Peak areas manual methods

B. Peak Area Manual methods for converting the area of the detector signal into numerical data historically have included (1) planimetry, (2) taking the product of peak height and peak width at half-height, (3) triangu-... 

Chromatographic peak areas were measured by both manual and mechanical integration methods. Total analysis time was less than one hour per sample. 

Acceptable hnearity, reprodudhihty and recovery was shown for the two active substances. Either peak-area or peak-height data may be used for quantification. Comparison of the manual and semi-automated dissolution tests have shown equivalency of the two methods. 

The manual triangulation method used to measure peak areas is no longer in use. However, it is useful to remember that for a Gaussian eluting peak, the product of its width at half height by its height corresponds to approximately 94% of the total area of the peak. 

Alternatively, peaks areas can be defined manually and the entire volume of the peak integrated. This method is more tolerant of imperfect gels. However, because the bands in native gels are often broad, it is critical to set a uniform criterion (such as pixel intensity) for defining peak boundaries and... 

The filter disc is transferred to a stopperred flask containing ethanol and propan-2-ol, and shaken to extract the water present which is then determined by gas-liquid chromatography the quantitity of water present is calculated from the ratio of the areas of the peaks for water (unknown) and ethanol (internal standard). The alkaloids are extracted from the filter disc using sulphuric acid and determined by a specially developed autoanalyser procedure based on the Koenig reaction this is shown schematically in Fig. 21. Since the procedure was developed specially for this application, results obtained from it were compared closely with those produced by the traditional manual method, a steam distillation technique, before it was adopted as a standard method. 

The methods of quantitative analysis are essentially tiiose inherited from gas chromatography. Peak hei t or pe area can be measured, either manually or with electronic devices. Peak height measiu-ements have the advantage of simplicity but are sensitive to changes in peak shape peak area measurements should always be used where peaks are broad and tailing. 

The DryLab model utilized in Waters AMDS has additional requirements The number of sample components should not exceed 12 peak area% should be greater than 1%. These requirements are necessary to achieve greater prediction accuracy only. Any discrepancies could be corrected manually in DryLab using the data entry screen by manually entering the retention of the components from the scouting runs (to assign the peaks with a certain number). DryLab has been used for the method development of model drug candidates... 

In contrast to the IS method, external standardization may be used in which several standard solutions of varying concentrations of the sample are prepared. Following constant volume injection of each standard solution, a plot of peak area (or height) versus concentration is made, and unknown sample concentrations are obtained from interpolation of the calibration curve. The success of this technique, however, is dependent upon the precision of injection volume, readily accomplished with automatic injection but less so when manual microliter syringes are used. 

The most accurate manual method to determine the area of a skewed peak is to construct a trapezoid. This is easy if you use a template (included), and harder if you do not. Refer to Figure 20-55. Determine the height as shown. You need two widths, one at 15% and the other at 85% of the height. 

What is the most accurate manual method to determine the area of a chromatographic peak ... 

Comparison of manual methods for determining peak areas... 

If the baseline shows any drift, peak heights are often more correctly determined manually than by an automatic system (important for trace analysis). The peak area is very seldom calculated manually, and if it is, a peak area equivalent is used, which is the product of peak height and peak width at baseline. In older publications, the peak area was often calculated by cutting out the peak and weighing it [88). This method also gave very good results. 

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