by gradient elution.     

Semimicro Size Exclusion Chromato-Graphy For Oligomers. II. Separation of Oligomers and Comparison With Conventional Columns

Abstract

Sixteen columns of 1.5 mm i.d. × 25 cm length packed with polystyrene gels of a particle diameter 10 ± 2 μm and having the exclusion limit of 8000 molecular weight as polystyrene were connected in series and used for the separation of oligomers such as oligostyrenes, epoxy resins, methylated melamine-formaldehyde resins, and phenol-formaldehyde resins of novolac and resol types. The observed overall value of the number of theoretical plates was 103000 plates/4 m. Separation results were compared with the conventional SEC which used two SEC columns of 8 mm i.d. × 30 cm length packed with PS gels of a particle diameter 6 ± 2 μm and having the same exclusion limit of the semimicro SEC. The value of N of this column was 17500 plates/30 cm. Oligostyrenes up to n = 11 (undecamer) were separated. Methyl ether derivatives of polynuclear methylol melamines up to penta-nuclear methylol melamine were separated. The possibility for separation of overlapping peaks by SEC having clearly higher efficiency was discussed.     

Optimization of chromatography to overcome matrix effect for reliable estimation of four small molecular drugs from biological fluids using LC–MS/MS

The article describes a systematic study to overcome the matrix effect during chromatographic analysis of gemfibrozil, rivastigmine, telmisartan and tacrolimus from biological fluids using LC–ESI–MS/MS. All four methods were thoroughly developed by the appropriate choice of analytical column, elution mode and pH of mobile phase for improved chromatography and overall method performance. Matrix effect was assessed by post-column analyte infusion, slope of calibration line approach and post-extraction spiking. The best chromatographic conditions established were: Acquity BEH C₁₈ (50 × 2.1 mm, 1.7 μm) column with 5.0 mm ammonium acetate, pH 6.0–methanol as the mobile phase under gradient program for gemfibrozil; Luna CN (50 × 2.0 mm, 3 μm) column with a mobile phase consisting of acetonitrile–10 mm ammonium acetate, pH 7.0 (90:10, v/v) for rivastigmine; Inertsustain C₁₈ (100 × 2.0 mm, 5 μm) column using methanol–2.0 mm ammonium formate, pH 5.5 (80: 20, v/v) as the mobile phase for isocratic elution of telmisartan; and Acquity BEH C₁₈ (50 × 2.1 mm, 1.7 μm) with methanol–10 mm ammonium acetate, pH 6.0 (95:5, v/v) as mobile phase for tacrolimus. The methods were thoroughly validated as per European Medicines Agency and US Food and Drug Administration guidance and were successfully applied for pharmacokinetic studies in healthy subjects.     

Dual-column GC analysis of mediterranean fish for ten organochlorine pesticides and sixty two chlorobiphenyls

The simultaneous analysis of α-HCH, β-HCH, γ-HCH, HCB, p,p′-DDD, p,p′-DDT, p,p′-DDE, o,p′-DDT, mirex, dieldrin and 62 chlorobiphenyl congeners on two parallel capillary GC columns of different polarity is described for nine Mediterranean fish species. Ten commercially available columns with stationary phases completely characterized in respect of their PCB elution patterns were considered for dual-column GC-ECD analysis. The combination of a 60 m × 0.25 mm i.d. column coated with a 0.25 μm film of 50% diphenyl dimethylsiloxane and a series combination of a 25 m × 0.25 mm i.d. column coated with a 0.25 μm film of 5% diphenyl dimethylsiloxane with a 25 m × 0.22 mm i.d. column coated with a 0.10 μm film of 1, 10-dicarba-closo-dodecarborane dimethylpolysiloxane furnished the highest number of separated chlorobiphenyl congeners (104). The dual-column GC system performed with high stability and reproducibility over a broad concentration range (1–3000 ng/g lipid) of the organochlorine compounds in the investigated fish.     

Improved congener-specific GC analysis of chlorobiphenyls on coupled CPSil-8 and HT-5 columns

The congener-specific analysis of polychlorinated biphenyls (PCB) by high resolution gas chromatography on a 50 m × 0.25 mm fused silica column coated with a 0.26 μm film of 5% diphenyl polydimethylsiloxane (CPSil-8) has been significantly improved by series coupling with a 25 m × 0.22 mm column coated with a 0.10 μm film of 1,2-dicarba-closo-dodecarborane polydimethylsiloxane (HT-5). Using helium as carrier gas, a total of 64 congeners in technical PCB mixtures could be analyzed as resolved peaks by ECD (86 by MS) with the CPSil-8 column, and 84 by ECD (108 by MS) with the combination. The high upper temperature limit for these stationary phases (>300°C) enabled fast temperature programming and rapid analysis (60 min).     

