Plasma D-penicillamine redox state evaluation by capillary electrophoresis with laser-induced fluorescence.
D-Penicillamine (D-Pen) is a thiol drug used in the treatment of Wilson's disease, rheumatoid arthritis, metal intoxication and cystinuria. We have recently described a new capillary electrophoresis (CE) method to measure physiological thiols, in which separation of total plasma homocysteine, cysteine, cysteinylglycine, glutathione is achieved using the organic base N-methyl-D-glucamine in the run buffer. In this paper, we present an improvement of our method that allows a baseline separation of total plasma D-Pen from the physiological thiols. Moreover, reduced, free and protein-bound forms of drug are measured by varying the order of disulfide reduction with tributylphosphine and proteins precipitation with 5-sulphosalicylic acid (SSA). After derivatization with 5-iodoacetamidofluorescein (5-IAF), samples are separated and measured by capillary electrophoresis with laser-induced fluorescence in an uncoated fused-silica capillary (57 x 75 microm i.d.) using a phosphate/borate run buffer pH 11.4. In these conditions, the migration time of D-Pen is about 7 min and the time required for each analysis is roughly 10 min. The proposed method has been utilized to measure the various forms of the drug in a D-Pen administered Wilson's disease patient.
Zinellu
Angelo
A
Chair of Clinical Biochemistry, University of Sassari, Viale San Pietro 43/B, I-07100 Sassari, Italy. angelozinellu@libero.it
Carru
Ciriaco
C
Sotgia
Salvatore
S
Deiana
Luca
L
D005453:Fluorescence; D006527:Hepatolenticular Degeneration / Q000097:blood / Q000188:drug therapy; D006801:Humans; D007834:Lasers; D010084:Oxidation-Reduction; D010396:Penicillamine / Q000097:blood* / Q000627:therapeutic use; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D010396:Penicillamine