Homemade Column Research Articles

Investigation of Retention Mechanism of B2 and B3 in Home-Made ZIC-HILIC Columns and Its Application for The Simultaneous Estimation in Multivitamin Tablets

In the present study, the retention behavior of two vitamins (B2 and B3) was investigated using a zwitterionic hydrophilic interaction liquid chromatography technique (ZIC-HILIC) on two home-made columns, and two analytical methods for these vitamins were established as a result. Several chromatographic parameters were carefully investigated and optimized. The best separation was achieved on two new home-made columns (ZIC-1 and ZIC-3) employing acetonitrile/sodium acetate buffer as the mobile phase (40 mM, pH 4.75). The target vitamins were detected at 254 nm. In the ZIC-1 and ZIC-3 columns, the linear ranges of the target vitamins (B2 and B3) were 0.06-11.0 and 0.08-7.0 µg/mL, respectively. For both target vitamins, the detection and quantitation limits of the suggested methods were 0.011-0.055 and 0.033-0.166 µg/mL, respectively. They also showed high precision (RSD ≤ 0.93), linearity (r2 ≥ 0.9988), and recovery (between 99.0 and 101.71%). The suggested procedures have been validated and tested on multivitamin tablets, proving that they are suitable for analyzing food supplements for these tablets, and acceptable results were obtained for the study.

Evaluation of organo-vermiculites as sorbent phases for solid-phase extraction of ibuprofen from water.

The study of ibuprofen (IBU) preconcentration was carried out making use of a homemade column for solid-phase extraction (SPE), using vermiculite (VT) or organo-vermiculites (OVTs) as sorbent phases. Aqueous samples (50.0 mL) percolated the column and IBU was sorbed onto the VT or OVT and then desorbed using acetonitrile. Employing this SPE system and OVT, calibration curves were generated for IBU, by spectrophotometric quantification using the α-naphthylamine method. R2 values higher than 0.9950 and LOD between 12 and 18 μg L-1 were observed, for real enrichment factors of 21 and 31, by using OVTs. The analytical protocol was applied to three water samples, which were spiked with IBU solutions to evaluate the precision and accuracy of the method. Recoveries between 77 and 110% at three different IBU concentrations and RSD lower than 18% were observed, even by using the spectrophotometric method. The protocol developed in this study demonstrated that the OVT was appropriate to work as a preconcentration phase for IBU determination in water samples.

Comparison of 624-type Capillary Columns

In our research, seven 624-type capillary columns were investigated. All the columns were the same in length, internal diameter, and film thickness (30 m × 0.32 mm × 1.0 µm). However, they were produced by different manufacturers or the same manufacturer but in different batches. Even though the manufacturers recommend them as "equivalent columns" this equivalence did not prevail even in the case of columns produced by the same manufacturer. Our examination criteria centered on the quantitative determination ability of the columns. A homemade column test mixture was compiled to represent all the second-order interactions that can occur between the analyte and stationary phase. Although theoretically these columns have the same stationary phase quality, they did not result in the same chromatograms. In addition to the origin and batch of the column, the "history of the column" contributes likewise to the different peak symmetry, retention order, and even peak areas that affect the quantitative determination. We quantified this quantitative determination ability with the effective carbon number (ECN) and the Limit of Quantitation (LoQ) values. Based on our results the attainable LoQ and ECN values depend at least as much on the origin and actual state of the stationary phase as on the measurement conditions to be optimized. In our paper, we demonstrate the extent to which the same stationary phases offered by different companies and/or different backgrounds can influence our detection limit and detector response even if the relevant columns have theoretically the same chemical structure.

