Limits of Detection and Determination

Detectability is the basic parameter that provides information concerning the applicability of a given technique in trace analysis. The quantitative characteristic of detectability is the detection limit (DL, LOD, Lq), which defines the smallest amount, or concentration, of a given analyte that can be experimentally stated with a given probability. A numerical value of has sense only when detailed experimental conditions for its evaluation are given. Sometimes the term instrumental limit of detection is used. It should refer to the particular specified instrumentation 

In analytical practice, the concept of a detection hmit for an anal3Tical method is useful. It should be evaluated in conditions directly corresponding to real conditions for the whole analytical procedure and take into account real situations that the analyst meets in performing analysis. Such information should take into account aU interferences that occur in the analytical process, including incompleteness of chemical and physical procedures (extraction, precipitation, distillation, etc.) and possible loss of analyte in the course of the whole analytical procedure. All these phenomena can directly influence the magnitude (and precision) of the final analytical signal. 

Comparison of various analytical procedures used in trace analysis needs comparable conditions of determination and the same procedure for detection limit calculation. In some special cases the calculated limit of detection for pure solutions can differ by one order of magnitude in analysis of biological objects or food, by two orders in analysis of simple inorganic materials, or by as much as three orders for very complex objects such as mineral or geological items [13]. 

In experimental evaluation of the detection limit, each measurement carries an accidental error. When there are sufficient individual results it is assumed that the distribution of errors is normal (i.e., Gaussian), which for very small signals is not strictly fulfilled. Under such conditions the spread of experimental results is characterized by the standard deviation at the background level, 5b- Because exact determination of 5b might be difficult, it is generally assumed that it does not differ significantly from 5b close to limit of detection. Then, 5b can be calculated as follows  

is a coefficient that depends on the assumed probability. Its value is usually k=3. Note that when the results of measurements are given in the units of the analytical signal (e.g., number of counts, absorbance), then the obtained result should be divided by the sensitivity of the method, defined as the ratio of concentration to signal and corresponding to the slope of the analytical graph. 

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The determination of OXO in Japanese oyster was realized using reversed-phase HPLC. Samples were extracted with LLE and SPE recoveries were 88.3% (193). Oyster samples were homogenized with a phosphate buffer adjusted to pH 7. After centrifugation, supernatants were concentrated using an SPE C-18 cartridge. Before use, the cartridge was activated with MeOH and phosphate buffer. After the sample had been passed, the cartridge was flushed with water and the analytes were eluted with MeOH-orthophosphoric acid (9 1). The eluate was evaporated, and the residues were dissolved in the mobile phase. The method developed was validated and the study of OXO stability was performed. The limits of detection and determination were 10 and 40 ng/ml, respectively. 

Under the optimized working conditions, a typical calibration curve for an enzyme system was obtained for gluconic acid, with a linear range between 6.0 x 10-7 and 2.0 x 10-5M (r — 0.999, slope — 1995+31 pA M-1, intercept = (7 + 1) x 10 4pA). The limits of detection and determination, calculated according to the 3Sb/m and 10s criteria, respectively,... 

Sensitivity is mainly a limiting factor in trace analysis. It is evident that the method applied should be sensitive enough so that the concentration of substance to be determined is accessible (limit of detection and determination) or the colonies of microbes countable. It must also deliver a measurable difference for a small change in content. When the sensitivity of the method of final detection is a limiting factor the analyst may have several possibilities which all will influence the selection, optimisation and validation of the other steps of the procedure. He may ... 

If TLC is used as an analytical method in quality control, the reproducibUity, i.e. lack of scatter (precision of both system and method), and also the accuracy of the analytical results must be determined. The GMP/GLP guidelines also require vahdation of the method, i.e. testing for linearity, selectivity, robustness and limits of detection and determination (see also Section 9.1 Validation of TLC Methods ). For method development, high demands are placed on the stationary phases of the chromatographic system ... 

Important characteristics of a method are the limits of detection and determination. The limit of detection is the smallest concentration giving a significant response of the instrument that can be distinguished as being present to above the blank or background response. The limit of determination is the smallest amount of measurand that can be measured with a stated precision. 

The limits of detection and determination can be calculated from calibration curves as recommended by DIN 32645 (1994) (see also Einax et al., 1997). A chromatogram for the MDI calibration concentration of 7.8 ng/ml is shown in Fig. 1.2-4. All relevant parameters for HPLC-analysis are summarized in Table 1.2-3. 

In ipy analytical chemistry paper I tried to give you some practical information about the variability of analytical systems at the trace levels where toxicologists and residue chemists must operate. The lower you go, the more variable will be the results. A copy of ny curve relating precision to concentration is repeated below. Eventually, and probably before the parts per trillion level, at the present state of the art (picograms), the results become so bad that the false positives and false negatives will determine the limit of detection and determination. 

Trace analysis Selectivity, limits of detection and determination, precision, recovery Calibration, trueness, precision, ruggedness... 

Signal-to-noise ratio The ratio of the intensity of the analytical signal to that of the noise. This is used in determining the limits of detection and quantitation. 

Sensitivity, Linearity, and Stability of Instruments. Quinine sulfate has been suggested as a standard for determining the limit of detection and linearity of Instruments (44). Some organic materials dissolved In plastics have been suggested as standards because of their stability (22), and many Inorganic Ions dissolved In glasses are quite stable. 

It is appropriate at this juncture to illustrate the power of chemiluminescence in an analytical assay by comparing the limits of sensitivity of the fluorescence-based and the chemllumlnescence-based detection for analytes in a biological matrix. The quantitation of norepinephrine and dopamine in urine samples will serve as an illustrative example. Dopamine, norepinephrine, and 3,4-dihydroxybenzy-lamine (an internal standard) were derivatized with NDA/CN, and chemiluminescence was used to monitor the chromatography and determine a calibration curve (Figure 15). The limits of detection were determined to be less than 1 fmol injected. A typical chromatogram is shown in Figure 16. 

Situation and Criteria A method was to be developed to determine trace amounts of cyanide (CN ) in waste water. The nature of the task means precision is not so much of an issue as are the limits of detection and quantitation (LOD, LOQ), and flexibility and ease of use. The responsible chemist expected cyanide levels below 2 ppm. 

Vogelgesang, J., Limit of Detection and Limit of Determination Application of Different Statistical Approaches to an Illustrative Example of Residue Analysis, Fresenius Z. Anal. Chem. 328, 1987, 213-220. 

In addition, each workbook contained a summary table of all results and limit of detection (LOD) determinations. The table was organized with sample identifications in the left-hand column. Eor each analyte, the analytical result and the LOD appeared in adjacent columns, and analyte recoveries appeared above the results columns. The summary table was generated automatically from the analytical results in the individual worksheets, without operator intervention or re-entry of any information. 

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