Filters Filtering, mobile phase

Pump a filtered mobile phase of 20% methanol in distilled, deonized water over the column at half of the recommended maximum flow... 

Filtering mobile phase prevents often replacing inlet frits and check valves. Degassing mobile phase prevents bubble formation. 

Give one reason why it is necessary to filter mobile phase solutions before use on an HPLC system. 

The dn/dc measurement should be carried out under the same temperature (7) and same wavelength (A.) as the LALLS experiment and at a constant chemical potential (p.). The conditions for a constant chemical potential can be achieved by dialyzing the polymer solution against the filtered mobile phase until the dn/dc of the polymer solution becomes constant. In addition, the final concentration of the polymer solution should be... 

In the case of thin-layer chromatography there is frequently no wait to establish complete equilibrium in the chamber before starting the development. The chamber is usually lined with a U-shaped piece of filter paper and tipped to each side after adding the mobile phase so that the filter paper is soaked with mobile phase and adheres to the wall of the chamber. As time goes on the mobile phase evaporates from the paper and would eventually saturate the inside of the chamber. 

Errors in the molecular weight data from HPSEC are usually due to improperly prepared samples, column dispersity, or flow rate variations. The sample to be analyzed should be completely dissolved in the mobile phase and filtered prior to injection onto the column. A plugged column inlet frit will invalidate results. In addition, do not load the column with excess sample. Column overloading affects the accuracy of data by broadening peaks, reducing resolution, and increasing elution volume. For best results, the concentration of the injected sample should be as low as possible while still providing adequate... 

Commercial grades of PVP, K-15, K-30, K-90, and K-120 and the quaternized copolymer of vinylpyrrolidone and dimthylaminoethylmethacrylate (poly-VP/ DMAEMA) made by International Specialty Products (ISP) were used in this study. PEO standard calibration kits were purchased from Polymer Laboratories Ltd. (PL), American Polymer Standards Corporation (APSC), Polymer Standards Service (PSS), and Tosoh Corporation (TSK). In addition, two narrow NIST standards, 1923 and 1924, were used to evaluate commercial PEO standards. Deionized, filtered water, and high-performance liquid chromatography grade methanol purchased from Aldrich or Fischer Scientific were used in this study. Lithium nitrate (LiN03) from Aldrich was the salt added to the mobile phases to control for polyelectrolyte effects. 

Ascending, one-dimensional development in a trough chamber with chamber saturatioiL After sample application the HPTLC plate was preconditioned for 30 min in the mobile phase-free trough of a twin-trough chamber (without filter paper lining). 

Samples with higher protein levels (yogurts), are initially treated with hydrochloric acid and after protein precipitation the supernatant is filtered and injected into the HPLC column. The separations performed with a LiChroCART RP18 column used a mixture of acetonitrile and formic acid as the mobile phase. A baseline quantification of the carminic acid was possible in the presence of other coloring agents, with excellent recuperation, selectivity, accuracy, and precision. ... 

For preparative separation, the mobile phase can be selected by performing preliminary analytieal TLC experiments. In PLC, the chromatographic chamber has to be saturated within 2 h beeause the development of preparative plates is much slower than the analytical development. In the analytical preassay during the selection of mobile phase composition, the chromatographic chamber must be hned with a sheet of filter paper to obtain a saturated atmosphere with mobile phase vapor. Then, the optimized analytical mobile phase can be transferred imchanged to preparative separations in the saturated developing chamber. 

Chamber saturation is recommended for better reproducibility of the separation — especially if multicomponent mobile phase mixtures are composed of solvents differing in volatility or polarity to a great extent. Moreover, chamber saturation can improve resolution of two components or reduce the formation of secondary fronts. For chamber saturation, the large tank sides are lined with a sheet of filter paper 20 X 20 cm each. Dnring the filling of the mobile phase into the chamber, it is poured onto the filter, which is then completely wetted and soaked by the mobile phase. Note that the wet filter paper is dipped into the mobile phase at the trough bottom. The prepared closed tank will become satnrated within 15 to 30 min depending on the volatihty of the solvent components (withont wetted filter paper it needs more... 

FIGURE 6.9 Horizontal chamber for fully online HPTLC by Nyiredy 1 — evaporator, 2 — diode-array detector, 3 — quartz glass cover plate, 4 — septum, 5 — injector block, 6 — mobile phase, 7 — filter paper, 8 — Teflon chamber, 9 — chromatoplate. (Modified from Nyiredy, Sz., J. Planar Chromatogr. 15, 454-457, 2002.)... 