Chromatographic Regulation by the Building Blocks for Triazine Polymer-Based Core–Shell Stationary Phases

Two versions of reversed-phase/hydrophilic interaction mixed-mode core–shell stationary phases for high-performance liquid chromatography using porous organic polymers as the shell material were investigated. The stationary phase named SiO2@CTPCC-TPB using triazine-triphenylbenzene-based porous organic polymers as the shell material has been reported previously. To regulate chromatographic performance, heptazine–biphenyl moieties with fewer benzene rings and higher nitrogen content were used as organic building blocks to grow a porous shell on the surface of silica to acquire a core–shell stationary phase, namely, SiO2@CHPCy-DB. Various techniques were used to characterize the developed core–shell microsphere, and nonpolar/polar solutes were utilized as probes to investigate the chromatographic behavior of the stationary phase. The results revealed that SiO2@CHPCy-DB exhibited reversed-phase/hydrophilic interaction mixed-mode with multiple retention mechanisms. Meanwhile, the separation behavior of the two homemade columns and a commercial column was compared to assess the role of the building blocks in porous organic polymers in regulating and improving separation. The successful determination of drugs of abuse in urine via the SiO2@CHPCy-DB column indicated its potential for the determination of trace analytes in complex samples.

Mechanically stable core-shell cellulose nanofibril/sodium alginate hydrogel beads with superior cu(II) removal capacity

Nanocellulose hydrogels are promising sustainable biosorbents for removing heavy metal ions for wastewater treatment. However, the nanocellulose hydrogels reported thus far typically suffer from inferior adsorption performance and/or poor mechanical stability, thus limiting their industrial applications. Achieving the goals of mechanical stability and high removal capability remains a crucial technical challenge, which may be addressed, as presented in this study, by developing novel core-shell carboxymethylated cellulose nanofibril (CMCNF)/sodium alginate (SA) hydrogel beads (CAbs). By immobilizing CMCNFs (shell) on the surface of the SA hydrogel bead (core) via electrostatic attractions and hydrogen bonding, a mechanically stable hydrogel bead with a core-shell configuration was constructed, which shows a Cu(II) removal capacity of up to 221 mg/g that exceeds that of CMCNFs and most other nanocellulose structural adsorbents. Furthermore, both the formation principle of the core-shell structure and the Cu(II) removal mechanism were explored in detail. Finally, we demonstrated a potential application of core-shell CAbs to treat drinking water with a low concentration of Cu(II) using a homemade column adsorption device. This work brings sustainable nanocellulose adsorbents a step closer to industrial applications for Cu(II) wastewater treatment.

Development of Narrow-Bore C18 Column for Fast Separation of Peptides and Proteins in High-Performance Liquid Chromatography.

Separation with high efficiency and good resolution is constantly in demand in the pharmaceutical industry. The fast and efficient separation of complex samples such as peptides and proteins is a challenging task. To achieve high efficiency with good resolution, chromatographers are moving towards small particles packed into narrow-bore columns. Silica monolith particles (sub-2 µm) were derivatized with chlorodimethyl octadecyl silane (C18) and packed into stainless steel columns (100 mm × 1.8 mm i.d) by a slurry-packing method. The developed columns were used for the separation of peptides and proteins. A separation efficiency (N) of 40,000 plates/column (400,000 plates/m) was achieved for the mixture of five peptides. Similarly, the fast separation of the peptides was carried out using a high flow rate, and the separation of the five peptides was achieved in one minute with high efficiency (N ≅ 240,000 plates/m). The limit of detection (DL) and the limit of quantification (QL) for each analyte were determined by developing a linear regression curve with relatively very low concentrations of the target compound. The average values of the QL for the peptide and proteins were 0.55 ng and 0.48 ng, respectively, using short C18 column (1.8 mm × 100 mm) UV (at 214 nm). The fast analysis of peptides and proteins with such high efficiency and good resolution has not been reported in the literature yet. Owing to high efficiency, these home-made columns could be used as an alternative to the expensive commercial columns for peptide and protein separation.

On-Site Determination of Methylmercury by Coupling Solid-Phase Extraction and Voltammetry.

