Local conformation of poly(1-adamantyl methacrylate) evaluated from intrinsic viscosity

The dilute-solution properties of poly(1-adamantyl methacrylate), which was prepared by radical polymerization in benzene at 60 °C and fractionated with benzene and ethanol as solvent and nonsolvent, respectively, were examined by the measurement of intrinsic viscosities in benzene, tetrahydrofuran, and cyclohexanone at 30°C. The Mark-Houwink-Sakurada equations were established for each solvent-polymer system. From the viscosity data in these three solvents by means of the Burchard-Stockmayer-Fixman plot, the characteristic ratio and steric factor of the chain were determined to be 16.7 and 2.9, respectively, which are higher than the values reported for other methacrylate polymers bearing cycloalkyl esters.     

Competitive thermal ene reaction and Diels-Alder reactions of 2-[N-(alk-2-enyl)benzylamino]-3-vinylpyrido[1,2-a]pyrimidin-4(4H)-ones

Thermal reaction of 2-[N-(alk-2-enyl)benzylamino]-3-(2-substituted and 2,2-disubstituted)vinylpyrido[1,2-a]pyrimidin-4(4H)-ones gave azepine, the desired ene products, and/or pyran derivatives. The formation of the latter was responsible for the [4+2] cycloaddition reaction between the α,β-unsaturated ester carbonyl moiety as a diene part and the alkenylamino moiety as an ene one. The reaction features depended upon the kinds of substituents both on the vinyl and alkenyl counterparts; strongly electron-withdrawing substituents on the vinyl moiety or an electron-donating substituent on the alkenyl one changed the reaction feature from the ene reaction to the hetero Diels-Alder reaction.     

ESR study of free radicals produced from the reaction of o-substituted phenyl methacrylates with tert-butoxy radical

ESR spectra of radicals produced from the reactions of phenyl methacrylate (PMA) and four o-substituted PMAs, 2,4,6-trimethyl-, 2,6-diisopropyl-, 2,6-di-tert-butyl-, and 2,6-di-tert-butyl-4-methyl-PMAs, with tert-butyoxy radical were measured in 2-methyltetrahydrofuran over the temperature range of ?53 to ?15°C. The coupling constants of the β-methylene protons observed varied with the bulkiness of the o-substituents, whereas the p-substitution did not affect the pattern of the spectra. 2,6-Diisopropyl- and 2,4,6-timethyl-PMAs, which can form homopolymers, gave 5- and 13-line spectra, respectively. For the radicals from 2,6-di-tert-butyl- and 2,6-di-tert-butyl-4-methyl-PMAs, the same 8-line spectrum was observed, indicating that the coupling constant of one of the β-methylene protons was too small to detect. Conformations of the radicals were deduced from the coupling constants of the β-methylene protons. Variation of the ESR spectrum according to the bulkiness of the o-substituent was interpreted as a consequence of steric interactions between the polymer chain bound to the β carbon and the substituents, and the α-methyl group.     

Quality check of heparin injections by 1H-nuclear magnetic resonance spectroscopy

The quality of commercial heparin injections was examined by 400-MHz proton nuclear magnetic resonance (1H-NMR) spectroscopy using several measuring modes. The signals of the N-acetyl protons, as well as the sugar-ring protons, attached to the sulfamino and sulfato group-bearing carbons could be easily distinguished from other proton signals and quantified. Measuring at a high temperature (60 degrees C) enabled clear isolation of the H-5 proton signal in the sulphated iduronic acid residue (Is-5) from other proton signals including that of water. The heparin contents of various heparin injections were estimated by using this signal as an index. However, the signal intensity was not parallel with anticoagulant activity. On the other hand, the N-acetyl proton signal was highly correlated to anticoagulant activity. The present method was also useful for concurrent identification of additives in heparin injections.     

Crystal engineering for topochemical polymerization of muconic esters using halogen-halogen and CH/pi interactions as weak intermolecular interactions

We now report the molecular and crystal structure design of muconic ester derivatives on the basis of crystal engineering using halogen-halogen contacts and CH/pi interactions. The solid-state photoreaction pathway of the dibenzyl (Z,Z)-muconates as the 1,3-diene dicarboxylic acid monomers depends on the structure of the ester groups. The substitution of a halogen atom for the aromatic hydrogen of a benzyl group induces topochemical polymerization to produce stereoregular polymers in a crystalline form, whereas the unsubstituted benzyl derivative isomerizes to yield the corresponding E,E isomer under similar conditions. The topochemical polymerization process is directly confirmed by the fact that the single-crystal structures before and after the polymerization are very similar to each other. From the crystal structure analysis for a series of substituted benzyl (Z,Z)- and (E,E)-muconates, it has been revealed that the planar diene moieties are closely packed to form a columnar structure in the crystals. The stacking of the polymerizable monomers is characterized by a stacking distance of 4.9-5.2 A along the columns. This structure is supported by a halogen-halogen interaction between the chlorine or bromine atoms introduced at the p position of the benzyl groups in addition to an aromatic stacking due to the CH/pi interaction between the benzylic methylene hydrogens and aromatic rings. The design of a monomer packing corresponds to the type and position of the introduced halogen atom and also the polymorphs. To make a stacking distance of 5 A using both halogen-halogen and CH/pi interactions as supramolecular synthons is important for the molecular design of muconic ester derivatives appropriate for topochemical polymerization.     