Screening and identifying antioxidants from Oplopanax elatus using 2,2ʹ‐diphenyl‐1‐picrylhydrazyl with off‐line two‐dimensional HPLC coupled with diode array detection and tandem time‐of‐flight mass spectrometry

The root of Oplopanax elatus (Nakai) Nakai has a well‐known history of use for the treatment of diseases such as neurasthenia, cardiovascular disorders, and cancer by the native people in northeast China. It is important to screen and identify the bioactive molecules from its root rapidly. Hereby, an off‐line two‐dimensional high performance liquid chromatography coupled with diode array detection and tandem time‐of‐flight mass spectrometry together with 2,2?‐diphenyl‐1‐picrylhydrazyl was established to screen antioxidants from the root of O. elatus. A Waters cyanogen column (150 × 3.9 mm, id, 4 μm) was used for the first dimensional liquid chromatography, while a Hypersil BDS‐C18 column (250 × 4.6 mm, id, 5 μm) was installed for the second dimension liquid chromatographic analysis. Twenty‐eight compounds had been tentatively identified from the methanol extract of the air‐dried root of O. elatus including six polyynes and eight phenolic derivatives were screened with antioxidant activity. The developed method could be expedient for screening and identifying antioxidants from O. elatus.     

Determination of betaine,l‐carnitine,and choline in human urine using a self‐packed column and column‐switching ion chromatography with nonsuppressed conductivity detection

A simple method for the determination of betaine, l ‐carnitine, and choline in human urine was developed based on column‐switching ion chromatography coupled with nonsuppressed conductivity detection by using a self‐packed column. A pretreatment column (50 mm × 4.6 mm, id) packed with poly(glycidyl methacrylate‐divinylbenzene) microspheres was used for the extraction and cleanup of analytes. Chromatographic separation was achieved within 10 min on a cationic exchange column (150 mm × 4.6 mm, id) using maleic anhydride modified poly(glycidyl methacrylate‐divinylbenzene) as the particles for packing. The detection was performed by ion chromatography with nonsuppressed conductivity detection. Parameters including column‐switching time, eluent type, flow rates of eluent, and interfering effects were optimized. Linearity (r ² ≥ 0.99) was obtained for the concentration range of 0.50–100, 0.75–100, and 0.25–100 μg/mL for betaine, l ‐carnitine, and choline, respectively. Detection limits were 0.12, 0.20, and 0.05 μg/mL for betaine, l ‐carnitine, and choline, respectively. The intra‐ and interday accuracy and precision for all quality controls were within ±10.11%. Satisfactory recovery was observed between 92.5 and 105.0%. The validated method was successfully applied for the determination of betaine, l ‐carnitine, and choline in urine samples from healthy people.     

An HPLC‐UV method for determining plasma dimethylacetamide concentrations in patients receiving intravenous busulfan

Dimethylacetamide (DMA) is a solvent used in the preparation of intravenous busulfan, an alkylating agent used in blood or marrow transplantation. DMA may contribute to hepatic toxicity, so it is important to monitor its clearance. The aim of this study was to develop an HPLC‐UV assay for measurement of DMA in human plasma. After precipitation of plasma proteins with acetonitrile followed by dilution (1:4) with water, the extract was injected onto the HPLC and detected at 195 nm. Separation was performed using a Cogent‐HPS 5 μm C₁₈ column (250 × 4.6 mm) preceded by a Brownlee 7 μm RP₁₈, pre‐column (1.5 cm × 3.2 mm). The mobile phase was 25 mm sodium phosphate buffer (pH 3), containing 2.5% (v /v) acetonitrile and 0.0005% (v /v) sodium‐octyl‐sulfonate. Using a flow rate of 1 mL/min, the retention times of DMA and the internal standard (IS), 2‐chloroacetamide, were 9.5 and 3.5 min, respectively. Peak area ratio (DMA:IS) was a linear function of concentration from 1 to 1000 μg/mL. There was excellent intraday precision (<5% for 5–700 μg/mL DMA), accuracy (<3% deviation from the true concentration) and recovery (74–98%). The limits of detection and quantification were 1 and 5 μg/mL, respectively. In eight children who received intravenous busulfan, DMA concentrations ranged from 110 to 438 μg/mL.     

