in vitro Metabolism Pathway of Effective Component of Bletilla striata as Militarine in Liver Microsomes and Kinetics Char- acteristics of Enzyme-catalyzed Reactions / 中国药房

OBJECTIVE： To study in vitro metabolism pathway of effective component of Bletilla striata as Militarine in liver microsomes and kinetics characteristics of enzyme-catalyzed reactions. METHODS： The in vitro incubation system of rat and human liver microsomes was established， and incubation reaction of Militarine was performed. UPLC-QTOF-MS was used to identify the structure of its metabolites in combination with UNIFI database and references. Using puerarin as internal standard， UPLC-Triple Quad-MS was used to quantitatively analyze metabolic transformation of Militarine in rat liver microsomes. The kinetic parameters （vmax， km， CLint） of Militarine enzyme-catalyzed reactions with/without reducing coenzyme Ⅱ （NADPH） were calculated by fitting the curves with GraphPad Prism 5.0 software. RESULTS： After incubation in rat and human liver microsomes， Militarine produced a chemical formula C21H29O11， which was presumed to be a metabolite of Militarine ester bond hydrolysis. The kinetic study of enzyme-catalyzed reactions showed that vmax of Militarine enzyme-catalyzed reactions with/without NADPH were 1.955， 2.129 nmol/（h·mg）； km were 8.601， 9.854 nmol/mL； CLint were 0.227 3， 0.216 1 mL/（h·mg）； there was no significant difference between with NADPH and without NADPH. CONCLUSIONS： The main metabolic pathway of Militarine in liver microsomes is the hydrolysis of C1 and C4 ester bonds. Its metabolism does not depend on the pathway of cytochrome P450 enzymes initiated by NADPH.

Modern research progress of Chinese materia medica diaphoretic processing method / 中草药

Diaphoretic processing is one of the essential methods aiming to speed up the drying herbs and help the preservation of medicinal materials in the initial processing of Chinese materia medica. In addition, modern research showed that diaphoretic processing also affects the chemical components pharmacological activities and pharmacodynamic action of medicinal materials. During the diaphoretic processing, the internal temperature of medicinal materials rose, which increased the enzyme activity and improved the enzyme-catalyzed reaction, and then these lead to the change of chemical components of medicinal materials. With the diaphoretic processing continues, continually elevated internal temperature of medicinal materials started a series anti-heat stress reaction and generated some secondary metabolites, which considered as main effective components of medicinal materials. In this paper, the mechanism of diaphoretic processing was revealed by analysis the biological enzyme action and anti-heat stress reaction, in order to provide a theoretical basis for the application and extension of the diaphoretic processing.

Antibody titre quantitation method by ELISA without standard substance based on kinetics of enzyme-catalyzed reactions / 重庆医学

Objective To establish antibody titre quantitation method by ELISwithoustandard substance .MethodThe tesgroupof non-specifi,negative control ,specifiand total antibodiewere sefodetecting serantibodieby ELIS.Aftelineafitting of the time-absorbance datof early developing ,the fitted slopewere used athe velocity of absorbance changing , which were denoted by ν0 ,νC,νS,νT.Based on thathe ν-value and the concentration ? of determinand were linearelationship while the substrate waexcessive ,the function with parameteof ν-valueforeflecting the multiple proportionof antibodiecon-centrationbetween specifiand negative control groupcould be deduced .The assessmenof specifiIgG antibodiein serof KM mice immunized with fish collagen waused aan instance .ResultThe function C/C= (ν-ν0 )/(νC-ν0 ) could calculate the mul-tiple proportion(titre) of antibodieconcentrationbetween specifiand negative control group.Conclusion The above method of antibody titre quantitation isuitable fosemi-quantitative analysiwithoustandard substance .

Study on the enzyme-catalyzed synthesis of ligands for ASGPR-mediated liver targeting liposomes / 中国药学杂志

OBJECTIVE: To synthesize asialoglycoprotein receptor ligand-targeted modifier which is used to insert the surface of liposome by enzyme-catalyzed amidation of lactobionic acid and stearamine. METHODS: The structure of the product was confirmed by IR, ESI-MS and H1-NMR. The effects of types and quantity of enzyme, organic solvents, molar ratio of substrate and temperature of reaction were studied. RESULTS: When using DMSO as reaction medium, Novozym 435 immobilized lipase at 400 U · mL-1, molar ratio of lactobionic acid to stearamine at 2:1, and reacting at 40°C for 24 h, the transformation of stearamine reached more than 99%. CONCLUSION: The enzyme catalysis is useful for synthesizing liver targeting liposomes.