Investigations On Bioaccumulation/Cell-sorption Of Ultra-trace Heavy Metals And Their Speciation

Solid phase extraction protocols with biomaterials as sorbent have attracted extensive attention, which provide promising potentials for separation and preconcentration of ultra-trace heavy metals species, attributed to the diversity of biological cells, abundant binding sites on cell surface such as amines, hydroxyls, thiols, carboxylates, thisoethers, as well as discriminative effects of various functional groups for metal species. At this point, it is highly desirable to investigate the mechanisms of bio-sorption and corresponding methodologies for improving the analytical performance in metal speciation and sample pretreatment when using biological substrates as sorbent.As a new solid phase extraction technique in sample pretreatment, the bioaccumulation/ cell-sorption is still in its infancy in separation/preconcentration of trace heavy metals. Conventional operating procedures not only cause large amount of sample and reagent consumption, but also deteriorate the performance of the entire analytical system. Flow injection/sequential injection analysis provides an excellent platform for the automation and minimization of on line solid phase extraction procedures. In addition, the renewable surface technique can readily be performed in a lab-on-valve system by employing bead injection technique, which not only provides a practical protocol for on line solid phase extraction exploiting living cells as sorbent, but also offers a new avenue for on-line and real-time monitoring of bio-sorption of trace heavy metals via metabolism-dependent uptake.The present work aims at investigating and elucidating various important issues during the process of cell sorption of metal species, including the accumulation mechanisms of different biological substrates to metal ions, the effects of intrinsic property of the cells and external parameters on the sorption performance, the changes of chemical states of the heavy metal species of interest. The final goal of the studies is to develop on-line separation/preconcentration and speciation procedures for heavy metal species via combination of flow injection/sequential injection and cell sorption.Chapter 1 of the thesis discussed the current situation of bioaccumulation/cell sorption for metal species.Chapter 2 of the thesis developed a sequential injection cell-sorption procedure for the separation and preconcentration of ultra-trace cadmium by using a microcolumn packed with Chlorella vulgaris and Saccharomyces cerevisiae cell immobilized silica beads. ETAAS was employed for quantification of the cadmium. The employment of a mixture of two cells significantly improved sorption efficiency of cadmium attributed to the fact that more binding sites were available in such a case. With a sampling volume of 1.0 ml, an enrichment factor of 38.6, a detection limit of 1.0 ngl-1 (3a) along with a sampling frequency of 20 h-1 were obtained within a linear range of 0.005-0.2μgl-1(Cd2+), achieving a precision of 2.3% RSD (0.05μgl-1,n=9).In Chapter 3 of the thesis, the cell-sorption properties of paramagnetic ions of Mn2+ and Cr3+ in the presence of a magnetic field (external effect) were investigated. Magnetic field retards the moving velocity of the metal ions which offers more chances for the paramagnetic ions to interact with various binding sites on the cell surfaces. As a results, the sorption efficiencies of the paramagnetic ions were significantly improved, i.e., the sorption efficiencies of Mn2+ and Cr3+ were improved to 80% and 90% respectively in a magnetic field of 240 mT, as compared to 45% and 60% achieved in the absence of the magnetic field. The system was applied to the separation/preconcentration of ultra-trace level of manganese with detection by ETAAS. With a sampling volume of 500μl, an enrichment factor of 21.2, a detection limit of 8.0ngl-1 (3σ) was obtained. The method was validated with determination of manganese contents in a certified reference material of riverine water (SLRS-4).Chapter 4 investigates selective preconcentration of chromium (VI) by using egg-shell membrane as sorbent. It also indicated that reduction of Cr(VI) was encountered during its sorption on the surface of egg-shell membrane. The reductive sorption process from Cr(VI) to Cr(III) was demonstrated by UV-Vis spectrophotometry and ETAAS. A novel procedure for chromium speciation was thus developed based on the selective retention of Cr(VI). Within a linear range of 0.05-1.25μgl-1,a detection limit of 0.01μgl-1 (3σ) and precision of 3.2% RSD (0.5μgl-1,n=9) were obtained.In the last chapter, sorption of arsenic species by using living HeLa cells cultured on the surface of Sephadex was investigated. The HeLa cell immobilized Sephadex beads were manipulated by employing bead injection in a lab-on-valve system. It has been demonstrated that the cell sorption of arsenic by HeLa cells involves both metabolism-independent biosorption on the surface of the cells and metabolism-dependent bio-accumulation process through the cell membrane. A procedure for the speciation of inorganic arsenic was developed based on the investigations by hyphenating the LOV cell sorption system with ETAAS. Within a linear range of 0.15-2.5μgl-1 and a sample volume of 450μl, an enrichment factor of 11, a detection limit of 0.05μgl-1 (3σ) were obtained.