Abstract
With the purpose of improvement of carrying out the quantitative determinations in forensic and toxicological analysis the possibility of application of the method of standard for UV-spectrophotometric determination of analytes in biological fluids has been studied. The procedure for determination and acceptability estimation of linearity, accuracy and precision for validation of such methods in the variant of the method of standard tested by the example of UV-spectrophotometric method of doxylamine quantitative determination in blood has been offered. The given procedure provides application of the normalized coordinates. For normalization of the experimental data obtained two approaches have been used: 1) Approach 1: the use of the reference solution with the concentration of the analyte corresponding to its concentration in the final spectrophotometric solution measured under the condition of zero losses for the point of 100% in the normalized coordinates. 2) Approach 2: the use of the reference sample with the concentration of doxylamine corresponding to its concentration for the point of 100% in the normalized coordinates. Estimation of linearity, accuracy and repeatability of the method at the first stage has been performed using the model solutions within the approach based on assumption of insignificance of the uncertainty of the analyte quantitative determination in model solutions in comparison with the complete uncertainty of the analysis results ΔAs. At the second stage determination of linearity, accuracy, repeatability and intermediate precision of the method has been carried out on the model samples prepared using the matrix for three parallel runs. It has been shown that the method of standard can be applied for UV-spectrophotometric determination of doxylamine in blood – Approach 1 is more acceptable.
Highlights
There is a number of international regulations given directive recommendations on carrying out the validation of bioanalytical methods – “Guidance for Industry: Bioanalytical method validation” (U.S FDA, 2001) [5], “Guideline on validation of bioanalytical methods” (ЕМА, 2011) [7], “Guidance for the Validation of Analytical Methodology and Calibration of Equipment used for Testing of Illicit Drugs in Seized Materials and Biological Specimens” (UNODC, 2009) [6] and “Standard Practices for Method Validation in Forensic Toxicology” (SWGTOX, 2012) [8]
In forensic and toxicological analysis we often meet with one-time examinations, and various biological fluids, organs and tissues are sent for the examinations, i.e. it is necessary to determine the analyte quantitatively in several various biological objects, but the necessity of carrying out such determinations can arise rarely enough
It is of interest to study the possibility of using the method of standard when carrying out UV-spectrophotometric determination of analytes in biological fluids; in this connection the procedure for determination and acceptability estimation of linearity, accuracy and precision for validation of such methods in the variant of the method of standard [3] developed according to [1] has been offered
Summary
There is a number of international regulations given directive recommendations on carrying out the validation of bioanalytical methods – “Guidance for Industry: Bioanalytical method validation” (U.S FDA, 2001) [5], “Guideline on validation of bioanalytical methods” (ЕМА, 2011) [7], “Guidance for the Validation of Analytical Methodology and Calibration of Equipment used for Testing of Illicit Drugs in Seized Materials and Biological Specimens” (UNODC, 2009) [6] and “Standard Practices for Method Validation in Forensic Toxicology” (SWGTOX, 2012) [8]. In forensic and toxicological analysis we often meet with one-time examinations, and various biological fluids, organs and tissues are sent for the examinations, i.e. it is necessary to determine the analyte quantitatively in several various biological objects, but the necessity of carrying out such determinations can arise rarely enough. In such situation plotting the calibration curve for each matrix demands rather nonrational time consumption, and to the moment of obtaining the results of analysis they can become irrelevant. The results of testing of the approaches offered by the example of the UV-spectrophotometric method of doxylamine quantitative determination in blood are given in this paper [2]
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