Refractometer, flow-differential

Other anomalies were observed, since Beer s law was fulfilled at 220 nm and between 0 and 2 g/L and not at other wavelengths. Considering the possibility of unattended anomalies of the HEC after fractionation, we had to abandon this detection method. For this reason, for the continuous concentration determination, we used a flow-differential refractometer (type R403, Waters Associates, Milford, MA, USA) connected to an integrating recorder. Since there is no monochromator on the apparatus, it was not possible to use the refractive index increment value used for the light-scattering measurements. 

X 0.75 cm) Ve i = 28 ml = 50 ml eluent 0.05 M NaCI flow rate 0.80 ml/min detection Optilab 903 interferometric differential refractometer applied sample mass/volume 200 /tl of 2-mg/ml aqueous solutions sum of individual chromatograms (theory —) and (theory/experimental) ratio (—) plotted for quantification of deviations in separation performance between narrow distributed samples and broad distributed samples. 

SEC measurements were made using a Waters Alliance 2690 separation module with a 410 differential refractometer. Typical chromatographic conditions were 30°C, a 0.5-ml/min flow rate, and a detector sensitivity at 4 with a sample injection volume of 80 fil, respectively, for a sample concentration of 0.075%. All or a combination of PEO standards at 0.05% concentration each were used to generate a linear first-order polynomial fit for each run throughout this work. Polymer Laboratories Caliber GPC/SEC software version 6.0 was used for all SEC collection, analysis, and molecular weight distribution overlays. 

Hie hydrolytic depolymerization of nylon-6 was followed by gel permeation chromatography (GPC), viscometry, and gravimetry. GPC determinations were performed on a Waters 150C chromatography system using benzyl alcohol as die eluant, two Plgel 10-p.m crosslinked polystyrene columns, and a differential refractometer detector. The flow rate was 1 mL/min. The concentration of the polymer solutions was 0.5 wt% and dissolution was accomplished at 130°C. 

Gel Permeation Chromatography. The instrument used for GPC analysis was a Waters Associates Model ALC - 201 gel permeation chromatograph equipped with a R401 differential refractometer. For population density determination, polystyrene powder was dissolved in tetrahydrofuran (THF), 75 mg of polystyrene to SO ml THF. Three y -styragel columns of 10, 10, 10 A were used. Effluent flow rate was set at 2.2 ml/min. Total cumulative molar concentration and population density distribution of polymeric species were obtained from the observed chromatogram using the computer program developed by Timm and Rachow (16). 

The second most widely used detector in HPLC is the differential refractometer (RI). Being a bulk property detector, the RI responds to all substances. As noted in Table 3 the detection limits are several orders of magnitude higher than obtained with the UV detector. Thus, one turns to the RI detector in those cases in which substances are non-UV active, e.g. lipids, prostaglandins. In addition, the RI detector finds use in preparative scale operation. Finally, relative to the UV detector, the RI is significantly more temperature and flow sensitive and cannot be used in gradient elution. 

The fluid from the tube or the core leaves the valve through port 5 and enters the inlet of the sample cell of the differential refractometer (made by Knauer of West Germany). The residue flows out of the sample cell to the waste. The reference cell contains... 

Chromatographic System. The isocratic liquid chromatograph used was a Waters Associates (Milford, MA) Model 24A alc which included a Model 6000A Solvent Delivery System, a Model 401 Differential Refractometer and a Model 440 Absorbance Detector operating at 254 nm and was fitted with a WISP automatic injector. The analog outputs of the UV absorbance detector or differential refractometer were recorded with a Model 730 Data Module (printer, plotter, integrator)(Waters). Eluent flow rate was 1.0 ml/min unless otherwise noted. 

On-line size exclusion chromatographic (SEC) analyses were performed with a Waters Model 401 differential refractometer (DR), a Waters Model 480 ultraviolet (UV) variable wavelength spectrophotometer and a Foxboro Miran lA infrared (IR) photometer, equipped with a zinc selenide ultramicro flowcell of 1.5 mm nominal pathlength and 4.5 /xl volume, purchased from the same supplier. A set of ten Mycrostyra-gel (Waters Associates) columns, regenerated by Analytical Sciences Inc. (ASI) and of nominal porosities 100, 500 (two) 10 (two), 10 (three), 10 and lO X, in the order given and a mobile phase flow rate of 1 ml/min was used. The column set had a specific resolution of 19.7 in 1,4-dioxane, as determined by the method of Yau(2). 

A gSe of two Waters ultrastyragel columns, designated 10 A and 10 A and a Waters pump (Model 590) for HPLC were used in this study. The elution solvent was tetrahydrofuran (THE) which was distilled in the presence of a small amount of CaH in order to remove the peroxide. The flow rate was maintained at 1 ml/min. The sample injection volume was -30 pi. The chromatogram detected by the differential refractometer (Waters R401) was recorded on a strip chart recorder. All experiments were performed at room temperatures with concentrations below the over-loading condition. 

High Performance Size Exclusion Chromatography. SEC was carried out on polystyrene-divinylbenzene gels with porosites ranging from 100 to 104A. The solvent, THF, had a flow rate of 1 ml/min. The detector was a differential refractometer. 

The weight average molecular weight(Mw) measurements were determined by utilizing the Chromatix low angle laser light scattering G.P.C. detector (14). The G.P.C. column set was similar to those mentioned above and the carrier solvent was tetrahydrofuran at 25°C with a flow rate of 1.5 ml/min. The (dn/dc) values of both polybutadiene and polyisoprene were determined in THF with the Chromatix KMX-16 differential refractometer. 

