Proteins, speciation analysis

The list of elements and their species listed above is not exhaustive. It is limited to the relatively simple compounds that have been determined by an important number of laboratories specializing in speciation analysis. Considering the economic importance of the results, time has come to invest in adequate CRMs. There is a steadily increasing interest in trace element species in food and in the gastrointestinal tract where the chemical form is the determinant factor for their bioavailability (Crews 1998). In clinical chemistry the relevance of trace elements will only be fully elucidated when the species and transformation of species in the living system have been measured (ComeUs 1996 Cornelis et al. 1998). Ultimately there will be a need for adequate RMs certified for the trace element species bound to large molecules, such as proteins. 

Another intensively studied element in speciation analysis is arsenic. The biological and environmental effects of arsenic species and their transformation pathways have been studied in numerous papers.40- 42 Both arsenite and arsenate accumulate in living tissues because of their affinity for proteins, lipids and other cellular compounds.43 Arsenic species can undergo transformation via... 

The applications based on point (1) are the so-called speciation analysis dealing with organome-tallic compounds, proteins, elements in different oxidation states and similar analytical problems... 

Lange-Hesse, K., Dunemann, L. and Schwedt, G. (1991) Development of a combined ultra- and diafiltration technique for use in speciation analysis of protein-bound cadmium in plants. Fresenius J. Anal. Chem., 339, 240-244. 

Se Speciation in Plants The presence of a number of volatile Se species was reported in edible allium plants such as garlic by GC-AES [79]. Selenomethionine, Se-methylselenocysteine, and y-glutamyl-Se-methyl-L-seleno cysteine were identibed in garlic and onion by HPLC-ICP-MS and ES-MS/MS [80]. Selenomethionine is the primary species found in all types of nuts (19D25 percent of the total Se) [30], sunBower [81], and mushrooms [36, 37], The distribution of Se among different fractions (lipid extract, low molecular weight, and protein fractions) of nuts and speciation analysis was studied [30]. Selenium was not detected in any of the lipid extracts obtained from the different types of nuts [30], Results obtained for Brazil nuts by SEC with on-line ICP-MS detection showed that approximately 12 percent of total Se was weakly bound to proteins [30],... 

Selenium Speciation in Edible Animal Tissues Reports on Se specia-tion analysis in edible animal tissues have been scarce. Speciation analysis of Se in cod muscle tissue was performed by separating the species using both RP- and SE-HPLC prior to ICP-MS detection. The main Se compound found in enzymatic hydrolysates was selenomethionine [42], This selenocompound was absent in MeOHDHCl extracts, indicating that Se was mainly incorporated into proteins. A number of unidentiFed Se species were also detected in cod muscle tissue, the separated Se compounds being quantised on-line by post-column isotope dilution [42], Soluble Se compounds extracted from muscles of chicken, turkey, duck, ostrich, lamb, cattle, and pig were separated by SEC with ICP-MS detection. Four peaks were observed, but distribution of Se among these peaks varied considerably in tissues from different animal species [86]. 

Aluminium is an environmentally abundant element and has no known biological function in living cells. On the other hand, A1 exhibits biological toxicity as demonstrated in experimental models." " Intake and output of aluminium for the healthy population from various sources, biological monitoring of occupational aluminium exposure in serum and speciation analysis of the amount and composition of A1 species with high and low molecular masses, have been reviewed by several groups. Besides the use of mass spectrometric and non-mass spectrometric techniques (e.g., ETAAS after separation of human serum proteins ) ESI-MS-MS analysis has also been performed to identify the A1 species. 

Anion exchange chromatography coupled with ICP MS was used in the simultaneous speciation analysis of As, Se, Sb and Te compounds in extracts of fish [230]. Size exclusion chromatography (SEC) coupled with specific detectors is frequently used to analyse species of trace elements in protein-rich materials, such as extracts of meat and plant tissues. For instance, SEC hyphenated with ICP MS was used for the speciation analysis of Cu and Zn in samples of leguminous plants [191]. The same technique was applied to the speciation analysis of Cu, Cd, Zn, Se, As and Ca in fish [220] and Fe, Zn, Cu, Ag, Cd, Sn and Pb in mussels [189]. SEC HPLC coupled with GF AAS mmed out to be very useful for determining levels of Fe species in baby food [312]. With gel permeation chromatography (GPC) GF AAS, the speciation forms of Cd were determined in two kinds of vegetables contaminated with this element [216]. 

Elements usually interact as constituents of macromolecules (proteins, enzymes, hormones, etc.) or according to their oxidation state. Thus, only problem-related speciation analysis promises to become a key for useful risk assessment of elements in the environment. Also activities such as diagnosis and therapy, as well as the depletion of trace elements, optimum levels of essential elements or toxicity of trace elements, will be estimated more reliably. 

The most complicated speciation analysis is in plants and biological samples. Besides the compounds and ions already mentioned metals in living systems participate in a lot more complicated species. They may be bound to amino acids, proteins, peptides and the separation methods may influence their original distribution. For the determination of organo-arsenic, selenium, lead and tin hydride generation combined with different preconcentration steps may be used (Dedina and Tsalev, 1995). 

MTs in invertebrates have also been named MT-like proteins, MLPs, because they differ significantly from those of vertebrates. In spite of these differences, both vertebrate and invertebrate MTs have been isolated for Cd speciation analysis using similar analytical procedures. A typical preparative procedure to extract MTs from animal tissues is given in Scheme 1. 

In recent years, many analytical strategies for elemental speciation analysis, particularly in (bio)medi-cal speciation, make use of LC-atomic detector hybrid techniques. Most frequently studied elements include As, Pb, Cd, Sn, Hg, Se, Cr, and A1 both in biological and environmental samples. The capability for multielemental detection of ICP-MS can be most useful in such analysis. An illustrative example of multielement trace-element speciation is metal-lothionein proteins (MT) from rabbit liver by LC-ICP-MS. Comparative profiles for Cd, Zn, Cu, and S obtained for MTs from rabbit liver, using a fast protein liquid chromatography (FPLC) column coupled to ICP-MS are shown in Figure 7, using molecular and specific detectors. 

Capillary electrophoresis (CE), as a complement to other separation methods, is widely applied in analytical laboratories. This method is versatile for diverse analytes because of its extraordinary resolution power of up to 500000 theoretical plates, and its ability to separate anions, cations, and neutral molecules in a single analysis. Because the coupling of CE to ICP-MS is introduced into speciation analysis, especially in recent years, several interfaces of CE-ICP MS have been developed, thus the approach has been proven to be potential for protein quantification. ... 

DNA analysis, performance of polymerase chain reactions, clinical assays for pH, enzymes, proteins, oxygen etc., trace pollution monitoring and other sorts of biological analyzes are at the focus of recent developments [5]. Another reference lists environmental monitoring (including speciation), clinical monitoring, and quality control in production processes as applications of pTAS equipment in chemical analysis [30]. 

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