A measurement and speciation procedure for the determination of total mercury (HgTOT), inorganic mercury (HgIN), and methylmercury (CH3Hg) was developed and the applicability for on-site determination was demonstrated. A simple, portable sample pretreatment procedure was optimized to extract the analytes. Home-made columns, packed with a new sorbent material called CYXAD (CYPHOS 101 modified Amberlite XAD), were used to separate the two forms of the analyte. HgTOT and CH3Hg were determined by anodic stripping voltammetry (ASV), using a solid gold electrode (SGE). Two certified reference materials (BCR-463 Tuna Fish and Tuna Fish ERM-CE 464) and eight fresh fishes were analyzed. Then, the results that were obtained following the optimized portable procedure were compared with the concentrations obtained, using a direct mercury analyzer (DMA). This quantification, using the two techniques, demonstrated the good performance of the proposed method.

Novel amyloid-like porous lysozyme skeletons as “green” superadsorbent presenting ultrahigh capacity and rapid sequestration towards hazardous Congo red

Finding routes to processing the water-soluble proteins into well-organized materials with rational structures holds a massive challenge for adsorption-based environmental remediation. Herein, one strategy to answer this challenge is to utilize the enhanced amyloidal denaturation protocol to achieve phase transition of egg-white lysozyme with the generation of a novel biosorbent with superadsorption ability for Congo red (CR) removal, namely porous lysozyme skeletons. The as-prepared superadsorbent exhibits three-dimensional skeletal geomorphology and highly porous structures (240.2 m2/g), both beneficial for adsorption. As expected, this amyloidal adsorbent shows remarkable uptake performances for CR with rapid kinetics (≤20 min) and an ultrahigh capacity of 2.00 × 103 mg/g in the form of multilayer adsorption, as well as the removal adaptability over wide ranges of solution chemistry variation. Additionally, a remarkable adsorption selectivity towards CR in the presence of another anionic/cationic dyes and robust recyclability after undergoing twenty successive cycles are discerned. More importantly, the resultant lysozyme skeletons can effectively realize immediate decoloration flowing across homemade column filtration. The scavenging mechanisms are concluded as the cooperation of the electrostatic attraction, hydrophobic interaction, H-bonding, π-π stacking, β-sheet affinity, and pore-filling effect. Overall, this “green” superadsorbent can be deemed as an attractive project for alleviating dyestuff pollution.

One-minute highly selective Cr(VI) determination at ultra-trace levels: An ICP-MS method based on the on-line trapping of Cr(III)

An analytical method derived from the coupling of frontal chromatography (FC) with Inductively Coupled Plasma – Mass Spectrometry (ICP-MS) is proposed for the fast determination of Cr(VI) ultra-traces. The insertion of a short, homemade column filled with a strong cationic exchange resin in the flow-path of a commercial ICP-MS allows on-line trapping of cationic Cr(III) and elution of anionic Cr(VI). As a result, only the Cr(VI) front reaches the detector. This separation mechanism enables the highly selective quantification of Cr(VI) ultra-traces (LOD = 0.026 µg/kg - defined as 3 s of 10 replicated measurements of a 0.050 µg/kg solution) over a wide linearity range (tested up to 1024 µg/kg), even in the presence of Cr(III) concentration as high as 50 mg/kg. Key advantages of the proposed method are the extremely short analysis time (one minute), together with the simplicity and cost-effectiveness of the modifications applied over a commercial ICP-MS instrumental configuration. No time- or chemical-consuming pretreatments are needed: it is only necessary to acidify the sample prior Cr(VI) determination, as normally performed for common ICP-MS analysis. The applicability of the method was demonstrated over mineral water samples and toy migration solutions.

Zwitterion Chromatography-Hydrophilic Interaction Chromatography for Separation and Quantitative of Rutin and Quercetin from Herbs and Bee Products

Quercetin and rutin have a number of useful pharmacological consequences and a possible medication of the future. The growth of cancer cells was observed in vitro and the production of tumors in experimental animals was decreased. In addition to preventing human cancer induction and progression. A set of four HILIC stationary phases, two home-made columns with different space chains between the ionic site groups, and two commercially stationary phases HALOand#174;HILIC 2.7 and ZICand#174;–pHILIC for chromatographic separation and quantitation of quercetin and rutin in herbs and bee products. The separation mechanisms are based on hydrophobic and hydrophilic for the quercetin and rutin, respectively. The validated method was successfully applied in bee products and herbs. The results showed that the HILIC methods were simple and reliable and could be used in bee products and herbs to detect the content of quercetin and rutin.

