Dilution linearity matrix effects

In this example, linearity of dilution identified matrix effects in dog urine samples. Minimum required dilution for dog urine was identified as 1 80, a dilution where recovery of the analyte shows reprodudbiUty of results. 

Because of the possibility that the herbicide alachlor could adulterate food if either poultry or livestock consumed contaminated materials, Lehotay and Miller evaluated three commercial immunoassays in milk and urine samples from a cow dosed with alachlor. They found that milk samples needed to be diluted with appropriate solvents (1 2, v/v) to eliminate the matrix effect. One assay kit (selected based on cost) was also evaluated for use with eggs and liver samples from chickens. Egg and liver samples were blended with acetonitrile, filtered, and diluted with water. Linear calibration curves prepared from fortified egg and liver samples were identical... 

Quantitative XRF analysis has developed from specific to universal methods. At the time of poor computational facilities, methods were limited to the determination of few elements in well-defined concentration ranges by statistical treatment of experimental data from reference material (linear or second order curves), or by compensation methods (dilution, internal standards, etc.). Later, semi-empirical influence coefficient methods were introduced. Universality came about by the development of fundamental parameter approaches for the correction of total matrix effects... 

Sample preconcentration was performed by means of an automated on-line SPE sample processor Prospekt-2 (Spark Holland, Emmen, The Netherlands). Oasis HLB cartridges (Waters, Barcelona, Spain) were used to preconcentrate cannabi-noids present in the water samples whereas isolation of the rest of the compounds was done in PLRPs cartridges (Spark Holland). Before extraction, influent samples were diluted with HPLC water (1 9, v/v) to reduce matrix interferences and to fit some analyte concentrations, e.g., cocaine (CO) and benzoylecgonine (BE), within the linear calibration range. A sample volume of 5 mL was spiked with the internal standard mixture (at 20 ng/L) in order to correct for potential losses during the analytical procedure, as well as for matrix effects. Elution of the analytes to the LC system was done with the chromatographic mobile phase. 

The ICP was a Perkin-Elmer 3000DV with an AS90 Autosampler, which has an instrument detection limit of about 1 ppb (for most elements) with a linear calibration up to 100 ppm (for most elements). Solid samples were prepared via microwave digestion in concentrated nitric and hydrochloric acids, then diluted to volume. The ICP was calibrated and verified with two independent, certified standard sets. Spikes and dilutions were done for each batch of samples to check for and/or mitigate any matrix effects. The ICP process ran a constant pump rate of 1.5 mL/min for all samples and standards during analysis. A 3 mL/min rinse and initial sample flush were used to switch between each sample and standard. The plasma was run at 1450 W with argon flow. Trace metal-grade (sub-ppb) acids and two independently NIST-certified calibration standard sets were used for calibration and method verification. 

Approaches have been described for using standard additions without diluting to a constant volume. Multiplying measured responses by a ratio of total to initial volume accomplishes a correction for dilution. However, the matrix concentration is also diluted by the additions, creating a nonlinear matrix effect that may or may not be transformed into a linear effect by the volume correction. Kalivas [83] demonstrated the critical importance of maintaining constant volume. 

Thrombin is added to plasma diluted 1 100 in a concentration range 0-200 nM. The response is linear in the tested concentration range (R2 = 0.990) and reproducibility was good (CV% = 13%). The matrix effect with a blank signal of-21 Hz is present but, despite the high complexity of the matrix, the increase in thrombin concentration could be detected (Fig. 6). 

Most LBAs require some level of sample dilution prior to analysis due to either the assay MRD or high analyte concentrations in the study samples. It is imperative to demonstrate during method development that the analyte, when present in levels above the ULOQ, can be diluted to concentrations within the quantitative range. This may be accomplished by illustrating that the analytical recovery of an ultrahigh matrix spike ( 100 1000 times ULOQ), diluted serially in assay matrix, remains acceptable over a wide concentration range (when corrected for the dilution factor). Dilutional linearity experiments often reveal the presence of a prozone or Hook effect, which is discussed in the next section. 