Preparation of Fritless Packed Silica Columns for Capillary Electrochromatography

Fritless packed silica gel columns were prepared using sol‐gel technology. A part of a 75 μm i.d. fused silica capillary was filled with a mixture of tetramethoxysilane and poly(ethylene glycol). After gelling at 40°C and heating at 300°C, the resultant silica gel was derivatized with dimethyloctadecylchlorosilane. A scanning electron micrograph of a cross‐section of the capillary column showed that the gel took the form of a spherical particle aggregate and adhered to the column inner wall. The column performance was evaluated for electrochromatography using acetonitrile–50 mM HEPES buffer (pH 6.6) (60/40 or 40/60, v/v) as the mobile phase. An electroosmotic flow of 1.0 mm/s was generated with (60/40, v/v) acetonitrile/HEPES buffer at a field strength of 546 V/cm. Using a sol‐gel‐derived packed column at an electroosmotic flow of 0.5 mm/s, efficiencies of up to 1.1×10⁵ plates/m were obtained for retained solutes.     

Simultaneous determination of residual polychlorinated biphenyls and organochlorine pesticides in fish and fish products by gas-liquid chromatography

A procedure is developed for the simultaneous determination of polychlorinated biphenyls and organochlorine pesticides in fish and fish products by gas-liquid chromatography with an electron capture detector on a capillary column DB-1701 (30 m × 0.25 mm × 0.25 μm) under temperature-programming conditions. The detection limit is 0.06 μg/kg of the test sample. Polychlorinated biphenyls and organochlorine pesticides were extracted from the samples using a hexane-acetone mixture (1 : 1). Lipids were removed by treatment with H₂SO₄. Eighteen samples were analyzed; the concentrations of pollutants were found to be below the prescribed values.     

Determination of key flavonoid aglycones by means of nano‐LC for the analysis of dietary supplements and food matrices

A method for the analysis of flavonoids (myricetin, quercetin, naringenin, hesperitin, and kaempferol), with interesting bioactivity, has been developed and validated utilizing nano‐LC technique. In order to find optimal conditions, capillary columns (75 μm id × 10 cm) packed with different types of stationary phases, Kinetex® C18 core–shell (2.6 μm particle size), Hydride‐based RP‐C18 (sub‐2 μm particle size), and LiChrospher® 100 RP‐18 endcapped (5 μm particle size) were evaluated. The method was validated using Hydride‐based RP‐C18 stationary phase, with sub‐2 μm particle size. A good chromatographic performance, expressed in terms of repeatability (RSD, in the range 1.63–4.68% for peak area), column‐to‐column reproducibility (RSD not higher than 8.01% for peak area), good linearity and sensitivity was obtained. In particular limit of detection values between 0.07 and 0.31 μg/mL were achieved with on column focusing technique. The method was applied to the determination of studied flavonoids in dietary supplements as well as in food matrices. The amount of quercetin found in the first analyzed dietary supplement, was in agreement to the labeled content. In the other samples, where the content of flavonoids was not labeled, most of the studied flavonoids were determined in amounts somewhere comparable to those reported in literature.     

Semimicro Size Exclusion Chromatography For Oligomers. I. Column Packing Procedure

Abstract

Polystyrene gels of a particle diameter 10 ± 2 μm for the use in oligomer separation were packed into 1.5 mm i.d. × 25 cm length columns by the balanced density slurry-packing technique under a constant flow rate of 500 μL/min. The slurry solvent was a mixture of toluene and chloroform (50.5/49.5, v/v). The example of the number of theoretical plates (N) of these columns was 8600 plates/25 cm (HETP = 29.1 μm) at flow rate of 40 μL/min by injecting 1 μL of 0.5% benzene solution. Sixteen columns were connected and the overall value of N was 103000 plates/4 m. A typical example of oligomer separation was demonstrated. A constant-flow technique is preferable to a constant-pressure technique. When two or three column blanks were packed together, the columns located at the outlet of the packer-column assembly had higher values of N. Optimum flow rate of the slurry solvent when three column blanks were packed together lay between 400 and 500 μL/min. The packing efficiency, that is, the probability of getting valid columns was about 60%. Viscous slurry solvents were not effective to get efficient columns. To pack gels in the less swollen state gave sometimes efficient columns. Pressure monitoring in progress of packing was very effective to foresee the column efficiency.     

Direct separation of the enantiomers of ramosetron on a chlorinated cellulose‐based chiral stationary phase in hydrophilic interaction liquid chromatography mode

Ramosetron is an enantiopure active pharmaceutical ingredient marketed in Japan since 1996 and later in a few Southeast Asian countries predominantly as an antiemetic for patients receiving chemotherapy. In this study, a simple and rapid high‐performance liquid chromoatography method for the separation of ramosetron and its related enantiomeric impurity by using hydrophilic interaction liquid chromatography mode is presented. Chiral resolution was performed on an analytical column (100 mm × 4.6 mm id) packed with 3 μm particles of cellulose‐based Chiralpak IC‐3 chiral stationary phase. Using a mobile phase containing acetonitrile–water–diethylamine (100:10:0.1, v/v/v) and setting the column temperature at 35°C, the resolution value was 7.35. At a flow rate of 1 mL/min, the enantioseparation was completed within 5 min. The proposed method was partially validated and it has proven to be sensitive with limit of detection and limit of quantitation of the (S)‐enantiomer impurity of 44.5 and 133.6 ng/mL.     

