Radial flow rapid pressure swing adsorption

A new PSA process has been proposed and experimentally verified. This process was operated with a radial flow geometry under a cycle time less than 30 seconds. It has been showed that enriched oxygen could be produced when air was fed inward. The same system showed virtually no separation effect if the feed direction was reversed. The change of separation efficiency upon flow reversal was most significant when small adsorbent particles were employed. A ø 200×75 mm annular packing with 3 µm particles of zeolite 5A was able to produce 60% purity oxygen from air. The effect of flow direction on system performance confirmed the importance of flow resistance distribution. In radial flow geometry, most of the flow resistance was located near the center of the disk. The relative small pressure gradient at the feed end enabled a better absorbent utilization during the inward feed step, and a more effective desorption during the vent step. The same principle could be extended to other geometric configurations.     

Capillary supercritical fluid chromatography with flame photometric detection using a large bore column SFC pumping system

A commercially available instrument with an SFC pumping system suitable for wide bore columns (4.6 mm i.d.) has been modified for capillary supercritical fluid chromatography (CSFC) by incorporating a double-stage flow splitter. The first flow splitter was installed in front of the sample injection valve in order to avoid a high solute split ratio. The second splitter was mounted in the column oven so that the injected sample (0.2 μL) would be split to the capillary column. In order to perform pressure programmed elution, a pressure regulating system equipped with a gradient programmer has been used. Flame photometric detection was optimized for the analysis of organosulfur compounds by CSFC. In this study, detection limits were found to be 6–14 ng and the experimentally determined exponent (n value) varied from 1.721 to 1.984 depending on the compounds tested. Sulfur- and phosphorus-containing thermally labile pesticides can be chromatographed and selectively detected by using CSFC/FPD in either sulfur- or phosphorus mode, respectively.     

Visualization of sample introduction in liquid chromatographic columns. Contribution of a flow distributor on the sample band shape

The contributions to the radial distribution of the sample concentration across the column inlet and to the axial band dispersion resulting from a column header containing a distributor were evaluated using a band-visualization process entailing matching the refractive indices of the stationary and mobile phases in a glass column. This study illustrates graphically how a distributor fitted to the column can increase the axial dispersion of the sample band compared to an inlet containing only a frit. The distributor did not provide a uniform sample distribution across the column. In fact, for 17-mm inner diameter columns and high-porosity frits, the distribution was no better than with the frit having no distributor. However, when low-porosity frits were employed, improved peak shapes were obtained with a distributor. In addition, we observed that the inlet header configuration influenced dramatically the flow stream established along the column. The radial distribution of the efficiency of the columns was nearly homogeneous for those having only a frit but not for those having also a distributor. For the latter, the efficiency decreased from the column axis to its wall.     

Are axial and radial flow chromatography different?

Radial flow chromatography can be a solution for scaling up a packed bed chromatographic process to larger processing volumes. In this study we compared axial and radial flow affinity chromatography both experimentally and theoretically. We used an axial flow column and a miniaturized radial flow column with a ratio of 1.8 between outer and inner surface area, both with a bed height of 5 cm. The columns were packed with affinity resin to adsorb BSA. The average velocity in the columns was set equal. No difference in performance between the two columns could be observed. To gain more insight into the design of a radial flow column, the velocity profile and resin distribution in the radial flow column were calculated. Using mathematical models we found that the breakthrough performance of radial flow chromatography is very similar to axial flow when the ratio between outer and inner radius of the radial flow column is around 2. When this ratio is increased, differences become more apparent, but remain small. However, the ratio does have a significant influence on the velocity profile inside the resin bed, which directly influences the pressure drop and potentially resin compression, especially at higher values for this ratio. The choice between axial and radial flow will be based on cost price, footprint and packing characteristics. For small-scale processes, axial flow chromatography is probably the best choice, for resin volumes of at least several tens of litres, radial flow chromatography may be preferable.     

A highly sensitive method (supercritical fluid chromatography coupled with ion mobility mass spectrometry) for determination of multiple compounds in radix curcumae

A highly sensitive method was developed for simultaneously separating and identifying multiple compounds in radix curcumae. The determination of these compounds was achieved by combining supercritical fluid chromatography with drift tube ion mobility quadrupole time-of-flight MS. Related parameters were optimized: the RX-SIL column was used as the stationary phase, methanol was selected as the organic modifier, back pressure was 120 bar, back temperature was 60°C, the mobile phase flow rate was 1.75 mL/min, the makeup solvent was 0.2% formic acid/methanol with a flow rate of 0.7 mL/min. Under optimal conditions, multipolar compounds were separated. Furthermore, these compounds were identified by the values of collision sectional areas. The established method was verified by related parameters and exhibited good linearity, sensitivity, precision and accuracy. It could be extended to analyze other curcuminoids and sesquiterpenoids in natural products.     

