Grandes mujeres científicas: Maud Leonora Menten / Great scientific women: Maud Leonora Menten

Maud Leonora Menten nació en Canadá, tuvo cuatro títulos universitarios: Bachiller en Artes, Master en Fisiología, médica y Doctora en Bioquímica. Trabajó en Estados Unidos, Alemania y Canadá. Trabajó en diferentes áreas: en la distribución de los iones cloruro en el sistema nervioso central, en tumores experimentales y su tratamiento con bromuro de radio, en el equilibrio ácido-base durante la anestesia, en el mecanismo hiperglucemiante de toxinas bacterianas, en el descubrimiento de un mecanismo de acoplamiento en química orgánica y hasta en la electroforesis de las hemoglobinas humanas. Sin embargo, el aporte por el cual es más conocida es su trabajo en el estudio de la cinética enzimática junto a Leonor Michaelis en 1913. El propósito de este trabajo es exponer la vida personal y académica de una científica conocida por la gran mayoría de los profesionales de la salud. La mujer que a principios del siglo XX trabajó con grandes investigadores de Canadá, Estados Unidos y Alemania, cuyos aportes científicos fueron reconocidos muchas décadas después. (AU)

Maud Leonora Menten was born in Canada; she had four university degrees, Bachelor of Arts, Master of Physiology, Physician and Doctor of Biochemistry. She worked in the United States, Germany, and Canada. Maud worked in different areas: the distribution of chloride ions in the central nervous system, experimental tumors and their treatment with radium bromide, the acid-base balance during anesthesia, the hyperglycemic mechanism of bacterial toxins, the discovery of a coupling mechanism in organic chemistry and even the electrophoresis of human hemoglobins. However, the contribution for which she is best known is for her work in the study of enzymatic kinetics with Leonor Michaelis in 1913. The aim of this paper is to expose the personal and academic life of a scientist known to the vast majority of Health professionals. The woman who, at the beginning of the 20th century, worked with great researchers from Canada, the United States and Germany, whose scientific contributions were recognized many decades later. (AU)

Kinetic Analysis of Enzymatic Reaction of Active Components from Xinshao Formula in Liver Microsomes of Sham-Operated and Cerebral Ischemia-reperfusion Injury Rats / 中国实验方剂学杂志

Objective: To investigate and compare enzymatic kinetics of scutellarin,apigenin-7-O-glucronide and paeoniflorin from Xinshao fomula in liver microsomes of sham-operated rats and middle cerebral artery occlusion(MCAO) rats with focal cerebral ischemia-reperfusion injury. Method: Xinshao fomula were incubated respectively with liver microsomes of sham-operated rats and MCAO rats,UPLC-MS and substrate elimination method was employed,Michaelis constant(Km),maximum velocity of enzymatic reaction(Vmax) and intrinsic clearance(CLint) of these three components from Xinshao fomula in liver microsomes of sham-operated rats and MCAO rats were calculated,these parameters between different groups were evaluated by statistical analysis. Result: The Km values of scutellarin,apigenin-7-O-glucronide and paeoniflorin in liver microsomes of MCAO rats were (0.798±0.031),(0.213±0.017),(0.499±0.029) μmol·L-1,which were quite different to these in liver microsomes of sham-operated rats.Compared with the sham-operated group,Vmax and CLint values of scutellarin and paeoniflorin in liver microsomes of MCAO rats were significantly reduced(PPVmax of apigenin-7-O-glucronide in liver microsomes of MCAO rat was also significantly reduced(PConclusion: Metabolic rates of these three active components from Xinshao fomula in liver microsomes of MCAO rats with focal cerebral ischemia-reperfusion injury decrease with low elimination rate.

