Quantification and Detection Limits

Best practices in establishing detection and quantification limits for pesticide residues in foods... 

The definitions of method detection and quantification limits should be reliable and applicable to a variety of extraction procedures and analytical methods. The issue is of particular importance to the US Environmental Protection Agency (EPA) and also pesticide regulatory and health agencies around the world in risk assessment. The critical question central to risk assessment is assessing the risk posed to a human being from the consumption of foods treated with pesticides, when the amount of the residue present in the food product is reported nondetect (ND) or no detectable residues . 

There are several factors involved in defining the limitations of an analytical method. Selecting the right method for defining these limitations can be as important as the actual definitions. Factors that must be taken into consideration in defining detection and quantification limits are ... 

Comparison of methods for calculating detection and quantification limits for analytical methods used for food... 

Several methods have been discussed for the determination of method limitations when evaluating procedures for the determination of pesticides in food. A brief comparison of the methods discussed for the determination of the detection and quantification limits of methods used for the analysis of food products can be found in Table 2. 

However, it is likely that in most cases standards set at levels below detection and quantification limits should be regarded as considerably less useful than those set above such limits. They should therefore fall toward the screening or tentative end rather than the mandatory end of the standards spectrum. It may be appropriate, in some cases, to use a detectable concentration as a cause for concern or a trigger for remedial action, although in such cases it is important that the standards and how they are implemented are reviewed as the analytical LODs are gradually improved to the point at which the detectable levels may not necessarily be unacceptable. Also, the required accuracy and precision of the method of analysis to be used should be specified to make sure that all those who are potentially affected are being regulated on an equivalent basis. 

Solid materials are commonly used to retain either analytes or interferents in solution in continuous analytical systems on account of their advantages. Thus, they afford efficient trace preconcentration in a convenient manner, thereby lowering detection and quantification limits and enabling the determination of the target analytes at the required levels. Also, they facilitate the removal of interferents (leachate clean-up) and the storage of analytes as the retained species remain unaltered over long periods due to the inert... 

Some definitions are contradictory, meaningless, without benefit or will cause much expenditure of personnel and measurement capacity, e.g. Limit of determination. This is the smallest analyte content for which the method has been vahdated with specific accuracy and precision . Apart from the fact that precision is included in the explanation of accuracy the definition manifests a fundamental inability to give a definition which is fit for practice. A useful definition of the detection and quantification limit is based on a statistical approach to the confidence hyperbola of a methods calibration curve, elaborated by the Deutsche Forschungsgemeinschaft [12]-... 

The quantification of ochratoxin A, at levels within the range 0.25-10 ng/ml from wine by HPLC-fluorescence detection, was described [193]. RP-HPLC - fluorescence method for the detection of ochratoxin A in wine with a detection limit of 0.05 ng/ml was also published [194]. A stable isotope dilution assay by LC-MS/MS was developed for quantification of the ochratoxin A by using [D5]-ochratoxin A as internal standard with a low detection and quantification limits of 0.5 and 1.4 pg/kg, respectively [195]. The LC-MS/MS method (ESI and APCI) was also applied to the analysis of contaminated coffee samples by ochratoxins A and B with absolute minimum detection limit around 10-20 pg per injection. Fragment ions from the [M+H]+ and [M+Na]+ ions of... 

The final step in the method development process is validation [559-564]. Full validation involves statistical testing of various aspects of method performance (e.g. accuracy, precision, sensitivity, specificity, detection and quantification limits, linearity. 

The HCP antiserum was also used to quantify HCP by ILA assay throughout the process of purification of P40. The reaction steps were the same as those described above for BBG2Na impurities (Fig. 6). The sensitivity range of the P40 HCP-ILA assay was comprised between 5 and 100 ng/ml (Fig. 15), and detection and quantification limits were 5 and 10 ng/ml respectively. 

All samples were extraeted with diehloromethane in an automatic extractor (Biichi B-811). Surrogate recovery standards (D8-naphthalene, DlO-phenanthrene, D12-perylene) were spiked on each PUF and QFF prior to extraction. The volume was reduced after extraction under a gentle nitrogen stream at ambient temperature, and fractionation achieved on a silica gel column. Samples were analyzed using a GC-MS (gas chromatograph coupled with a mass spectrometer) HP 5975 with a J W Scientihc fused silica column DB-5MS, 5% Ph for PAHs. Terphenyl was used as an internal standard. Detection and quantification limits were controlled by laboratory and field blanks, are site dependent and were reported elsewhere (Lammel et al. 2009b). 

