Toxicology of Industrial Compounds

76 METHODS FOR THE DETERMINATION OF REACTIVE COMPOUNDS
Figure 6.3 Modified Edman degradation of alkylated N-terminal valine in
haemoglobin (Törnqvist et al., 1986).
respective diol epoxides results in the formation of relatively stable
electrophiles which also alkylate cysteine residues in proteins.
Upon mild acid treatment these adducts are liberated as the respective
tetrols. Similar to adducts from aromatic amines, the tetrols can be
extracted and analysed by HPLC with specific detection methods, or by GC
with electron capture detection or by GC/MS after derivatisation with
electrophores (Shugart and Kao, 1985; Weston et al., 1989; Day et al.,
1990).
Alkylating agents
Adducts of alkylating agents with the thiol group of cysteine, histidine or
the N-terminal amino acids resist alkaline or acid hydrolysis. To determine
the alkylated amino acids the protein is hydrolysed with 6 N HCl and the
amino acids are separated on a anion exchange column. The fractions
containing the alkylated amino acids are derivatised with electrophores and
analysed by GC/MS (van Sittert et al., 1985; Bailey et al., 1987).