High-Performance Liquid Chromatographic Determination of Roxithromycin in Tablets

Abstract

For the quantitative determination of Roxithromycin in tablets a rapid, and simple HPLC assay was developed. Reversed-phase chromatography was conducted using a RPC 18 (3.9 × 150 mm, 5 μm) and guard column RP C 18 (3.0 × 3.9 mm, 5 μm), mobile phase of 0.067 M phosphate buffer, pH 4.0 and methanol (65:35), and UV detection at 210 nm. Mean recovery of 100.90% and percentual coefficient of variation (CV%) of 1.51% was obtained from five commercial samples of Roxithromycin. The calibration graph was a straight line (r = 0.9995). Linearity is observed in the concentration range from 50.0 to 250.0 μg/ml. The excipients did not interfere in the determination.     

Separation of complex sugar mixtures on a hydrolytically stable bidentate urea‐type stationary phase for hydrophilic interaction near ultra high performance liquid chromatography

A stationary phase bearing both bridged bis‐ureido and free amino groups (USP‐HILIC‐NH₂–2.5SP) for high‐speed hydrophilic interaction liquid chromatography separations was prepared using a one‐pot two‐step procedure starting from 2.5 μm totally porous silica particles. Highly polar compounds, such as polyols, hydroxybenzoic acids, and sugars, were successfully analyzed in shorter times and with higher peak efficiency, when compared to results obtained with a bidentate urea‐type column packed with 5 μm particles. Increased sugarophilicity and better peak shape were attested for the USP‐HILIC‐NH₂–2.5SP column (100 × 3.2 mm id) when compared with two commercially available UHPLC columns, namely an acquity BEH amide packed with totally porous 1.7 μm microparticles and a HILIC Kinetex column packed with core–shell 2.6 μm particles. Finally, the new column was employed in the separation of complex mixture of sugars (mono‐, di‐, and oligosaccharides) and in the analysis of beer samples. The resulting chromatograms showed good selectivity and overall resolution, while the catalyzing effect of the free amino moieties resulted in excellent peak shapes and in the absence of split peaks due to sugar anomerization phenomena.     

High Performance Liquid Chromatography Separations Using Columns Packed with Spherical ODS Particles. III. Effect of Column Dimensions on the Resolution of a Complex Mixture

Abstract

Separations on short columns, (3 and 5 cm, packed with 3 μ ODS spherical materials) and somewhat larger ones (10 cm and 20 cm columns having 2.1 mm and 4.6 mm diameters packed with 5 μ ODS spherical materials) were compared using Aroclor 1254. With simple mixtures, the results showed that short columns can give separations comparable with those on longer columns when the percentage of the organic modifier in the mobile phase is adjusted. This was not so with more complex mixture. The results also showed that columns which have a comparable volume do not produce comparable separation. The longer column, 200 mm × 2.1 mm gave better resolution than the shorter 50 mm × 4 mm column. Also a shorter column, (100 mm × 4.6 mm), which had double the volume of a longer column. (200 mm × 2.1 mm), gave better resolution of the Aroclor 1254 test solution.     

Cryogenic modulation fast GC × GC–MS using a 10 m microbore column combination: Concept,method optimization,and application

The present research is based on the concept of using a 10 m × 0.1 mm id column for cryogenic‐modulation fast comprehensive two‐dimensional gas chromatography with quadrupole mass spectrometry. Specifically, an 8.9 m × 0.1 mm id low‐polarity column was used as the first dimension, and a 1.1 m × 0.1 mm id medium‐polarity column was used as the second dimension. The main scope of the investigation was to develop a high peak‐capacity method, with an analysis time of approximately 10 min. Various aspects related to method optimization are discussed, as well as separation parameters such as peak capacity (in each dimension, and as a total value), first‐dimension sample capacity, peak widths, modulation ratio, sensitivity enhancement, and number of spectra per peak. The fast approach was evaluated in applications involving a mixture of cosmetic allergens and a sample of perfume. The approach proposed enables high‐resolution separations in a short time (across the C₈–C₂₃ alkane range), as well as a considerable reduction of the consumption of gases for modulation cooling and heating.