Chromatographic peak testing

Results are presented in the percentage. Percentage of single chromatographic peak areas was measured on the basis of area of the single peaks to the total peak area ratio. Selected results were statistically evaluated using a t-test at the 0.05 level. The intervals of significance are presented in tables. 

Peak purity tests are used to demonstrate that an observed chromatographic peak is attributable to a single component. Mass spectrometry is the most sensitive and accurate technique to use for peak purity evaluation because of the specific information derived from the analysis. However, a good number of HPLC methods use mobile phase conditions that are incompatible with mass spectrometry detection. In this case, PDA spectrophotometers using peak purity algorithms may be used to support the specificity of the method. Almost all commercially available diode array detectors are equipped with proprietary software that will perform these calculations. Although this technique is more universal in application to HPLC methods, the data provided is neither particularly... 

Before the ruggedness test could be contemplated it was essential to fully validate the method with respect to other method characteristics. Hence the following tests were carried out, specificity, spectral purity of chromatographic peaks, linearity of detector response, and repeatability over 100 injections. Satisfactory results were achieved for all these experiments before we continued to the ruggedness test. 

Purity of EGDN. The area under the gas chromatographic peak for acetone shall be less than 0.6% of the area for EGDN. The sum of the areas of all other impurity gas chromatographic peaks shall not exceed 0.05%°f the area of the peak of EGDN. Test as specified in 4.3.2... 

Critical separations in chromatography should be investigated at an appropriate level. For critical separations, selectivity can be demonstrated by the resolution of the two components that elute closest to each other. Peak purity tests using diode array or mass spectrometric detectors may be useful to show that the analyte chromatographic peak is not attributable to more than one component. 

Reliable quantitative results are obtained by external calibration if automatic injectors or autosamplers are used. This method involves direct comparison of the peak responses obtained by separately chromatographing the test and reference standard solutions. If syringe injection, which is irrepro-ducible at the high pressures involved, must be used, better quantitative results are obtained by the internal calibration procedure where a known amount of a noninterfering compound, the internal standard, is added to the test and reference standard solutions, and the ratios of peak responses of the analyte and internal standard are compared. 

Fritted-glass crucible A filtering crucible equipped with a porous glass bottom also called a sintered-glass crucible. Fronting Describes a nonideal chromatographic peak in which the early portions tend to be drawn out compare with tailing. F-test A statistical method that permits comparison of the variances of two sets of measurements. 

Vouros and co-workers recently demonstrated the effectiveness of nanospray to improve both the S/N quality concurrent with a reduction in matrix effects such as ion suppression [102]. Rather than use a nanobore LC column, the authors constructed a unique postcolumn flow that allowed for nanospray flow rates with conventional column formats. A cylindrically symmetric flow splitter was shown to preserve chromatographic peak shape at split ratios as high as 2000 1 thus, effluent from a conventional 2.1-mm Cl8 column at 200 pi. /min was reduced to 100 nL/min. The heart of the flow splitter was a fused-silica needle emitter with an orifice diameter of 5 pm. Tlie nanoflow splitter was applied to the in vitro metabolite analysis of a test... 

A comprehensive study of coated anion-exchange resins provides some valuable insights [20]. Of several coating materials tested, cetyl pyridinium chloride (CPI) was found to be the most satisfactory. Static coating was employed with 1-3 x I0 M CPCI in 3 % (v/v) acetonitrile-97 % water. Coating of a polyacrylate resin (Rohm Haas XAD-8) gave sharper chromatographic peaks for anions than the more hydro-phobic PS-DVB resin (Hamilton PRP-1). 

The efficiency of chromatographic columns is usually measured under conditions of linear adsorption equilibria from pulse tests. The Height Equivalent to a Theoretical Plate (HETP) is calculated from the first moment /ij and the variance of a chromatographic peak obtained in a column of length I by ... 

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