The testing of impnrities in active pharmacentical ingredients has become an important initiative on the part of both federal and private organizations. Franolic and coworkers [113] describe the utilization of PLC (stationary phase — silica gel and mobile phase — dichloromethane-acetonitrile-acetone (4 1 1, v/v)) for the isolation and characterization of impurities in hydrochlorothiazide (diuretic drug). This drug is utilized individually or in combination with other dmgs for the treatment of hypertension. The unknown impurity band was scraped off the plate and extracted in acetonitrile. The solution was filtered and used for LC/MS and NMR analysis. The proposed procedure enabled the identification of a new, previonsly nnknown impurity. It was characterized as a 2 1 hydrochlorothiazide-formaldehyde adduct of the parent drug substance. 

Mobile phase The HPLC mobile phase is made up as follows. Prepare 2 L of acetate buffer by dissolving 13.6 g of sodium acetate and 6 mL of glacial acetic acid in 2 L of deionized water. Adjust the solution to pH 4.8 with concentrated sodium hydroxide solution (or glacial acetic acid) if necessary. Mix 2 L of buffer with 1.6-2 L (the amount depends on the particular commodity) of methanol. Eilter the solution through a 0.22-pm Nylon 66 filter membrane before using the mobile phase Absolute ethanol Aaper Alcohol and Chemical Co. (200 proof)... 

HPLC allows a quantitative determination with relatively simple extractions. In many cases, extraction only involves a heating of the commodity with water, followed by filtration and injection onto an HPLC column. In the determination of caffeine, theobromine, and theophylline in cocoa, coffee, or tea, as well as in other foods, there is scarcely a month that passes without a new paper on this assay. Kreiser and Martin provide typical conditions for analysis.28 In their studies, samples were extracted in boiling water and filtered prior to injection onto the HPLC column. The HPLC conditions used a Bondapak reversed phase column and a mobile phase of water methanol acetic acid (74 25 1) with detection at 280 nm. This method is accurate, precise, and conserves time. It has also been adopted by the AOAC as an official method for the determination of theobromine and caffeine in cocoa beans and chocolate products.29... 

A French Standard, which corresponds to ISO 10095 issued in 1992, specifies a method for determination of caffeine in green or roasted coffee or in coffee extracts (decaffeinated or not).33 Caffeine is extracted with water at 90°C in the presence of MgO. The extract is filtered, then cleaned-up on a mini-column packed with a silica phenyl group derivative, and analyzed by HPLC on a C18 column with a methanol/water (30 70) mobile phase and a UV detector operating at 254 to 280 nm. 

High Performance Liquid Chromatographic (HPLC) Analysis. A Waters HPLC system (two Waters 501 pumps, automated gradient controller, 712 WISP, and 745 Data module) with a Shimadzu RF-535 fluorescence detector or a Waters 484 UV detector, and a 0.5 pm filter and a Rainin 30 x 4.6 mm Spheri-5 RP-18 guard column followed by a Waters 30 x 3.9 cm (10 pm particle size) p-Bondapak C18 column was used. The mobile phase consisted of a 45% aqueous solution (composed of 0.25% triethylamine, 0.9% phosphoric acid, and 0.01% sodium octyl sulfate) and 55% methanol for prazosin analysis or 40% aqueous solution and 60% methanol for naltrexone. The flow rate was 1.0 mL/min. Prazosin was measured by a fluorescence detector at 384 nm after excitation at 340 nm (8) and in vitro release samples of naltrexone were analyzed by UV detection at 254 nm. 

The small particle size of the packing makes 3x3 columns susceptible to plugging problems. Samples and mobile phases should be filtered before use, and the column should be protected with guard and scavenger columns (see Section 5.3.2). 

A microparticulate hplc column is a very efficient filter, and if the mobile phase contains any particulate matter, or acquires it from the pump and/or the injection valve due to wear, it will collect at the top of the column. If this happens, the pressure drop across the column for a given flow will gradually increase, and the column may eventually become completely blocked. To prevent this happening, the mobile phase should always be filtered before use, preferably through a 0.5 p,m porosity filter, and guard and scavenger columns should be used as a matter of routine (see Section 5.3.2). 

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