Triphenylmethane dyes malachite green (MG) and crystal violet (CV) have been used as antimicrobial, antiparasitic and antiseptic agents in aquaculture. However, MG and CV, as well as their metabolites leucomalachite green (LMG) and leucocrystal violet (LCV) are potential mutagens and carcinogens. Thus, the efficient determination of dye residues is of great concern. Considering the complexity of the aquatic products, the sample pretreatment is significant for decreasing matrix interference and improving detection sensitivity. In this study, a simple and rapid QuEChERS procedure was developed and combined with HPLC analysis for the simultaneous determination of the four dyes in fish tissue. An XCharge C18 column was applied in HPLC analysis to achieve good peak shape and selectivity. The pretreatment method involved the extraction of dyes from fish tissue and further clean-up with dispersive solid phase extraction (d-SPE) material. The extraction volume, extraction time as well as d-SPE materials were systematically optimized. The results indicated that reversed-phase/strong anion exchange (C18SAX) adsorbent in the d-SPE procedure could effectively improve the recovery compared with conventional C18 or C18 incorporated with primary secondary amine (PSA) material. Under optimized conditions, good linearity was achieved in the concentration range of 0.5-100 mg/L with R2 greater than 0. 998. The recoveries were 73%-91% and the precisions were 0.66%-5.41%. The results demonstrated the feasibility and efficiency of QuEChERS procedure incorporated with HPLC for dye monitoring.

A native beta-cyclodextrin (beta-CD) stationary phase was prepared by covalent bonding of beta-CD on silica particles via thiol-ene click chemistry. The resulting beta-CD bonded silica (Click TE-CD) was characterized by elemental analysis, which proved the successful immobilization of beta-CD on the silica support with thiol-ene click chemistry. Click TE-CD was chromatographically evaluated with a set of flavone glycosides under hydrophilic interaction chromatography (HILIC) mode, reversed-phase chromatography (RPLC) mode and supercritical fluid chromatography (SFC) mode. The acetonitrile content dependent "U" retention curves indicated its HILIC/RPLC mixed-mode retention behavior. The difference of the separation selectivity between HILIC, RPLC and SFC was described as orthogonality by using geometric approach. The orthogonalities between HILIC/RPLC, HILIC/SFC, and RPLC/SFC reached 69. 8%, 50. 8% and 50. 8%, respectively. The separation of Chinese traditional medicine Lignum Dalbergia Odorifera extract under HILIC, RPLC and SFC modes indicated the potential of Click TE-CD stationary phase in the analysis of complex samples. The mixed-mode HPLC properties and excellent orthogonality demon strated its flexibility in HPLC method development and its great potential in two-dimensional liquid chromatography separation on one HPLC column by different separation modes.

The digest of the standard protein alpha-casein was used as the objective to investigate the influence of pH value of mobile phase on phosphopeptide enrichment selectivity by using Click OEG-CD matrix. Disodium phenyl phosphate was primarily selected as the model sample. The results indicated that phosphate group was difficultly to be ionized when the pH value of mobile phase was below its pKa value. The retention of disodium phenyl phosphate on the Click OEG-CD matrix was not based on ion exchange interaction, leading to the weak retention. The influence of pH value (2, 4 and 6) of mobile phase under hydrophilic interaction liquid chromatography (HILIC) mode on phosphopeptide enrichment selectivity was investigated taking the digest of alpha-casein as a sample. Phosphopeptides could not be enriched on the matrix with the pH value of 2; phosphopeptides could be enriched on the matrix and the elution window was narrow with the pH value of 4; phosphopeptides could be enriched on the matrix while the elution window was broad with the pH value of 6. According to the investigation, Click OEG-CD matrix could be better applied in phosphopeptide enrichment.