Parallelism and Dilutional Linearity to Evaluate Matrix Effects... 

Some workers will interpret this as the ability to reproduce calibration curves for a particular assay that are parallel to each other when plotted graphically whether linear or nonlinear, as opposed to a demonstration of analyte recovery without bias, when sample matrix is diluted to assay samples whose concentrations lie above the analytical range of the method. Parallelism and other potential matrix effects should be investigated separately. 

Linearity has been described by some workers in a way which, by the current authors, would be interpreted as matrix parallelism, whereas others will use the term to describe the extent to which a calibration curve is linear in nonligand-binding assays. For the purpose of this chapter, the term linearity or dilution linearity is used to describe the results of experiments conducted using spiked samples to demonstrate the potential for high-concentration samples to be able to be diluted into the analytical range and read with acceptable accuracy and precision. It is often used to give an indication that matrix effects will not cause a problem upon sample dilution in circumstances where incurred or volunteer samples are not available with concentrations of analyte sufficiently high to conduct parallelism experiments. 

As a result of dilution, the element or elements being determined are now present as minor constituents, so the intensity-to-concentration relationship is linear and thus minimizes the matrix effects. 

X-ray tube and for excitation with the filtered Bremsstrahlung spectrum from a tungsten X-ray tube. The data shown were obtained from diluted aqueous solutions which can be considered to be virtually free from any matrix effects. A detection limit of 10 pg for a 10 pi sample corresponds to a concentration of 1 pg/1. A linear dynamic range of four orders of magnitude is obtained for most elements for example, lead at concentrations of 2-20,000 pg/1 using cobalt as an internal standard at 2000 pg/1. 

The most witlely used quantitative method of ICPMS uses a set of calibration standards for preparing a calibration curve. Simple aqueous standards are usually adequate if the unknown solutions are sufticiently dilute — less than 2000 pg/mL of total dissolved solids. With higher concentrations of matrix elements, attempts are often made to match the matrix elements in the samples with those in the standards. To compen-.sate for instrument drift, instabilities, and matrix effects, an internal standard is usually introduced into the standards and the unknowns. The internal standard is an clement that is absent from the samples and that has an atomic mass and ionization potential near those of the analytes. Two elements frequently used for internal standards are indium and rhodium. Both produce ions in the central part of the mass range ( In, "In, and " Rh) and are not often found naturally occurring in samples. Generally, log-log plots of ion current, ion count, or intensity ratios for sample and internal standards are linear over several orders of magnitude of... 

An LC-MS-MS method for simultaneous determination of metronidazole and spiramycin I concentrations in human plasma, saliva, and gingival crevicular fluid (GCF), preceded by liquid-liquid extraction (LLE). Omidazole is used as an internal standard. A C18 column is used with an eluent of acetonitrile, water, and formic acid. Intra- and interbatch precision 7, 12, and 9% in plasma, saliva, and GCF, respectively. Accuracy was lowest in saliva (15.4%) and better in plasma (8.7%) or in GCF (10.7%). Linearity, specificity, recovery, matrix effect, dilution process, stability in human plasma and saliva after three freeze-thaw cycles, stability in human plasma and saliva at ambient temperature, and stability of the extracts in the automatic injector of the HPLC system have been studied. Useful for pharmacokinetic evaluations. 

In a long series of papers on the master equation, Pritchard and his coworkers elucidated for the first time the effects of rotational and vibrational disequilibrium on the dissociation and recombination of a dilute diatomic gas. Ultrasonic dispersion in a diatomic gas was analyzed by similar computational experiments, and the first example of the breakdown of the linear mixture rule in chemical kinetics was demonstrated. A major difficulty in these calculations is that the eigenvalue of the reaction matrix (corresponding to the rate constant) differs from the zero eigenvalue (required by species conservation) by less than... 

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