N-acetylation of amino sugar methyl glycosides for gas-liquid chromatographic analysis.

A procedure is described for rapid and quantitative N-acetylation of amino sugars, particularly suitable for gas-liquid chromatographic analysis of constituent carbohydrates in glycoproteins.     

A rapid procedure for detection of bacterial amino acid decarboxylases

The detection of decarboxylases of arginine, glutamic acid, histidine, lysine, ornithine, phenylalanine, tryptophan, and tyrosine in bacteria by thin-layer chromatography on polyamide sheets is described. The bacteria were grown on agar medium plates supplemented with eight amino acids at pH 5.5 for induction of amino acid decarboxylases, then transferred to amine-production media. The decarboxylation products in the spent media (amines and/or γ-amino-n-butyric acid) were dansylated and the dansyl derivatives were separated by thin-layer chromatography on polyamide sheets. This method requires only two separate incubations of the decarboxylase-induced bacteria in amine-production media for 1 h at 37°C for simultaneous detection of eight bacterial amino acid decarboxylases using 0.4 μl of the spent media.     

A procedure for the quantitative analysis of the sulphur amino acids of rat tissues

1. A method is described for the quantitative separation of the sulphur compounds in a single sample of tissue by passing an extract through a serial assembly of ion-exchange resins in the order: Dowex 2 (Cl⁻ form), Dowex 1 (CO₃²⁻ form), Amberlite CG-50 (H⁺ form) and Zeo-Karb 225 (H⁺ form). 2. Groups of sulphur amino acids were eluted separately from each column; the recovery of sulphur compounds after their labelling with ³⁵S in vivo by injection of l-[³⁵S]-methionine was 91–106%. Individual sulphur compounds were further resolved by one-dimensional or two-dimensional paper chromatography. 3. Evidence is presented on the occurrence of S-adenosylmethionine and S-adenosylhomocysteine in rat liver and brain. Rat liver and brain contained 83·6 and 31·4mμ-moles/g. respectively of S-adenosylmethionine.     

The role of sugar configuration in the acetolysis of 6-deoxyhexose methyl glycosides

The acetolysis of several perbenzylated 6-deoxyhexose methyl glycosides under two mild conditions (10 equiv ZnCl₂ in 2:1 v/v Ac₂O-AcOH at 5 °C; 10:10:1 v/v/v Ac₂O-AcOH-TFA at 70 °C) was studied. We focused on the effect of sugar configuration on the competition between mechanisms with activation at exocyclic or endocyclic oxygen site. No effect was detected in acetolysis using the TFA protocol promoting an exo-activation mechanism, which affords 1-O-Ac-pyranosides regardless of sugar configuration. On the contrary, it has a primary role in determining the endo- versus exo-product distribution on ZnCl₂-promoted acetolysis.     

A rapid procedure for purification of EcoRI endonuclease.

A convenient and rapid procedure has been developed to purify restriction endonuclease Eco RI. The method involves sonication of cells at low ionic strength, precipitation of the endonuclease with Polymin P (a polyethyleneimine), elution of the enzyme from the Polymin P precipitate, ammonium sulfate precipitation and chromatography on phosphocellulose. The purified restriction endonuclease is free of exonuclease and other endonucleases.     

A rapid deprotection procedure for phosphotriester DNA synthesis.

An equimolar solution of aldoxime and tetramethylguanidine at 70 degrees C removes both the base and phosphate protection from oligonucleotides prepared by solid phase phosphate triester technology. The rate of cleavage from the succinyl linkage commonly used for solid phase synthesis is also increased. The method is simpler, faster and more easily automated than existing methods.     

A procedure for the rapid preparation of mitochondria from rat liver.

A technique for the rapid preparation of mitochondria from rat liver is described. Tissue fractionation is performed by a single centrifugation step with a discontinuous Percoll density gradient. Total preparation times of 5--6 min are achieved by using this method. The mitochondrial fraction obtained is relatively free of contaminating organelles, as judged by marker-enzyme activity determinations. Mitochondria isolated by Percoll-density-gradient centrifugation differ from mitochondria obtained by differential centrifugation [Taylor, Prpić, Exton & Bygrave (1980) Biochem. J. 188, 443--450] in that the former exhibit a higher acceptor control ratio and a higher calcium content. Values obtained for the protonmotive force are not significantly different between the two preparations. The technique described may be widely applicable for studies requiring the rapid preparation of functionally intact and relatively uncontaminated mitochondria.     

A rapid procedure for the isolation, identification and estimation of citrinin

A rapid procedure for the isolation, identification, and estimation of citrinin, the antibiotic cum nephrotoxin, has been proposed. The colorimetric procedure proposed for estimation makes use of the phenolic function of the compound. This method was found to be applicable to a wide variety of materials.     

A rapid and quantitative determination of amino acid phenylthiohydantoins by direct densitometry on thin-layer chromatogram.

Thin-layer chromatograms on silica gel plate of amino acid phenylthiohydantoins were simultaneously used for identification and quantitation. With a scanning, two-wavelength densitometer, linear calibration curves have been obtained for 15 different derivatives, which can be employed for a rapid and simple quantitation of sample spots. A satisfactory application of the method to the determination of amino-terminal residues of bovine α-chymotrypsin is described.     

A rapid purification procedure for L-asparaginase from Vibrio succinogenes.

A simple procedure has been developed for the purification of L-asparaginase from Vibrio succinogenes. Only two steps of ion-exchange chromatography are required. A higher yield and higher specific activity are obtained than previously reported.