The intensity of the current produced by analyte ions is relevant in quantification. Limits of detection are improved when fragmentation is reduced or eliminated and the ion current, attributable to the analyte, is present as a single species. For instance, using Cl often improves both detection and quantification limits when compared to El, although the controlled fragmentation used in selected reaction monitoring can also improve detection limits. Fragmentation as it applies to specific quantification techniques for small molecules is discussed in connection with the quadrupole family of instruments (Sections 3.3.3.1 and 3.3.5). Quantification for biopolymers, particularly proteins, is presented in Section 3.5.1.9. 

A two-step approach was used for the determination of method detection and quantification limits for the sulfur analytes, as described in Lee Aizawa (2003). The two step approach takes into consideration several factors that affect the analyte signal, including instrumental noise, variability in instrumental sensitivity, and variability in method efficiency, matrix effects and interference, and is simple to follow. Other methods, such as the Hubaux-Vos approach for the calculation of the detection limit can also be used, as reported in Fedrizzi et al. (2007). However, this later approach is complicated, time consuming and does not take either the variabiUty in method efficiency or the matrix effects into consideration (Lee Aizawa, 2003). A brief discussion on how to conduct the method vahdation using the two steps approach is mentioned in this section. 

The detection and quantification limits determined here (cl and cq) do not take the matrix interferences into account, because RMSE was determined from calibration standards. The value cq is used in the next step to spike the blank to compute the LOD and LOQ of the method, which incorporates instrumental variations. Consequently, both matrix/analytes and the extraction/analysis are taken into account for the determination of LOD and LOQ. 

Table 4 summarises the p>arameters obtained from the calibration graphs for all of the reductive sulfur analytes, along with the method detection and quantification limits for each compoimd. Linear regression analysis revealed that very good linearities (R > 0.992) were obtained in the calibration graphs for all of the reductive sulfur compounds. The method provided very good detection limits, which were well below the sensory thresholds (See Table 1) of the analysed sulfur compounds. 

The performance of a method is expressed in terms of its specificity, sensitivity, accuracy, precision, detection and quantification limits, linearity, range, robustness, and ruggedness. In addition to the parameters listed above, other factors to be considered are simplicity and cost. The latter may be of less importance for methods used at the product development stage or for regulatory analysis but may be of considerable importance for routine quality control methods. 

In the manuscript that we are describing in depth in this chapter [73], repeatability, reproducibility, and accuracy were checked, and the calibration curves were established, allowing the calculation of the detection and quantification limits. RSD values for repeatability were lower than 7.01% accuracy values oscillated between 97.2% and 102.0% and limits of detection were low, ranging from 1.64 to 730.54 ppb (negative polarity) and from 0.51 to 310.23 ppb (positive polarity). Neither matrix effect nor ion suppression were detected. The authors proved that the method developed for UHPLC-UV-ESI-TOF MS was a very valuable tool, because it was able to determine a wide number of metabolites in a single run and it was reliable from an analytical point of view. For this reason, it was also applied for the quantitative analysis of the 26 samples under study. 

In practice, identification ( yes ) means establishment of chemical structure, determination of detection and quantification limit. If the impurity is unusually toxic then it has to be qualified in all above cases, meaning the demonstration of biological safety. An impurity may stay below the threshold, making any action superfluous. 

The sensitivity and selectivity of GC-MS was used for the development and validation of a novel analytical method for the rapid and simultaneous detection of scopolamine and atropine in buckwheat Fagopyron esculentum) samples and related food products [98]. Low detection and quantification limits were achieved in the selected-ion mode (SIM) by derivatization (silylation) of the analytes. The LOD for atropine and scopolamine were found to be 0.3 and 1 pg kg, respectively, while the LOQ were 1 and 6 pg kg , respectively, which corresponds to less than one D. stramonium seed per million buckwheat fruits. 

Although HPLC is one of the most used techniques, it presents some limitations in detection and quantification limit. Also in complex matrix such as crude plant extracts, identification based on the UV detection and the retention time of standards can lead to a wrong identification of the compounds. 

---