Zwitterion Chromatography-Hydrophilic Interaction Chromatography for Separation and Quantitative of Rutin and Quercetin from Herbs and Bee Products

Quercetin and rutin have a number of useful pharmacological consequences and a possible medication of the future. The growth of cancer cells was observed in vitro and the production of tumors in experimental animals was decreased. In addition to preventing human cancer induction and progression. A set of four HILIC stationary phases, two home-made columns with different space chains between the ionic site groups, and two commercially stationary phases HALOand#174;HILIC 2.7 and ZICand#174;–pHILIC for chromatographic separation and quantitation of quercetin and rutin in herbs and bee products. The separation mechanisms are based on hydrophobic and hydrophilic for the quercetin and rutin, respectively. The validated method was successfully applied in bee products and herbs. The results showed that the HILIC methods were simple and reliable and could be used in bee products and herbs to detect the content of quercetin and rutin.

Preparation and Characterization of Glycidyl Polymethacrylate Monolith Column and its Application for Simultaneous Determination of Paracetamol and Chlorzoxazone in Their Combined Pharmaceutical Formulations

A home-made column (3.2 mm i.d., 100 mm length) was prepared in which glycidyl polymethacrylate was used. The optimum value of the initiator corresponded to 5 mg/mL. The column morphology was characterized by scanning electron microscopy. The permeability was evaluated using acetonitrile and water as a mobile phase and uracil as an unretained substrate. A simple and economical reversed-phase HPLC method has been developed for the simultaneous estimation of paracetamol and chlorzoxazone in their pharmaceutical formulations. Components were determined using a UV detector at 270 nm. The mobile phase was composed of 1% formic acid solution and acetonitrile (65/35, v/v); 0.7 mL/min flow rate and 5.0 µL injection volume were used. Peak resolution was 1.96. All findings allow concluding that the prepared stainless steel conventional HPLC column and the novel validated method are applicable for quality control and routine analysis.

An improved pollen number counting method using a cell counter and mesh columns

BackgroundThe determination of pollen number is important in evolutionary, agricultural, and medical studies. Tree species of the Cupressaceae family cause serious pollinosis worldwide. Although Japanese cedar (Cryptomeria japonica) is the most important forestry species in Japan, it is also the biggest cause of pollinosis in the country. Japanese cedar trees have been selected for growth speed and superior morphological traits and then cloned. These clones may vary in their pollen production, but there has been little research on how many pollen grains are produced by a single male strobilus (flower). A recently reported method for counting pollen number with a cell counter was applicable to Arabidopsis species and wheat, but was not suitable for Japanese cedar because the strobilus does not open with heating (e.g. 60 °C, overnight).ResultsHere, we report an improved pollen counting method for Japanese cedar using a precise and rapid cell counter in combination with home-made mesh columns. The male strobilus was gently crushed using a pestle. Large and small debris were then removed using 100- and 20-μm mesh columns, respectively. We successfully detected pollen sizes and numbers that differed between two clones using this method.ConclusionsThis improved method is not only suitable for counting pollen from Japanese cedar, but could also be applied to other species of the Cupressaceae family with hard scale tissue covering the pollen. Moreover, this method could be applied to a broader range of plant species, such as wheat, because there is no need to wait for anthesis and debris can be removed efficiently.

