Peak Width Calibration

Peedc-to-peak resolution in SEC can be calculated by the ratio of peak separation at the peak maxiaut to the sum of the baseline peak widths. This general definition of resolution is less useful in SEC, where a measure of the ability of the column to separate solutes of different molecular weight is required. For this purposes, we define a new term, the specific resolution factor, R, which relates peak resolution to sample molecular weight, assuming all measurements are made within the linear region of the molecular weight calibration curve, equation (4.41)... 

Tables 6.27 and 6.31 show the main characteristics of ToF-MS. ToF-MS shows an optimum combination of resolution and sensitivity. ToF-MS instruments provide up to 40000 spectra s-1, a mass range exceeding 100000 (in principle unlimited), a resolution of 5000, and peak widths as short as 200 ms. This is better than quadruples and most ion traps can handle. Unlike the quadrupole-type instrument, the detector is detecting every introduced ion (high duty factor). This leads to a 20- to 100-times increase in sensitivity, compared to QMS used in scan mode. The mass range increases quadratically with the time range that is recorded. Only the ion source and detector impose the limits on the mass range. Mass accuracy in ToF-MS is sufficient to gain access to the elemental composition of a molecule. A single point is sufficient for the mass calibration of the instrument. ToF mass spectra are commonly calibrated using two known species, aluminium (27 Da) and coronene (300 Da). ToF is well established in combination with quite different ion sources like in SIMS, MALDI and ESI.

Vinas et al. [47] determined penicillamine routinely by using batch procedures and FIA. A capsule was dissolved in water, diluted to 250 mL, and a suitable portion of the solution treated with 1 mM Co(II) solution (2.5 mL) and 2 M ammonium acetate (2.5 mL). The mixture was diluted to 25 mL and the absorbance of the yellow complex was determined at 360 nm. Calibration graphs were linear for 0.02-0.3 mM of penicillamine. The method was modified for flow injection analysis using peak-height or peak-width methods, but in both cases the flow rates were maintained at 3.3 mL/min. For the peak-height technique, calibration graphs were linear for 0.1-2 mM, and the sampling frequency was 150 samples per hour. For the peak-width method, the response was linear for 50 pM to 0.1 M, and this method was particularly useful for routine determinations. 

Calibration parameters are instrument parameters whose values do not vary with the type of experiment, such as peak widths, peak shapes, mass assignment, and resolution versus sensitivity. 

The ELS detector was previously also referred to as a mass detector, pointing to the fact that the response is (mainly) determined by the mass of the sample rather than by its chemical structure. Van der Meeren et al., though, demonstrated that the ELSD calibration curves of phospholipid classes were also dependent on the fatty acid composition (52). The dependence on the fatty acid composition is, however, completely different in nature and much less pronounced than for UV detection. The reason for this behavior is to be found in the partial resolution of molecular species, even during normal-phase chromatography. Thus, the peak shape depends not only on the chromatographic system but also on the fatty acid composition and molecular species distribution of the PL sample (47). Because it was shown before, based on both theoretical considerations and practical experiments, that the ELS detector response is generally inversely proportional to peak width (62,104), it follows that the molecular species distribution of the PL standards used should be similar to the sample components to be quantified. It was shown that up to 20% error may be induced if an inappropriate standard is used (52). 

When you are in a clinical environment where reproducibility is critical, you need to know and understand the variables that the calibration selects. An integrator should be able to integrate close neighbors about four times more accurately than you can by hand. The integrator sets three variable levels peak width, slope rejection, and noise rejections. It also makes decisions on how to integrate unresolved peaks. As peaks widen (or narrow) in later parts of the chromatogram, the integrator doubles (or halves) the peak width value to include the whole peak. 

From the calibration curve, the resolution power of the column set may also be evaluated. Resolution between two adjacent peaks, 1 and 2, is defined in terms of their retention volumes, V, and peak widths, w ... 

Processing method Controls and documents integration parameters, component names, calibration and quantitation information Integration threshold, peak width, minimum peak area, integration events Component table component names, retention times, response factors... 

Van Dooren and Muller (158) studied the peak width, peak height, and shape index of the melting peak of indium used as a calibrant in DSC. 

In the calibration curve, Aris plotted on the ordinate versus log Cao or log Cs on the abscissa At (min) is directly read off the recorder paper as the peak width b (mm)—measuring the peak width of all peaks at the same level, approximately halfway between the baseline and the top of the peak—that is, b = At Mr, where Ur is the recorder chart speed (mm/ min). 

J. F. Tyson, Extended Calibration of Flame Atomic Absorption Instruments by a Flow Injection Peak Width Method. Analyst, 109 (1984) 319. 

S. R. Bysouth and J. F. Tyson, A Microcomputer-Based Peak-Width Method of Extended Calibration for Flow-Injection Atomic Absorption Spectrometry. Anal. Chim. Acta, 179 (1986) 481. 

We have since verified that there has been no noticeable loss of mass resolution or sensitivity for the Esquire 3000+ using a modified ring electrode that has 2x1.5 mm laser holes and a single 2 mm fluorescence hole. Mass calibration and tuning the phase between the drive frequency and the auxiliary AC were necessary to re-establish ion trap performance. Our experimental peak width (both with and without ring electrode holes) is about 0.3 Th for rhodamine 101, or about half that of the simulation. Possible reasons for this discrepancy include inaccuracies in the shapes of the model electrodes, insufficient cooling time in the model prior to ion ejection (only ca 2 ms was used to shorten the length of the simulation), or an incomplete description of the ion ejection waveform. 

The increase in the peak width as well as the amplitude and slope of the skew term over time is a natural consequence of detector aging, and can be tracked by the periodic calibration of these parameters. The appearance of a long left skew and a signiftcant, irremovable right skew can be signs of the damage to the semiconductor crystal, which can temporarily be fixed by adjusting the pole/zero in the RC-feedback preamplifier. 

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