Calibration and quantification

Exciting the sample at 310 nm usually results in a rather featureless spectrum consisting of a broad elevation with a fluorescence maximum near 360 nm. Therefore, the intensity of the fluorescence emission at 360 nm is often used to calculate the sample concentration. Under certain conditions, e.g., when a major portion of the pollutant is a diesel fuel rich in monocyclic and dicyclic aromatics but poor in more highly condensed PAH, higher sensitivities may be obtained by selecting lower excitation and emission wavelengths (e.g., 280 ex/ 330 em). 

When the source of the fluorescing material extracted from the water is known and this material can be obtained in substance, e.g., in the case of an accidental discharge of oil, the oil should be used as the reference. When the source is unknown and the spectrum has the usual shape of a broad elevation with a maximum near 360 nm, experience accumulated over several years indicates that a light Arabian crude oil is a useful reference material. For calibrating analyses of moderately oil contaminated waters, a solution is prepared of approximately 20 mg of this oil per 10 mL spectroscopic grade n-hexane. 

Sample oil content is determined from the FI readings (mV detector output) at a dilution within the linearly calibrated range. The sample FI s) is corrected for the reading of the blank hexane FI a,), then multiplied by the dilution used to take the reading and by the slope (5) of the regression line. The concentration is then calculated by taking into acount the toatl volume Vy, [mL] of the sample extract and the volume Vs[L] of the extracted water. 

For analysis of dilute samples, 1 mL of the sample is transferred into the quartz cell. Its FI is read. Then 1 mL of hexane is added and the solution is re-read. The second reading should be approximately A of the first. If the solution is too coneentrated it will have to be diluted until it reads linearly with dilution. For more concentrated solutions it is useful to add 1 mL of hexane to the cell and then read 10 pL additions of the sample extract. 

21 Determination of petroleum residues dissolved and/or finely dispersed in surface seawater 

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Table 8.76 shows the main characteristics of voltammetry. Trace-element analysis by electrochemical methods is attractive due to the low limits of detection that can be achieved at relatively low cost. The advantage of using standard addition as a means of calibration and quantification is that matrix effects in the sample are taken into consideration. Analytical responses in voltammetry sometimes lack the predictability of techniques such as optical spectrometry, mostly because interactions at electrode/solution interfaces can be extremely complex. The role of the electrolyte and additional solutions in voltammetry are crucial. Many determinations are pH dependent, and the electrolyte can increase both the conductivity and selectivity of the solution. Voltammetry offers some advantages over atomic absorption. It allows the determination of an element under different oxidation states (e.g. Fe2+/Fe3+). 

Several successful attempts were done to transfer classical CEIA to a microchip-based format. This kind of miniaturization is a trend that can overcome the limitations of CE in high-throughput systems. On-chip CE offers both parallel analysis of samples and short separation times. Koutny et al. showed the use of an immunoassay on-chip (32). In this competitive approach fluorescein-labeled cortisol was used to detect unlabeled cortisol spiked to serum (Fig. 8). The system showed good reproducibility and robustness even in this problematic kind of sample matrix. Using serum cortisol standards calibration and quantification is possible in a working range of clinical interest. This example demonstrated that microchip electrophoretic systems are analytical devices suitable for immunological assays that can compete with common techniques. 

Calibration and quantification procedures are easier in LA-ICP-MS compared to other solid-state mass spectrometric techniques because the laser ablation and the ICP ion source operate at normal pressure and the laser ablation of solid samples and ionization of analytes are separated in space and time. Therefore the advantage of solution calibration in ICP-MS can be applied in this solid-state analytical technique. The introduction of solution based calibration, which is only possible in LA-ICP-MS, was an innovative step in the development of this sensitive mass spectrometric technique. A number of different calibration approaches using aqueous standard solutions in the dual gas flow technique have been discussed by various authors.74 75 In the dual gas flow injection technique , the nebulized standard solution and the laser ablated sample material are mixed in the -piece and the two gas flows from the nebulizer (e.g. ultrasonic nebulizer) and laser ablation chamber are added. Using solution based calibration with the addition of a standard solution, Leach et alP determined minor elements in steel reference materials with a relative accuracy of a few %. In comparison to the so-called dual gas flow technique proposed in the literature, where the argon flow rates through the nebulizer and ablation cell add up to 11 min-1 (e.g. 0.451 min-1 and... 

Analytical chemists know that two quantification methods are possible with chromatography quantification with external calibration and quantification with internal calibration. The first mode is only briefly presented in this chapter because it is not adapted to precise quantification. Priority is therefore given to quantification with internal calibration. The development of dosing methods is discussed elsewhere in this chapter. 

