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In order to meet the analytical requirements of the European Medicines Agency (EMA), a new HPLC-FLD method was successfully developed using dansyl chloride for the derivatization and determination of the genotoxic impurities N-Nitrosodimethylamine (NDMA) and N-Nitrosodiethylamine (NDEA) in Lisinopril API and its final product. Samples' pretreatment includes liquid–liquid microextraction, denitrosation, and derivatization steps. To optimize the process, the parameters contributing to high sensitivity and yielding reliable results were thoroughly studied and optimized using one-factor-at-a-time and experimental design approaches. The analytes were pre-column derivatized with Dansyl-Cl and analyzed by HPLC-fluorescence (λem/λem = 340/530) using a C18 column and a mixture of phosphate buffer (pH = 2.8; 20 mM)/acetonitrile 55:45 v/v as the mobile phase. The six-level concentration calibration was shown to be linear, with R equal to 0.9995 for both analytes. The limit of detection (LOD) was satisfactory and equal to 4.7 and 0.04 ng/mL for NDMA and NDEA, respectively. Precision was less than 13.4% in all cases, and the average recoveries were equal to 109.2 and 98.1% for NDMA and NDEA, respectively. The proposed procedure is relatively easy, rapid, and suitable for the determination of the two nitrosamines in routine analysis tests. [ABSTRACT FROM AUTHOR]

In order to meet the analytical requirements of the European Medicines Agency (EMA), a new HPLC-FLD method was successfully developed using dansyl chloride for the derivatization and determination of the genotoxic impurities N-Nitrosodimethylamine (NDMA) and N-Nitrosodiethylamine (NDEA) in Lisinopril API and its final product. Samples’ pretreatment includes liquid–liquid microextraction, denitrosation, and derivatization steps. To optimize the process, the parameters contributing to high sensitivity and yielding reliable results were thoroughly studied and optimized using one-factor-at-a-time and experimental design approaches. The analytes were pre-column derivatized with Dansyl-Cl and analyzed by HPLC-fluorescence (λem/λem = 340/530) using a C18 column and a mixture of phosphate buffer (pH = 2.8; 20 mM)/acetonitrile 55:45 v/v as the mobile phase. The six-level concentration calibration was shown to be linear, with R equal to 0.9995 for both analytes. The limit of detection (LOD) was satisfactory and equal to 4.7 and 0.04 ng/mL for NDMA and NDEA, respectively. Precision was less than 13.4% in all cases, and the average recoveries were equal to 109.2 and 98.1% for NDMA and NDEA, respectively. The proposed procedure is relatively easy, rapid, and suitable for the determination of the two nitrosamines in routine analysis tests.

• Extensive cross-comparisons conducted for Dansyl, OPA, Fmoc, Dabsyl, and Marfey's reagent. • Elution pH in LC–MS analysis greatly affects the performance of derivatization strategies. • With Coptis chinensis extract, Dansyl labeling gives the greatest number identified compounds. • With Coptis chinensis extract, each reagent yields a rather distinct compound coverage profile. • No single reagent offers is clearly superior, and case-by-case assessment is required. Current metabolomics research utilizes liquid chromatography-tandem mass spectrometry (LC–MS/MS) analyses to handle biological samples that contain thousands of quantifiable metabolites. However, no LC–MS/MS condition is suitable for directly analyzing all metabolites. An alternative approach is to derivatize metabolites to impart desirable properties such as better chromatographic separation, enhanced ionization efficiency, or fluorescence detection. An important category of metabolites is amine-containing compounds, which includes amino acids, neurotransmitters, alkaloids, biogenic amines, etc. Various derivatization methods have been developed for amine groups, but few studies have compared their relative strengths and weaknesses. We chose Dansyl-Cl, o-phthalaldehyde (OPA), Fmoc-Cl, Dabsyl-Cl, and Marfey's reagent to systematically compare their reactivity, absorbance, fluorescence, chromatographic separation, and ionization efficiencies under three pH conditions—2.6, 5.0, and 8.0. Their MS/MS fragmentation patterns were also examined under different collision energies. Overall, Dansyl-Cl is a very versatile derivatization method, generating products with fluorescence and high ionization efficiency. Fmoc-Cl is similarly useful under highly acidic chromatography conditions. Dabsyl-Cl may be a good alternative at weakly acidic and weakly basic conditions. OPA is a versatile fluorogenic reagent and its chemistry may be fine-tuned by incorporating different thiol molecules. Marfey's reagent is suboptimal in general, but its chiral property is useful for the separation of enantiomers. All five were applied to the analyses of Coptis chinensis , a Chinese medical herb, identifying hundreds of amine-containing metabolites through MS/MS analyses. None of the five methods is clearly superior, and their compound coverage profiles are rather distinct. A combination of multiple derivatization reagents is required for comprehensive coverage. Our comparative data provide useful guidelines for designing more efficient metabolomics experiments for different analytical goals. [ABSTRACT FROM AUTHOR]