AIM: To establish the quality standard of Agui Yangxue Tablets (Radix Angelicae sinensis, Radix Codonopsis, Rhizoma Atractylodis macrocephalae, etc.) METHODS: Radix Astragali, Radix et Rhizoma Glycyrrhizae and Radix Rehmanniae praeparata were identified by TLC. The content of ferulic acid and paeoniflorin was determined by HPLC. The HPLC condition of ferulic acid was as follows: column was Diamonsil~ TM C_ 18 (4.6 mm?200 mm,5 ?m), the mobile phase consisted of acetonitrile -0.085% phosphoric acid (22 ∶ 78), the UV detection wavelength was at 322 nm, the flow rate was 1.0 mL/min with column temperature at 35 ?C . The HPLC condition of paeoniflorin was as follows: column was Diamonsil~ TM C_ 18 (4.6 mm?200 mm,5 ?m), the mobile phase consisted of acetonitrile -0.1% phosphoric acid (15 ∶ 85), the UV detection wavelength was at 229 nm, the flow rate was 1.0 mL/min with column temperature at 25 ?C . RESULTS: The developed TLC spots were quite clear. The calibration curve of ferulic acid was linear with 0.012 9-0.154 8 ?g(r=0.999 7). The average recovery was 99.32% with RSD of 1.74% (n=9). The calibration curve of paeoniflorin was linear with 0.106 8-1.602 0 ?g(r= 0.999 9). The average recovery was 98.68% with RSD of 0.98 (n=9). CONCLUSION: The established TLC and HPLC methods are exclusive, reproducible and suitable for the quality control of Agui Yangxue Tables.

Praseodymium hexacyanoferrate film was modified on the graphite electrode by cyclic voltammetry in the solution of PrCl_3 and K_3Fe(CN)_6. Its electrochemical properties, including the influence of different scan rates, ions, and K~+ concentrations on the film, were studied. The membrane was characterized by IR and XPS. In IR spectrum, the vibration of cyano group verified from the formation of the film on the electrode. In XPS spectrum, the splitting of Fe2p_(1/2) )and Fe2p_(3/2)) showed that the valence of iron has been changed in the film formation, and a proper electropolymerization mechanism was put forward. The results showed the PrHCF film have some electrocatalytic activity to the oxidation of cysteine. The detection conditions for cysteine such as the potential and pH were also discussed.

OBJECTIVEGenistein, a major soy isoflavone metabolite (SIF), inactivates oxidation activity of bovine lactoperoxidase (LPO). Modification of the heme moiety of LPO by nitrogen-containing compounds has been shown to inactivate LPO. In contrast, SIF mediated inactivation of LPO does not involve a heme modification and the mechanism of SIF inhibition is poorly understood.

METHODSAfter inactivation of LPO by genistein in the presence of H(2)O(2), trypsin-digested LPO peptide fragments were collected and analyzed by MALDI-TOF-MS to characterize the chemical binding of genistein(s) to LPO.

RESULTSThe heme moiety of LPO was not modified by genistein. A covalent binding study showed that (3)H-genistein bound to LPO with a ratio of ∼12 to 1. After HPLC analysis and peak collection, trypsin-digested peptide fragments were analyzed by MALDI-TOF-MS. The 3H-genistein co-eluted peptide fragments (RT=24 min) were putatively identified as 199IVGYLDEEGVLDQNR214 with two bound genistein molecules or a genistein dimer (2 259 Da), 486TPDNIDIWIGGNAEPMVER504 with two bound genistein molecules or a genistein dimer (2 663 Da), and 161ARWLPAEYEDGLALPFGWTQR182 with three bound genistein molecules or a genistein trimer (3 060 Da). The fragment with a mass of 1 792 Da (RT=36 min) was identified as 132CDENSPYR139 with three genistein molecules or a genistein trimer.

CONCLUSIONSThe results suggest that LPO was inactivated by irreversible covalent binding of genistein or genistein polymers to particular peptide fragments constituting regions of the outward domain. No genistein interaction with the prosthetic heme moiety of LPO was observed.

Animals ; Cattle ; Enzyme Activation ; drug effects ; Genistein ; metabolism ; Hydrogen Peroxide ; pharmacology ; Isoflavones ; pharmacology ; Lactoperoxidase ; metabolism ; Placental Lactogen ; Protein Binding