Historically, measurements have classified ambient hydrocarbons in two classes methane (CH4) and all other nonmethane volatile organic compounds (NMVOCs). Analyzing hydrocarbons in the atmosphere involves a three-step process collection, separation, and quantification. Collection involves obtaining an aliquot of air, e.g., with an evacuated canister. The principal separation process is gas chromatography (GC), and the principal quantification technique is wdth a calibrated flame ionization detector (FID). Mass spectroscopy (MS) is used along with GC to identify individual hydrocarbon compounds. 

GC/MS. GC/MS is used for separation and quantification of the herbicides. Data acquisition is effected with a data system that provides complete instrument control of the mass spectrometer. The instrument is tuned and mass calibrated in the El mode. Typically, four ions are monitored for each analyte (two ions for each herbicide and two ions for the deuterated analog). If there are interferences with the quantification ion, the confirmation ion may be used for quantification purposes. The typical quantification and confirmation ions for the analytes are shown in Table 4. Alternative ions may be used if they provide better data. 

The detection and quantification capabilities of analytical methods often are important if they are used at trace levels of analytes. The description of the standard addition method, a special calibration in the sample finatises the chapter. 

The purpose of an analytical method is the deliverance of a qualitative and/or quantitative result with an acceptable uncertainty level. Therefore, theoretically, validation boils down to measuring uncertainty . In practice, method validation is done by evaluating a series of method performance characteristics, such as precision, trueness, selectivity/specificity, linearity, operating range, recovery, LOD, limit of quantification (LOQ), sensitivity, ruggedness/robustness, and applicability. Calibration and traceability have been mentioned also as performance characteristics of a method [2, 4]. To these performance parameters, MU can be added, although MU is a key indicator for both fitness for purpose of a method and constant reliability of analytical results achieved in a laboratory (IQC). MU is a comprehensive parameter covering all sources of error and thus more than method validation alone. 

The amounts of injected compounds are logarithmically proportional to the peak area. Calibration curves in the range from 0.08 to 10 nmol showed a regression coefficient from 0.991 to 0.998. Sensitivity was in the subnanomolar range. This method results in a versatile and cost-effective procedure for the detection and quantification of BAs. 

Equation (3.17) is the fundamental equation of the CLS model that allows calibration and prediction. The calibration step consists of calculating S, which is the matrix of coefficients that will allow the quantification of future samples. S is found by entering the spectra and the known concentration of a set of calibration samples in eqn (3.17). These calibration samples, which can be either pure standards or mixtures of the analytes, must contain in total all the analytes that will be found in future samples to be predicted. Then, eqn (3.17) for I calibration samples becomes... 

Because SPME extracts compounds selectively, the response to each compound must be calibrated for quantification. A specific compound can be quantified by using three GC peak area values from solvent injection, static headspace (gas-tight syringe), and SPME. The solvent injection is used to quantify the GC peak area response of a compound. This is used to quantify the amount of the compound in the headspace. The SPME response is then compared to the quantified static headspace extraction. These three stages are necessary because a known gas-phase concentration of most aroma compounds at low levels is not readily produced. A headspace of unknown concentration is thus produced and quantified with the solvent injection. Calibration must be conducted independently for each fiber and must include each compound to be quantified. 

The detection of products derived from the N-oxygenation of C=N functionalities presents many problems, which illustrate difficulties that are associated with the isolation, identification and quantification of small amounts of water-soluble metabolites. Spectrophotometric methods19 as well as differential pulse polarographic techniques20 previously used to determine oximes, nitrones and N-oxides frequently lack sensitivity and/or specificity. Improved analytical methods for the quantification of these N-oxy compounds include chromatographic techniques taking into account the chemical peculiarities of the individual N-oxygenated C=N functionalities. These procedures usually require the chemical synthesis of authentic material for comparison with data obtained with the isolated metabolites, and also for the construction of calibration curves. 

In creams, the important viscosity of the media disabled to identify the constituents involved in the electrochemical signal of a commercial product by comparison of the electrochemical characteristic of the species in aqueous media [27]. Moreover, mixing and adding an amount of solution in the cream changes its structure and quantification with an internal calibration is not conceivable. Identification and quantification of species implied to make home-made creams with different compositions. 

One of the important aspects of any analytical method is its calibration and, therefore, much effort has been put into SPME calibration. As it is not always practicable to employ traditional calibration methods (external standards, internal standards, and standard addition) owing to the sometimes significant matrix effects in complex samples, equilibrium calibration has been suggested as an alternative. In SPME, however, it would normally take rather a long time to achieve equilibrium calibration. If sensitivity were not a concern in an analysis, reduction of extraction time would be desirable, that is, the extraction could be stopped before equilibrium but this would thus demand a new approach to calibration. In this regard, as a way of circumventing matrix effects in environmental analysis, several diffusion-based calibration methods have been recently developed for quantification in SPME.30... 

Huber W, Von Heydebreck A, Sultmann H, Poustka A, Vingron M. Variance stabilization applied to microarray data calibration and to the quantification of differential expression. Bioinformatics 2002 18(suppl 1) S96—S104. 

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