Cloning and enzymatic function characterization of a novel tropinone reductase I (DaTRI 2) in Datura arborea / 药学学报

Tropinone reductase I (TRI) is a key branch point enzyme in the midstream of tropane alkaloids (TAs) biosynthesis pathway and represents an important target for TAs metabolic engineering, which can lead to metabolic flux of substrate tropinone to TAs. A novel TRI gene was isolated from Datura arborea, a woody resource plant, and designated as DaTRI2 (GenBank accession number is MH705164). The full-length cDNA of DaTRI2 with 1 135 bp exhibits a high sequence homology (96.8%) with DaTRI, and is predicted to encode a protein of 347 amino acids. Deduced DaTRI2 protein contain a conserved TGXXXGXG motif involved in NADPH binding, the catalytic N-S-Y-K tetrad motif and eleven amino acid residues important for binding to its substrate tropinone. The phylogenetic analysis revealed that DaTRI2 and other TRIs from Solanaceous plants belong to the same cluster and DaTRI2 exhibited closest phylogenetic proximity to TRIs from Datura. DaTRI2 was expressed in E. coli and the purified recombinant protein can catalyze both tropinone reduction and tropine oxidation with an optimum pH value of 8.0 and 9.6, respectively. When tropinone was used as the substrate, the Km and Vmax values of DaTRI2 at pH 6.4 were 210.05 μmol·L-1 and 69.6 nkat·mg-1 protein respectively, while the Km and Vmax values for tropine as the substrate were 188.03 μmol·L-1 and 114 nkat·mg-1 protein respectively, at pH 9.6. DaTRI2 transcript was most abundant in the young leaf, followed by the root. Cloning of DaTRI2 gene and biochemical analysis of recombinant DaTRI2 facilitate further research on the molecular mechanism on TAs biosynthesis in woody plants and provide a more potent candidate for TAs metabolic engineering.

Enzyme Kinetics of TM-2 in Rat, Beagle Dog and Human Liver Microsomes / 中国药学杂志

OBJECTIVE: To study the enzymatic kinetics of TM-2 in rat, Beagle dog and human liver microsomes by LC-MS/MS. METHODS: TM-2 was incubated with liver microsomal incubation system. LC-MS/MS method was established for quantitative analysis of TM-2 with cabazitaxel as internal standard. The enzyme kinetics parameters Vmax and Km was calculated by the GraphPad Prism 5.0 software. RESULTS: A rapid and sensitive LC-MS/MS method was developed to study the enzyme kinetics of TM-2 in rat, Beagle dog and human liver microsome. The corresponding enzymatic kinetic parameters in rat, Beagle dog and human were as follows: Vmax values were 16.3, 354.6 and 154.8 nmol•min-1•mg(protein)-1, respectively; Km values were 25.7, 313.8 and 89.4 μmol• L-1, respectively; CLint values were 0.63, 1.13 and 1.73 mL•min-1•mg(protein)-1, respectively. CONCLUSION: The result indicates that there are species differences in the activity of metabolic enzyme and the affinity of TM-2 to the metabolic enzyme. Enzyme kinetic parameters obtained of TM-2 provide important parameters for the further study.

Analysis of the metabolites of genistin and enzyme kinetics of genistin in rat liver microsome / 中国药学杂志

OBJECTIVE: To characterize the metabolism of genistin and study its enzymatic kinetics in rat liver microsome by HPLC-MS. METHODS: Genistin was incubated with rat liver microsomal incubation system. HPLC-MS method was used to characterize the metabolites. A metabolite generation method was established for quantitative analysis of genistein with sulfamethoxazole as internal standard. The enzyme kinetics parameters Vmax and Km was calculated by the GraphPad Prism 5.0 software. RESULTS: The metabolites in vitro incubation system was identified as genistein. The optimal time in rat liver microsomes incubation time of 40 min, the optimal protein concentration of 1 mg · mL-1, a substrate concentration of 50 μmol · L-1. The enzyme kinetics parameters of genistein were as follows; Vmax=(0.1042 ± 0.0033) μmol · min-1 · mg(pro)-1, Km=(28.96 ± 2.80) μmol · L-1. CONCLUSION: The results indicate that genistin can be metabolited as the form of hydroxylation in rat liver microsome. Metabolite generation method is a reliable and simple method for determination of kinetic parameters of hepatic microsomal enzymes, and enzyme kinetic parameters obtained of genistin provide important parameters for further study.