Sample clean-up method for analysis of complex-type N-glycans released from glycopeptides

N-Glycans in glycoprotein can be liberated either from glycoproteins or from their glycopeptides with glycoamidases. The latter approach is preferable, because it requires a smaller amount of the enzyme, and yields N-glycans in excellent yields. Moreover it alleviates the necessity of removing from the reaction mixture the detergents needed to denature the glycoproteins. On the other hand, this approach necessitates removal of interfering peptidic materials, because some of the peptide peaks often overlap with the peaks of carbohydrate chains in high-performance anion-exchange chromatography (HPAEC). These peptidic materials also hinder labeling of N-glycans by reductive amination. We have tried to remove the interfering peptidic materials by several different methods--octadecyl (C18) silica cartridge, cation-exchange resin column, and graphitized carbon cartridge. Unfortunately, none of these could completely remove the interfering peptidic materials. Therefore, we resorted to modify the amino groups of the peptidic materials with sodium 2,4,6-trinitro-benzene-1-sulfonate (TNBS) to render them more hydrophobic, so that they can be retained more strongly on the C18 or graphitized carbon cartridges. In the model study presented here, we were able to obtain N-glycans for HPAEC analyses without any interfering materials by a combination of TNBS reaction and graphitized carbon treatment.     

Preparation of new nitrogen-bridged heterocycles 72. A new approach to 1-acyl-3-(substituted methylthio)thieno[3',4':4,5]imidazo[1,2-a]pyridine derivatives. [corrected

The alkaline treatment of the pyridinium salts, readily available from the S-alkylations of 3-amino-4-(1-pyridinio)thiophene-5-thiolates with various alkyl halides, in chloroform at room temperature afforded the corresponding thieno[3',4':4,5]imidazo[1,2-a]pyridine derivatives in low to moderate yields via the intramolecular cyclization of the resulting 1,5-dipoles followed by the aromatization of the primary cycloadducts. Interestingly, the reactions using unsymmetrical 3-amino-4-[1-(3-methylpyridinio)]thiophene-5-thiolates afforded only 8-methylthieno[3',4':4,5]imidazo[1,2-a]pyridines and the other 6-methyl derivatives were not formed at all. In addition the isolation of a byproduct in the condensation reaction of pyridinium salt with the solvent (CHCl?) is also discussed.     

Charge heterogeneity of a therapeutic monoclonal antibody conjugated with a cytotoxic antitumor antibiotic, calicheamicin

A robust and highly reproducible capillary isoelectric focusing (cIEF) method for the evaluation of charge heterogeneity of monoclonal antibody (mAb) pharmaceutical which contains covalently bound antitumor compounds was developed using a combination of commercially available dimethylpolysiloxane-coated capillary and carrier ampholyte. In order to optimize major analytical parameters for robust mobilization, experimental responses from three pI markers were selected. The optimized method gave excellent repeatability and intermediate precision in estimated pI values of charge variants with relative standard deviations (RSDs) of not more than 0.06% and 0.95%, respectively, when using IgG(4) as a model. Furthermore, RSDs of charge variant compositions were less than 5.0%. These results suggest that the proposed method can be a powerful tool for reproducible evaluation of charge variants of both naked mAbs and their conjugates with high resolution, and it is applicable to quality testing and detailed characterization in the pharmaceutical industry. In addition, it should be noticed that the method provided non-linear pH gradient within the tested ranges, from pI 9.50 to 3.78, and the pH gradient caused the inconsistency of estimated pI ranges between cIEF and gel IEF. This result indicates that selecting appropriate pI markers based on the target pI ranges of charge variants for each mAb related pharmaceutical is highly recommended for the precise determination of pI values.     

Reactions of 2-amino-, 2-alkylamino-, and 2-piperidino-1-azaazulenes with aryl and chlorosulfonyl isocyanates

Reaction of 2-amino-1-azaazulene with phenyl isocyanate gave 3-phenyl-2H-3,4-dihydro-1,3,4a-triazabenz[5,4-a]azulene-2,4-dione. Reactions of 2-alkylamino-1-azaazulenes with aryl isocyanates gave 2-(N-ethyl-N′-arylureido)-1-azaazulenes initially, which rearranged to N-aryl-2-alkylamino-1-azaazulene-3-carboxamides and successive reaction with another molar amount of aryl isocyanate furnished uracil-fuzed 1-azaazulenes. Reaction of 2-piperidino-1-azaazulene with aryl isocyanate gave N-aryl-2-piperidino-1-azaazulene-3-carboxamide. Reaction of 2-(substituted amino)-1-azaazulenes with chlorosulfonyl isocyanate gave 3-cyano- and 3-chloro-2-(substituted amino)-1-azaazulenes.