Pyridine, reactions with—continued potassium

A iridine traces in aqueous solution can be determined by reaction with 4-(p-nitroben25l)pyridine [1083-48-3] and potassium carbonate [584-08-7]. Quantitative determination is carried out by photometric measurement of the absorption of the blue dye formed (367,368). Alkylating reagents interfere in the determination. A iridine traces in the air can be detected discontinuously by absorption in Folin s reagent (l,2-naphthoquinone-4-sulfonate) [2066-93-5] (369,370) with subsequent chloroform extraction and hplc analysis of the red dye formed (371,372). The detection limit is ca 0.1 ppm. Nitrogen-specific thermal ionisation detectors can be used for continuous monitoring of the ambient air. 

Preparation of 1,2,3,4-tetrahydro-l-naphthylideneamino p-slyrenesulfonate (NISS) To a solution of 1-tetralone oxime (13.7g, 0.085 mol) in pyridine (32 ml.) below 10 °C was slowly added p-styrenesulfonyl chloride (19g, 0.094 mol), whidi was freshly prepared from potassium p-styrenesulfonate and PClj according to the method reported ly Iwakura et al (15). The reaction was continued with... 

B. 1,2-Bis n-butylthio)benzem. In a 1-1., round-bottomed, three-necked flask fitted with a reflux condenser, a mechanical stirrer, and a thermometer which reaches into the reaction mixture is placed a solution of 59.0 g. (0.25 mole) of o-dibromobenzene in a mixture of 250 ml. of quinoline and 80 ml. of pyridine. To this solution is added 84.0 g. (0.55 mole) of cuprous -butylmer-captide, and the mixture is stirred and heated under reflux (Note 5) for 3.5 hours (Note 6). Heating is stopped and the reaction mixture is allowed to cool to about 100°. It is then poured into a stirred mixture of 1500 g. of ice and 400 ml. of concentrated hydrochloric acid occasional stirring is continued for about 2 hours. The aqueous part is then decanted from the dark brown, gummy residue and is extracted twice with 400 ml. portions of ether. The ether extract is added to the residue, and the resulting mixture is stirred for about 5 minutes. The ether solution is then decanted from the residue and is filtered. The residue is extracted twice more with 400-ml. portions of ether (Note 7). The combined ether extract is washed twice with 100-ml. portions of 10% hydrochloric acid, once with water, and twice with 100-ml. portions of concentrated ammonia (Note 8). After a final wash with water, the ether solution is dried over anhydrous potassium carbonate. The potassium carbonate is collected on a filter, and the ether is removed from the filtrate by distillation. The remaining brown oil is distilled in vacuum, giving a pale orange oil, b.p. 123-124°/0.3 mm., d 1.5684. The yield is 46.5-56.0 g. (73-87%) (Note 9). 

Work performed in our laboratory over the last several years has systematically addressed many of the problems associated with the thiocyanate chemistry. The use of sodium or potassium trimethylsilanolate for the cleavage reaction provided a method for rapid and specific hydrolysis of the derivatized C-terminal amino acid, which left the shortened peptide with a free C-terminal carboxylate ready for continued rounds of sequencing (3). The use of diphenylphosphoroisothiocyanatidate (DPP-ITC) and pyridine combined the activation and derivatization steps and... 

The chemical scheme for C-terminal sequencing is shown in Figure 2. The first step involves treatment of the peptide or protein sample with diisopropylethylamine in order to convert the C-terminal carboxylic acid into a carboxylate salt. Derivatization of the C-terminal amino acid to a thiohydantoin is accomplished with diphenylisothiocyanatidate (liquid phase) and pyridine (gas phase). The peptide is then extensively washed with ethyl acetate and acetonitrile to remove reaction by-products. The peptide is then treated briefly with gas phase trifluoroacetic acid, followed by water vapor in case the C-terminal residue is a proline (this treatment has no effect on residues which are not proline). The derivatized amino acid is then specifically cleaved with sodium or potassium trimethylsilanolate to generate a shortened peptide or protein which is ready for continued sequencing. In the case of a C-terminal proline which was already removed by water vapor, the silanolate treatment merely converts the C-terminal carboxylic acid group on the shortened peptide to a carboxylate. The thiohydantoin amino acid is then quantitated and identified by reverse-phase HPLC. 

To a three-necked flask equipped with gas inlet and outlet were added a stir bar, 6.0 g sulfur, and 40.0 g l-methyl-4-piperidone. The flask was cooled with ice, and under stirring ammonia gas was bubbled into the mixture. The temperature of the reaction was maintained between 40 and 50°C. The introduction of ammonia was continued until the last traces of sulfur disappeared (usually 2 h). The excess ammonia was removed by connecting the gas outlet to vacuum while stirring. The warm, viscous liquid was diluted with 200 mL 50% potassium carbonate solution, and the mixture was extracted with five 100-mL portions of ether. The ether solution was dried over potassium carbonate, filtered, and treated with hydrogen chloride. A yellow precipitate of l,6 -dimethyl-2, 4 5 6 7 7a -hexahydrospiro[piperidine-4,2 -thiazolo[5,4-c]-pyridine]dihydrochloride formed immediately, which was filtered, washed with ether, and dried under reduced pressue over CaCb to afford 53.5 g of product, m.p. 200-205°C. An analytical sample was prepared by... 

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