Inhibition and labeling of enzymes and abzymes by phosphonate diesters

Reactive phosphonate diesters were designed and prepared as inhibitors of serine proteases and esterases. Inactivation of trypsin, chymotrypsin, and butyrylcholinesterase was determined by residual enzymaticactivity as well as by the release of a chromogenic or fluorogenic product of the inhibition reaction. Second-order rate constants were determined from rates of nitrophenol formation. Application of the reaction for active-site titration of enzyme preparations is demonstrated. A basic functional group present in the nitrophenyl tropane phosphonate diester was shown to confer selectivity for inactivation of try psin and chymotrypsin. Biotinylated derivatives of the phosphonate diesters were prepared to permitanalysis of proteins modified in the inhibition reaction. Labeled polypeptides were resolved by SDSPAGE, electroblotted, and detected by streptavidin-peroxidase staining. A detection limit of less than 4 ng, corresponding to 20 nM of trypsin, was demonstrated. Pretretment of enzymes with DFP or nonbiotinylated phosphonates specifically blocks the labeling. This technique permits identification of serine proteases in complex mixtures with good sensitivity and specificity.     

Kinetic model ofCO oxidation on nonuniform surface analyzed regardingCOads spillover, 2. homotopic method. Effect ofm1 surface portion

The homotopic method has been used to analyze the kinetic model of three-stageCO oxidation on two nonuniform surface patches conjugated byCO ₂ spillover. Diagrams of steady states depending on the portion of surface patchm ₁ at various temperatures and pressure ratiosP(O ₂)/P(CO) have been constructed. The ratios of different type patches corresponding to the maximum overall reaction rate have been found.     

Methods and equipment of industrial activation analysis

Expanding application of activation analysis in industry resulted in exposure of features related to higher requirements to productivity, reliability, automation level, metrological support of analytical methods and equipment. Based on the application of neutron generators, radioisotope neutron sources, nuclear reactors, electron accelerators as activating radiation sources, high-productivity activation analytical systems used directly in analytical laboratories and plants were constructed. Level of development of the above works makes it possible to conclude that industrial activation analysis has formed as an independent trend of nuclear analytics and has considerable prospects.     

Potential function of internal rotation of (CH3)2CClNO,CH3CCl2NO,and their deuterium analogs

The potential function has been constructed for internal rotation of the NO-group in (CH₃)₂CClNO, CH₃CCl₂NO, and their deuterium analogs. It has been established that the N=O bond in the cis_Cl conformer of CH₃CCl₂NO does not eclipse the C-Cl bond, rather forming an angle of approximately 30. It has been shown that a change in substitutent on the -carbon atom may produce a substantial change in the form of the potential function of internal rotation (PFIR) of the NO group.Translated from Izvestiya Akademii Nauk SSSR, Seriya Khimicheskaya, No. 3, pp. 550–555, March, 1990.     

Dual reactivity of diphenylphosphinous and phenylphosphonous acid amides in reactions with N-acetoxymethyl-substituted diethylamine,benzamide, and acetamide

Conclusion N-Acetoxymethyldiethylamine reacts with tetraethyldiamidophenylphosphonite and diethylamidodiphenylphosphinite, and N-acetoxymethylbenzamide with tetraethyldiamidophenylphosphonite at the phosphorus atom only to form Arbuzov reaction products. The reaction of N-acetoxymethylacetamide with tetraethyldiamidophenylphosphonite and diethylamidodiphenylphosphinite, and the reaction of N-acetoxymethylbenzamide with diethylamidodiphenylphosphinite proceed at two reaction centers at the phosphorus atom to form the Arbuzov reaction products, and at the nitrogen atom to form acetoxyphosphinites (phosphonites) and N-diethylaminomethylamides.Translated from Izvestiya Akademii Nauk SSSR, Seriya Khimicheskaya, No. 12, pp. 2765–2769, December, 1986.     

Determination of biologically active low-molecular-mass thiols in human blood. III. Highly sensitive narrow-bore isocratic reversed-phase high-performance liquid chromatography with fluorescence detection

The fast isocratic narrow-bore reversed-phase high-performance liquid chromatographic method employing fluorescence detection is described for the precise reproducible simultaneous measurement of total homocysteine, cysteine and glutathione in human blood. Sample preparation involves conversion of disulfides to free thiols with triphenylphosphine, precipitation of proteins with sulfosalicylic acid, and conjugation of thiols with monobromobimane. Optimized sample preparation conditions as well as chromatographic conditions allowed to obtain reliable quantitative results within the concentration range corresponding to the levels of these thiols in human blood in norm and pathology. The detection limit was approximately 70 amol for all labeled aminothiols. The proposed method for these compounds analysis includes simple sample preparation, high selectivity, good linearity (r2>0.998), high reproducibility (within-run precision for derivatized aminothiol peaks area RSD<1.8% for three times consequently injected sample); high reliability and the small sample volume (1 microl) required for analysis make it suitable for clinical studies.     

Electronic Structure of Substituted Magnesium Malonates and Malonic Aldehyde Dianion Associates

Ab initio calculations using a molecular pseudopotential and an all-electron approximation have been carried out with various basis sets for Mg complexes and for dianions of malonic aldehyde and its imino and chloro derivatives. The valence levels are reproduced most adequately, whereas the atomic charges and bond characteristics depend heavily on the basis used. Employing a pseudopotential does not change the quality of the electronic characteristics of the system, slightly increasing the existing charge separation. The Cl atom in the -position is involved in -conjugation of the dicarbonyl cycle, causing antibonding in the dianion. Introduction of Mg considerably increases the mesomeric effect of Cl and electron density on the N and O atoms, slightly changing the charges on the carbon atoms. According to the calculations, the Mg–O and Mg–N bonds are slightly covalent. The metal AO is not included in the -system of the ring.