Exploration of the Selectivity and Retention Behavior of Alternative Polyacrylamides in Temperature Responsive Liquid Chromatography

Temperature responsive liquid chromatography (TRLC) allows for separation of organic solutes in purely aqueous mobile phases whereby retention is controlled through temperature. The vast majority of the work has thus far been performed on poly[N-isopropylacrylamide] (PNIPAAm)-based columns, while the performance of other temperature responsive polymers has rarely been compared under identical conditions. Therefore, in this work, two novel TRLC phases based on poly[N-n-propylacrylamide] (PNNPAAm) and poly[N,N-diethylacrylamide] (PDEAAm) are reported and compared to the state of the art PNIPAAm based column. Optimal comparison is thereby obtained by the use of controlled radical polymerizations, identical molecular weights, and by maximizing carbon loads on the silica supporting material. Analysis of identical test mixtures of homologue series and pharmaceutical samples revealed that PNNPAAm performs in a similar way as PNIPAAm while offering enhanced retention and a shift of the useable temperature range toward lower temperatures. PDEAAm offers a range of novel possibilities as it depicts a different selectivity, allowing for enhanced resolution in TRLC in, for example, coupled column systems. Reduced plate heights of 3 could be obtained on the homemade columns, offering the promise for reasonable column efficiencies in TRLC despite the use of bulky polymers as stationary phases in HPLC.

The polystyrene-divinylbenzene stationary phase modified with poly (Amine-Epichlorohydrin) for ion chromatography

A novel anion exchange stationary phase was synthesized based on polystyrene-divinylbenzene beads and poly (amine-epichlorohydrin) cationic polyelectrolyte. Epichlorohydrin and methylamine as monomers were used to prepare quaternized cationic polyelectrolyte and hyperbranched layers with a denser charged density and high hydrophilcity. Scanning electron microscopy (SEM), Brunauer–Emmett–Teller (BET), Fourier transform infrared spectra (FTIR) and elemental analysis were utilized to characterize the particle surface and chemical composition of modified microspheres. The chromatographic evaluation was investigated by separating the analytes containing of common anions (fluoride, chloride, nitrate, phosphate and sulfate) and polarizable anions (iodide, thiocyanate and thiosulfate). High column efficiency was obtained with the plate number per meter up to 20,607 N/m for chloride. Fast separation of five common anions and three polarizable anions was accomplished within 11 min. Finally, the contents of thiosulfate in water reducing agent samples were determined with the homemade column. The linearity of the method was satisfactory over a concentration range of 5–120 mg/L with a correlation coefficient of 0.9972. The relative standard deviations were all less than 1.50% for retention time, peak height and peak area. The recoveries over the range of 93.67–116.15% were obtained by analyzing real spiked samples. The result suggested that the prepared stationary phase exhibited effective separation, desirable reliability and considerable stability for application in common anions and polarizable anions analysis.

Simple quantitative analytical methods for the determination of alkaloids from medicinal and edible plant foods using a homemade chromatographic monolithic column

A polymer-based chromatographic monolithic column was prepared via in-situ radical polymerization using tetrahydrofurfuryl methacrylate as the monomer. The homemade column was used for the separation and quantitative analysis of alkaloids, including piperine from Piper longum (fruit of Piper longum Linn.) and pepper (fruit of Piper nigrum L.), hydroxy-α-sanshool, and hydroxy-γ-sanshool from zanthoxylum (fruit of Zanthoxylum bungeanum Maxim), as well as caffeine from Wuyi rock tea. The chromatographic fractions were identified by mass spectrometry. Single factor test and orthogonal test were both carried out to optimize the extraction conditions. The method validation indicated that the accuracy represented by spiked recovery ranged in 98.89%–102.06%, the correlation coefficients in 0.99986–0.99999. These results show that the prepared monolithic column can be successfully used to quantitatively analyse alkaloids from the real medicinal and edible plant foods with reversed-phase mechanism, which can avoid the long analytical time using traditional packed C18 column. The present method is a simple, and inexpensive method for quantitatively analysing alkaloids from medicinal and edible plant foods, exhibiting good specificity and durability.

