RESUMO
A capillary electrophoresis method and a durable choline biosensor were developed for measuring serum cholinesterase (EC 3.1.1.8) activity, a useful clinical index for liver function. The former is based on separation of benzoate and benzoylcholine (the artificial substrate of cholinesterase) in an uncoated fused-silica capillary. The migration time of benzoylcholine and benzoate was 1.3 min and 5.5 min, respectively. By the peak areas of A(233) signals, the linear dynamic ranges for both analytes were 0.01-50.0 mM, and the relative standard deviations of 1.0 mM benzoylcholine and benzoate were less than 4% and 6%, respectively. The FIA-choline sensor was constructed with the working electrode of the flow cell covered with a natural chitinous membrane purified from Taiwanese soldier crab, Mictyris brevidactylus. The biomembrane served as the supporting material for enzyme immobilization (choline oxidase, EC 1.1.3.17), and also prevented protein adsorption on the electrode surface. The calibration curve was linear between 0.05 and 5.0 mM (r=0.999). The relative standard deviations for 1.0 mM choline (n=7) were less than 3%, and the activity of the bioactive membrane lasted for about 2 months. The analytical results of both methods correlated well (r=0.940).
Assuntos
Técnicas Biossensoriais/instrumentação , Colina/metabolismo , Colinesterases/sangue , Eletroforese Capilar/métodos , Análise de Injeção de Fluxo/métodos , Animais , Benzoatos/isolamento & purificação , Benzoatos/metabolismo , Benzoilcolina/isolamento & purificação , Benzoilcolina/metabolismo , Quitina/química , Colinesterases/metabolismo , Eletroquímica , Humanos , Concentração de Íons de Hidrogênio , Membranas Artificiais , Fatores de TempoRESUMO
A special component is isolated from Semen sinapis Albae (white mustard seed), a traditional Chinese medicine. According to the physical and chemical investigation and spectroscopic identification, this component can be known as p-hydroxybenzoylcholine bisulfate, a choline base. This component in the drug is also determined by RP-HPLC. A reversed-phase C(18) column is used to separate the p-hydroxybenzoylcholine with an eluent of methanol-0.05 mol/L monopotassium phosphate solution (30:70) (adjusted by phosphoric acid to pH 3.6) at the flow rate of 0.5 mL/min. Detection is carried out with a UV detector operated at 285 nm, and the column temperature is 25 degrees C. It reveals that there is 0.021% (w/w) of p-hydroxybenzoylcholine bisulfate in Semen sinapis Albae and 0.037% (w/w) in stir-baked Semen sinapis Albae.