A Waters Associates Anaprep GPC fitted with one 4 ft X 2.4 inches od Styragel column having a nominal porosity of 104 A was used for the preparative fractionation of the PMMA blend in tetrahydrofuran at a temperature of 25 °C and at a flow rate of 30 ml/min. The degasser and differential refractometer were operated at 35° and 25 °C, respectively. Samples having concentrations of 0.25 wt-vol % were respectively, automatically injected from a 100 ml loop over a 5-minute period. Ten 125 ml fractions were automatically collected for each sample injection. Upon... 

Sugar and polyols in aqueous media were analyzed by HPLC. (column, sugar pack waters eluant H O flow rate 0.5 ml/min. temperature 90°C differential refractometer detector). 

A preparative chromatograph fitted with an ST/2000-1 guard column (IX 2-in. ID) and an ST/2000B preparative column (25 X 2-in. ID), each packed with ST MACROBORE Cl8 (25-/xm particle size), was employed for separation of esters. The system was equipped with a differential refractometer with a flowing reference (as opposed to an air-filled reference) in order to minimize baseline drift. 

An RP-HPLC procedure for the analysis of w-3 fatty acid esters that could eventually be adapted to the preparative isolation of quantities of EPA and DHA was also developed by Beebe et al. (48). The separations were performed at room temperature on an ODS-3 RAC II column (100 X 4.6-mm ID, 5-/zm particle size) with a guard column (30 X 4.6-mm ID dry-packed with SUPELCOSIL LC-18 40-yttm material). The esters were eluted with a mobile phase of acetonitrile/tetrahydrofuran/water (9 5 11) at a flow rate of 2.0 ml/min (detector differential refractometer). 

Procedure (See Chromatography, Appendix BA.) Use a suitable high-performance liquid chromatograph equipped with differential refractometer, autosampler injection unit, mobile-phase degasser, column heating block or oven, and a computing integrator. The column is Lichrosorb RP-18 250-mm x 4.5 mm (id) (GL Science, Inc., or equivalent) and YMC-Pack ODA-A A-303 250-mm x 4.5 mm (id) (YMC Company, Ltd., or equivalent) connected in a series, or equivalent, maintained at 50°. Use 80 20 acetone acetonitrile as the eluent, at a flow rate of 2 mL/min. 

Chromatographic System Determine as directed under Chromatography, Appendix HA, but use a liquid chromatograph equipped with a differential refractometer detector and a 30-cm x 7.8-mm (id) column packed with 25-pm diameter beads of silver bonded to sulfonated divinyl benzene-styrene copolymer (Aminex HPX-42A, Bio-Rad Laboratories, or equivalent). Maintain the column at a constant temperature of 65° 10°, and the flow rate at 0.3 to 1.0 mL/min. Use deionized water as the mobile phase. 

Differential refractometer detectors measure the difference between the refractive index of the mobile phase alone and that of the mobile phase containing chromatographed compounds as it emerges from the column. Refractive index detectors are used to detect non-UV absorbing compounds, but they are less sensitive than UV detectors. They are sensitive to small changes in solvent composition, flow rate, and temperature, so that a reference column may be required to obtain a satisfactory baseline. 

Conventional Size Exclusion Chromatography, SEC, was performed on a Spectra Physics apparatus with two PL gel columns (5 pm particle size, 300 mm length, one with 50 A and one with 100 A pore size) and a Styragel HR2 column (7.8mm internal diameter x 300mm length). The detection was achieved with a SP8430 differential refractometer. The tolnene was elnted at a flow rate of 0.8mL.min . 

Figure 9.72 Chromatograms of the action patterns of maltoheptaose after the indicated periods of incubation with a-amylase and a-glucosidase. Peaks 1, glucose 2, maltose 3, maltotriose 4, maltotetraose 5, maltopentaose (x) compound A 6, maltohexaose 7, maltoheptaose. (A) Pure maltoheptaose used for the assay. (B) Blank sample before the addition of substrate. (C-H) Chromatograms after 1, 5, 10,15, 20, and 30 minutes, respectively, of incubation. Chromatographic conditions column, 10 jum Nucleosil SA (250 mm X 4 mm) solvent, acetonitrile-water (72.527.5) flow rate, 0.7 mL/min temperature, 27°C detection, differential refractometer, full scale = 2 X 10-6 refractive index units. (From Haegel et aL, 1981.)...

The initial pulse of polymer solution which was injected into the column entry becomes diluted and attenuated as the different species are separated on the gel packing. The column effluent is monitored by detectors which respond to the weight concentration of polymer in the flowing eluant. The most common detector is a differential refractometer. Spectrophotometers, which operate at fixed frequencies, are also used as alternative or auxiliary detectors. Some special detectors which are needed particularly for branched polymers or copolymers are mentioned in Section 3.4.4. 

The opportunity to measure the dilute polymer solution viscosity in GPC came with the continuous capillary-type viscometers (single capillary or differential multicapillary detectors) coupled to the traditional chromatographic system before or after a concentration detector in series (see the entry Viscometric Detection in GPC-SEC). Because liquid continuously flows through the capillary tube, the detected pressure drop across the capillary provides the measure for the fluid viscosity according to the Poiseuille s equation for laminar flow of incompressible liquids [1], Most commercial on-line viscometers provide either relative or specific viscosities measured continuously across the entire polymer peak. These measurements produce a viscometry elution profile (chromatogram). Combined with a concentration-detector chromatogram (the concentration versus retention volume elution curve), this profile allows one to calculate the instantaneous intrinsic viscosity [17] of a polymer solution at each data point i (time slice) of a polymer distribution. Thus, if the differential refractometer is used as a concentration detector, then for each sample slice i. 

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