Screening carbonyl reductases with the ability to catalyze the reduction of complex carbonyl compounds is of great significance for the biosynthesis of R-tolvaptan(R-TVP). In this study, the target carbonyl reductase in the crude enzyme extract of rabbit liver was separated, purified, and identified by ammonium sulfate precipitation, gel-filtration chromatography, ion exchange chromatography, affinity chromatography, and protein mass spectrometry. With the rabbit liver genome as the template, the gene encoding the carbonyl reductase rlsr5 was amplified by PCR and the recombinant strain was successfully constructed. After RLSR5 was purified by affinity chromatography, its enzymatic properties were characterized. The results indicated that the gene sequence of rlsr5 was 972 bp, encoding a protein with a molecular weight of 40 kDa. RLSR5 was a dimeric protein, and each monomer was composed of a (α/β)₈-barrel structure. RLSR5 could asymmetrically reduce 7-chloro-1-[2-methyl-4-[(2- methylbenzoyl)amino]benzoyl]-5-oxo-2,3,4,5-tetrahydro-1H-1-benzazepine (prochiral ketone, PK) to synthesize R-TVP. The specific activity of the enzyme was 36.64 U/mg, and the optical purity of the product was 99%. This enzyme showcased the optimal performance at pH 6.0 and 30 °C. It was independent of metal ions, with the activity enhanced by Mn²⁺. This study lays a foundation for the biosynthesis of tolvaptan of optical grade.
Animals ; Rabbits ; Alcohol Oxidoreductases/biosynthesis* ; Recombinant Proteins/metabolism* ; Escherichia coli/metabolism* ; Liver/enzymology*

Six crystalline components were isolated from the lipophilic fraction of Artemisia annua L. They have been identified as four sesquiterpenes, one flavonol and one coumarin. Qinghaosu I and III are new sesquiterpenes. Five main constituents, camphene, iso-artemisia ketone, 1-camphor, β-carophyllene, and β-pinene were identified from the volatile oil of this herb.
Artemisia annua ; chemistry ; Artemisinins ; chemistry ; isolation & purification ; Bridged Bicyclo Compounds ; chemistry ; isolation & purification ; Camphor ; chemistry ; isolation & purification ; Monoterpenes ; chemistry ; isolation & purification ; Oils, Volatile ; chemistry ; Sesquiterpenes ; chemistry ; isolation & purification ; Terpenes ; chemistry ; isolation & purification

Six crystalline components were isolated from the lipophilic fraction of Artemisia annua L. They have been identified as four sesquiterpenes, one flavonol and one coumarin. Qinghaosu I and III are new sesquiterpenes. Five main constituents, camphene, iso-artemisia ketone, 1-camphor, β-carophyllene, and β-pinene were identified from the volatile oil of this herb.

The article mainly introduces the teaching content and teaching methods of medical ethics education for international students according to the investigation of their basic information,culture background,and learning background in Tianjin Medical University and Tianjin TCM University,and sets up a perfect teaching system with the international standard in order to realize internationalization of medical education.