主题：HPLC techniques

HPLC techniques have been established for the separation of various dye classes; however, they are not necessarily compatible with the mass spectrometer or they have not been applied to single fibers. Griffen et al. (5) developed an isocratic solvent system for the separation of an unknown mixture of basic dyes. Azo sulfonated acid dyes were separated by Chen et al. (6) using a methanol and aqueous ammonium acetate mobile phase, while azo and anthraquinone acid dyes were separated by Holčapek et al. (7) with a similar gradient. Acid dyes were also separated by a complex mobile phase consisting of methanol, water, tetramethylammonium chloride, and ammonium dihydrogenphosphate by Laing et al. (8). Speers et al. (9) used acidified acetonitrile and water for the separation of acid, basic, and disperse dyes. Success was also reported using a methanol and water mobile phase to separate disperse, acid, basic, vat, and direct dyes (10–12). Many of the aforementioned methods have been applied to either wastewater analysis or the extraction of bulk fabric samples, not to single fibers. Additionally, many of the HPLC solvents and reagents used are nonvolatile and therefore inappropriate for integration into the mass spectrometer. Thus, in order to apply the LC/MS in a forensic setting, existing HPLC methods for dye analysis must be modified and the analysis of single fibers must be viable.
Any method applied to the analysis of dyes extracted from fibers must be adequately sensitive for forensic applications. The amount of dye in a fiber length of 2–10 mm is c. 2–200 ng, with some of the lightest shades containing significantly less (8). Speers et al. (9) determined the limits of detection (LOD) for the HPLC analysis of basic, disperse, and acid dye using 5-μL injections. An LOD range was found of 40–800 pg for basic dyes, 230–1250 pg for acid dyes, and from 80 to 1280 pg for disperse dyes. Thus, the LC/MS meets the criteria for analyzing textile dyes from single fibers.
The focus of this study was to develop a methodology for LC/MS analysis of textile dyes in forensic applications that is both appropriate for MS detection and casework-sized samples. The development of a single HPLC method for the separation of both basic and disperse dye classes, as well as methods that can be used on dyes extracted from fibers ≤0.5 cm in length was investigated. Existing HPLC methods were adapted for the LC/MS and verified by extracting single fibers.