Purification of antibody against Ara h 2 by a homemade immunoaffinity chromatography column

ABSTRACTAntibodies are used extensively in numerous applications both in vivo and in vitro. To purify anti-Ara h 2 polyclonal antibody, a homemade immunoaffinity chromatography (IAC) column method was established. The properties of homemade column were compared with those of the mAb affinity protein G (MPG) agarose high flow, a commercially available column successfully used in capturing polyclonal antibodies. During antibody purification from rabbits’ antiserum against Ara h 2, the column capacity, recovery, and purification factor were characterized for IAC and MPG. The homemade IAC could separate the corresponding antibody with higher specificity and lower cost but with lower recovery and column capacity than those of MPG. Thus, the homemade IAC is a specific, inexpensive, and suitable method that can be used for various laboratory purifications.

Mathematical modeling of the whole expanded bed adsorption process to recover and purify chitosanases from the unclarified fermentation broth of Paenibacillus ehimensis.

In this study, a general rate model was applied to the entire process of expanded bed adsorption chromatography (EBAC) for the chitosanases purification protocol from unclarified fermentation broth produced by Paenibacillus ehimensis using the anionic adsorbent Streamline® DEAE. For the experiments performed using the expanded bed, a homemade column (2.6cm×30.0cm) was specially designed. The proposed model predicted the entire EBA process adequately, giving R2 values higher than 0.85 and χ2 as low as 0.351 for the elution step. Using the validated model, a 33 factorial design was used to investigate other non-tested conditions as input. It was observed that the superficial velocity during loading and washing steps, as well as the settled bed height, has a strong positive effect on the F objective function used to evaluate the production of the purified chitosanases.

Analytical Method Development and Validation for Some Persistent Organic Pollutants in Water and Sediments by Gas Chromatography Mass Spectrometry

Many of the persistent organic pollutants (POPs) with endocrine disrupting properties are monitored regularly by risk assessors with limited resources, where analytical procedures are usually laborious, expensive, and not ecofriendly. Moreover, these analyses were frequently advanced aiming one class of pollutants, consequently inefficient to correspond the demand of monitoring a quickly rising number of pollutants in the environment. The objective of this study was to develop a single sample extraction procedure and multiple gas chromatography-mass spectrometry runs for the detection of various groups of semi volatile organics; Polychlorinated Biphenyls (28, 52, 101, 118, 138, 153, 180), Polybrominated Diphenyl Ethers (17, 47, 66, 100, 153, 183) and Organochlorine Pesticides (α-HCH, HCB, γ-HCH, Heptachlor, p,p-DDD, p,p-DDE, p,p-DDT) in sediment and water samples simultaneously. Extraction for water involved solid phase extraction using C18 and for sediment using homemade column with florisil, primary secondary amine and magnesium sulfate with ultrasonication step by acetone. This procedure was validated and applied to water samples from tap, river and lake; and sediment samples from river and lake. For both matrices and all analytes, high linearity, recovery (88-106%) with all relative standart deviation values <20% and limit of quantification levels below the tested limits were achieved. This reliable and cost effective procedure for monitoring selected multiple POP levels in water and sediment; do not require complicated device nor intensive manual efforts, which would also minimize the depletion of the organic solvents, could be used for routine detection of selected POPs.

Recovery and purification of chitosanase produced by Bacillus cereus using expanded bed adsorption and central composite design.

This study presents a system for expanded bed adsorption for the purification of chitosanase from broth extract in a single step. A chitosanase-producing strain was isolated and identified as Bacillus cereus C-01 and used to produce chitosanases. The expanded bed adsorption conditions for chitosanase purification were optimized statistically using STREAMLINE(TM) DEAE and a homemade column (2.6 × 30.0 cm). Dependent variables were defined by the quality criteria purification factor (P) and enzyme yield to optimize the chromatographic process. Statistical analyses showed that the optimum conditions for the maximum P were 150 cm/h load flow velocity, 6.0 cm settled bed height, and 7.36 cm distributor height. Distributor height had a strong influence on the process, considerably affecting both the P and enzyme yield. Optimizing the purification variables resulted in an approximately 3.66-fold increase in the P compared with the value under nonoptimized conditions. This system is promising for the recovery of chitosanase from B. cereus C-01 and is economically viable because it promotes the reduction steps.