Stage-frit: A straightforward sub-2 μm nano-liquid chromatography column fabrication for proteomic analysis

In this work we demonstrated a facile method for the fabrication of C18 coordination polymer gel in a capillary, called stage-frit, which was efficiently applied to pack sub-2 μm C18 beads into the capillary by a high pressure bomb for the online separation of proteolytic peptides. The back pressure of the column with 10 cm × 75 μm i.d. is regularly lower than 170 bar at a flow rate of 300 nl/min, which could be operated on a common nanoLC system instead of nanoUPLC system due to the good permeability, low back pressure and high mechanical stress of the frit that will totally reduce the cost for the purchase of instrument. The stage-frit allows long-term continuous flow of the solvent and no significant beads loss or pressure instability was observed during the period. The repeatability of retention time for fifteen BSA tryptic peaks was found to be less than 1.08% (RSD) in six time nanoLC-ESI-MS/MS experiments. The average full width at half maximum (FWHM) of peptide peaks is 5.87 s. The sub-2 μm stage-frit nanoLC column showed better sensitivity than the commercial available for large scale proteomic analysis of total tissue proteins from human spleen. The number of identified peptides is approximately 0.4-fold and 0.2-fold higher than that obtained by utilizing commercial columns packed with 3 μm and 1.8 μm C18 materials, respectively. In the field of analytical chemistry, particularly the use of nanoLC systems, stage-frit nanoLC column offers a great potential for the separation of complex mixtures.     

Characteristics of elution profile in radial chromatography under linear conditions

Radial chromatography is one of the most efficient tools for the fast analytical and preparative separation of complex samples. Compared to conventional axial liquid chromatography, it has obvious advantage espe- cially in the preparation of biological sa…     

Temperature‐programmed multicapillary gas chromatograph microcolumn for the analysis of odorous sulfur pollutants

We report the fabrication and performance of a silicon‐on‐glass micro gas chromatography eight‐capillary column based on microelectromechanical systems technology that is 50 cm long, 30 μm wide, and 300 μm deep. According to the theory of a gas chromatography column, an even gas flow among different capillaries play a vital role in the peak broadening. Thus, a flow splitter structure is designed by the finite element method through the comparison of the velocity distributions of the eight‐capillary columns with and without splitter as well as an open tubular column. The simulation results reveal that eight‐capillary column with flow splitters can receive more uniform flow velocity in different capillaries, hence decreases the peak broadening and in turn increases the separation efficiency. The separation experiment results show that the separation efficiency of about 22 000 plates/m is achieved with the chip column temperature programmed for analysis of odorous sulfur pollutants. This figure is nearly two times higher than that of the commercial capillary column coated the similar stationary phase. And the separation time of all the components in the microcolumn is less than 3.8 min, which is faster than the commercial capillary column.     

超临界流体色谱法分析非对映异构体d4T-5’-N-磷酰化苯丙氨酸甲酯

采用超临界CO2流体色谱技术,分析d4T-5’-N-磷酰化苯丙氨酸甲酯手性磷的非对映异构体。色谱柱为Hpersil ODS2(250 mm×4.6 mm,5μm),流动相为夹带改性剂甲醇、乙醇和异丙醇的超临界CO2流体。以容量因子、选择性和分离度为指标,考察改性剂、背压和柱温对分离的影响。在甲醇、乙醇和异丙醇3种改性剂中,甲醇为最好的改性剂,其中在7%甲醇改性剂下,该化合物的分离度可达到3.35。在7%甲醇改性剂条件下,考察了压力(10～20 MPa)和温度(303.15～318.15 K)的影响。在优化的分离条件(改性剂为7%甲醇,流速为2 mL/min,柱温为308.15 K,背压为15 M Pa)下,d4T-5’-N-磷酰化苯丙氨酸甲酯的两种非对映异构体完全达到基线分离,分离时间约15 min。     

Improved liquid chromatography-Online radioactivity detection for metabolite profiling

If very-high-pressure liquid chromatography (VHPLC) is to replace conventional HPLC as the ultimate separation tool for metabolism studies in development, coupling it efficiently with online radioactivity detection (RAD) is needed. We describe the successful combination of VHPLC/RAD, facilitated by improvements in online radioactivity detection, as well as in column loading and peak capacity. The sensitivity of (14)C detection was improved by the use of a variable scintillation flow achieved via a simple modification to the classical online radiochemical detection set-up. A modification of the flow-through cell design in which internal diameter of the tubing was reduced further increased the sensitivity and resolution by decreasing peak tailing. The injection of relatively large injection volumes was made possible by the use of columns packed at ultra-high pressure with 2.2 microm particles. Because of the reduced back pressure generated using these larger particle sizes, two 150 mm x 3 mm columns could be coupled, allowing 4-fold larger injection volumes and a 50% increase in theoretical plate number at a similar back pressure compared to a standard 150 mm x 2.1mm Waters UPLC column. The value of the methodology described was demonstrated by the analysis of in vitro and in vivo metabolism samples of (3)H- and (14)C-labeled compounds and compared with conventional radio-HPLC. We have shown that metabolite separation can be achieved with increased efficiency while maintaining a sensitivity comparable to that of conventional HPLC/RAD.     

