The quantification and analysis of free amino acids (FAAs) in serum is important for illness diagnosis due to its content varies with the body’s condition. In this study, a new sensitive analytical method based on liquid chromatography electrospray ionization mass spectrometry (LC-ESI-MS/MS) was optimized for simple and rapid simultaneous detection of 20 amino acids by pre-column derivatization. Amino acid derivatized by 9-fluorenyl methyl chloroformate (FMOC-Cl) were eluted and separated by simple mobile phase reagent in 20 min via gradient elution using a traditional reverse phase C18 column (150 × 4.6 mm, 5 μm), at the flow rate of 0.8 mL/min. After optimizing derivatization, LC-MS/MS conditions, and sample pre-treatment, this method was verified with LODs and LOQs ranging from 0.0237 to 0.2391 ng/mL and 0.079 to 0.7971 ng/mL. The intra- and inter-day precision was no more than 10 %. The overall accuracy and recovery varied from 88.4-111 % and 88.7–114 %, respectively. Those methods were first applied to mice serum with ulcerative colitis. The results showed that the levels of Ala, Arg, Trp, Lys, Cys, Tyr, and Phe significantly increased in pre-treated group by egg white peptides, which will participate in metabolism. This study provides a valuable reference for further research on the pathogenesis of ulcerous colitis and the development of functional foods to relieve inflammation and balance metabolism.
Functionalization of the surfaces of silica particles is often the first step in their various applications. An improved heterogeneous Fmoc-Cl fluorescent assay using an aqueous solution was developed to detect the number of amino groups on solid-phase supports. The fluorescent Fmoc-Cl method is 50-fold more sensitive than the current UV assay using an organic solvent. This method, together with the homogeneous fluorescamine and OPA assays, is used to detect amino groups on the silica particle surface. The accuracy and effect factors of these methods were examined and the assays were optimized. The results showed that the amine groups on silica particles can produce stronger fluorescence than small amine molecules in solution, because the porous structure of the particle surface is a more hydrophobic environment. The number of active amino groups that can be conjugated with biomolecules is much less than the total number of amino groups on the silica particle. Compared with physical methods, chemical assays involving direct reaction with amino groups would furnish the closest result to the number of active amino groups on the particle surface.
Fluorescence emission of Fmoc-Cl in aqueous solution and linear correlation between intensity and concentration
A prototype of fluorescence sensor, based on 1,4-dihydroxyanthraquinone (1,4-DHAQ) derivative and 9-fluorenylmethyl chloroformate (Fmoc-Cl) co-modified Fe 3 O 4 magnetic nanoparticals (MNPs) (denoted as 1,4-DHAQ/Fmoc-Cl co-modified Fe 3 O 4 MNPs), has been achieved via solvothermal synthesis of amine-functionalized Fe 3 O 4 MNPs and subsequent surface co-modification of 1,4-DHAQ derivative and Fmoc-Cl onto the Fe 3 O 4 MNPs. The 1,4-DHAQ/Fmoc-Cl co-modified Fe 3 O 4 MNPs can be used for not only simple rapid qualitative detection of Cu 2+ , Cd 2+ , Zn 2+ and Hg 2+ in aqueous solution, but also quantitative analysis of the above four heavy metal ions with lower detection limits of 8 × 10 −9 , 1.5 × 10 −8 , 10 −8 and 10 −8 M, respectively. This is the first prototype of fluorescence sensor that has potentials in the detection of Cu 2+ , Cd 2+ , Zn 2+ and Hg 2+ in aqueous solutions and even in drinking water . The sensor designing idea might be extended to other multifunctional chemosensors .
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