Automated semipreparative purification with mass spectrometric fraction collection trigger: modeling and experimental evaluation of a setup employing passive splitting

A semipreparative HPLC setup was evaluated for automated fractionation with both photometric- and mass spectrometric trigger. The goals of the work were to systematically study and optimize the flow-splitting setup for mass-directed purifications by mathematical modeling and experimental verification. The system comprised a passive splitting device with make-up flow capability, which directed a small fraction of the column effluent to the mass spectrometer and the remainder to the fraction collector. Tubing lengths and diameters of the splitter as well as make-up flow rates were varied in order to address and optimize peak dispersion, delay times between mass detector and fraction collector, and mass spectrometric signal quality. A paraben standard mixture was analyzed and purified on both microparticulate and monolithic columns with 10 mm inner diameter and at typical flow rates of 5-10 mL/min. Fraction purities and recoveries close to 100% were achieved. The system allowed mass-triggered fractionations on a 1 mg scale at flow rates of 10 mL/min in combination with monolithic columns in less than 2 min. Finally, the system was successfully applied to the fully automated isolation of milligram quantities of degradation products in a pharmaceutical preparation to successfully allow for structure elucidation with NMR spectroscopy.     

Peak tailing and column radial heterogeneity in linear chromatography

The correlation between the radial heterogeneity of a column and the tailing of the elution profiles of chromatographic peaks was studied using a numerical method. A parabolic distribution of the linear flow velocity of the mobile phase and of the column efficiency in the radial direction were assumed. Moment analysis showed that peak tailing takes place under such experimental conditions and that it increases with increasing range of radial variations of the flow velocity and the column efficiency. It was also found that the higher the column efficiency, the larger the effect of a given degree of radial heterogeneity on the extent of peak tailing. Peak tailing behavior of columns having different efficiencies could be correlated with each other by an equation. Some characteristic features of tailing peaks were analyzed in connection with the column radial heterogeneity.     

High-Speed Large Scale Chromatographic Purification of Plasmid DNA with a Novel Giant-Pore Stationary Phase

A rigid spherical giant-pore poly (glycidyl methacrylate-co-ethylene dimethacrylate) matrix has been prepared by radical suspension–polymerization on the basis of a novel porogenic mode using superfine particles of calcium carbonate as a solid porogen. Scanning electron microscopy reveals that the bead has pores as large as 10 μm. The hydrodynamic properties show that this polymeric material has good strength and a low back pressure of 1.0 MPa at a flow velocity of 3,000 cm h⁻¹. After being modified to be an anion-exchange material, high dynamic binding capacity of plasmid DNA of above 1,000 μg plasmid per mL of bed by a column of this material, could be obtained comparing to the 150 μg plasmid per mL of bed with a Q-Sepharose FF column at the same flow rate. Large-scale preparative plasmid separations (2–20 mL) from cell lysate were investigated. A 75% yield and 94.9% purity of SC plasmid DNA were obtained by a 20 mL column of giant-pore beads at a flow rate of 600 cm h⁻¹.     

Columns switch recycling size exclusion chromatography for high resolution protein separation

Columns switch recycling size exclusion chromatography (csrSEC) was proposed to achieve high resolution protein separation with good biocompatibility. Proteins were firstly separated by two serially coupled SEC columns, and fractions were in sequence switched back to the first column by two-position valves for further separation in terms of close-loop recycling until satisfactory resolution was achieved. Compared to SEC, the separation window was broadened by increasing column length via cycling without further increase on back pressure. Compared to recycling SEC (rSEC), the overtaking of later eluted components by early eluted ones after several cycles could be avoided for complex sample analysis, by parking fractions in the second SEC column before transferred in turn back to the first one for cycling ordinally. In our experiments, the baseline separation of five proteins with molecular weight ranging from 10 kDa to 80 kDa was achieved by csrSEC. Furthermore, a multidimensional csrSEC–RPLC platform was constructed, and peak capacity up to 3600 was obtained for protein separation. All these results demonstrated that csrSEC is a promising protein separation mode with good biocompatibility, broadened separation window and improved resolution.     

Synthesis of Monodisperse Poly(chloromethylstyrene divinylbenzene)Particle and Its Applications

Poly(styrene-divinylbenzene)(PS-DVB)particlesarethemostcommonlyusedpolymericchromatographicstationaryphase.However,theirhydrophobicnaturelimitstheirusageinbiochemicalseparations.ChemicalmodificationofPS-DVBmaybeusedtoincreasetheirpolarityfortheseparationofbiomacromolecules.Becausepoly(chloromethylstyrenedivinylbenzene)(PS-CH2Cl)containsreactivechloromethylgroups,ithasmanyadvantagesoverPS-DVBinchemicalmodification.However,untilnow,theyhavebeenpreparedbysuspensionpolymerization.Thismeth…     

Supercritical fluid chromatography with a slurry-packed capillary column

A versatile system with a slurry-packed capillary column was developed for supercritical fluid chromatography, which is capable of programming both inlet and outlet pressure independently, as well as using a restrictor to apply back pressure. This system revealed the relationships between pressure drop, flow rate, and linear velocity in pressure-programmed supercritical fluid chromatography. In the restrictor system, both the pressure drop and the flow rate increased almost linearly with inlet pressure, while under conditions of constant pressure drop characteristic behavior was observed which depended on the density-viscosity relationships of supercritical fluid. Resolution in the separation of polysiloxane oligomers was found to be increased by increasing the ratio of pressure drop to pressure-programming rate, although the sensitivity decreased due to the increase in peak volume. The system controlling both inlet and outlet pressure has distinct advantages over the restrictor system controlling both inlet and outlet pressure has distinct advantages over the restrictor system in practical in practical operations.     

硅胶整体柱的制备及对苯取代物的分离

研究了一种制备硅胶整体柱的新方法。该工艺以四甲氧基硅烷和聚乙二醇为原料,采用改进的溶胶—凝胶技术,建立了消除柱体开裂和变形弯曲的方法及中孔的构建方法,考察了流速对柱压降的影响,并在正相色谱模式下应用制备的硅胶整体柱对苯取代化合物进行了分离。结果表明采用新工艺所制备的硅胶整体柱不开裂,不变形,并具有通孔和中孔双孔结构,渗透性强,柱压降低,机械强度高,柱效高等特点。这些特性尤其适合化合物的高效快速分离。     

A new approach to live reaction monitoring using active flow technology in ultra‐high‐speed HPLC with mass spectral detection

A new type of chromatography column referred to as a parallel segmented flow (PSF) column enables ultra‐high‐speed high‐performance liquid chromatography‐MS to be undertaken. This occurs because the separation efficiency obtained on PSF columns has been shown in prior studies to be superior to conventional columns, and the flow stream is split radially inside the outlet end fitting of the column, rather than in an axial post‐column flow stream split. As a result, the flow through the column can be five times higher than the flow through the MS. In this work, the degradation of amino acids in dilute nitric acid was used to illustrate the process. Separations were obtained in less than 12 s, although the reinjection process was initiated 6 s after the previous injection. The degradation rate constant of tryptophan, in the presence of tyrosine and phenylalanine, was determined. Copyright © 2015 John Wiley & Sons, Ltd.     

Improving splitless injection with electronic pressure programming

An experimental injection port has been designed for split or splitless sample introduction in capillary gas chromatography; the inlet uses electronic pressure control, in order that the column head pressure may be set from the GC keyboard, and the inlet may be used in the constant flow or constant pressure modes. Alternatively, the column head pressure may be programmed up or down during a GC run in a manner analogous to even temperature programming. Using electronic pressure control, a method was developed which used high column head pressures (high column flow rates) at the time of injection, followed by rapid reduction of the pressure to that required for optimum GC separation. In this way, high flow rates could be used at the time of splitless injection to reduce sample discrimination, while lower flow rates could be used for the separation. Using this method, up to 5 μl of a test sample could be injected in the splitless mode with no discrimination; in another experiment, 2.3 times as much sample was introduced into the column by using electronic pressure programming. Some GC peak broadening was observed in the first experiment.     

Novel separation medium spongy monolith for high throughput analyses

Sponge-like material was utilized as novel chromatographic media for high throughput analyses. The pore size of the sponge-like material was several dozen micrometer, and was named spongy monolith because it consists of continuous structured copolymers, which was made of poly(ethylene-co-vinyl acetate), such as monolithic materials including silica monoliths and organic polymer monoliths. The spongy monolith was packed into a stainless steel column (100 mm × 4.6 mm I.D.) and evaluated in liquid chromatography (LC) with an on-line column-switching LC concentration system. The results indicate that the packed column could be used with high flow rates and low back pressure (9.0 mL/min at 0.5 MPa). Furthermore, bisphenol A was quantitatively recovered by on-line column-switching LC concentration with the spongy monolithic column. Additionally, the adsorption capacity and physical strength of the media was enhanced via chemical modification of spongy monoliths using glycerol dimethacrylate. The results compared with original spongy monolith demonstrated that a higher adsorption capacity was achieved on a shorter column, and a stable low back pressure was obtained at high throughput elution even with a longer column.