ethyl (2S)-2-amino-4-methylsulfanylbutanoate

Conditions: temp:90.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: A solution of 2.2 g of L-lysine in 13.0 ml of ethyl alcohol and 1.6 ml of conc. sulfuric acid was heated under reflux at 90° C. for 5 hours, obtaining L-lysine ethyl ester in a yield of 76.5%. Separately, L-methionine ethyl ester was prepared from 0.8 g of L-methionine in the same manner as in Example 1. After 66 mg of papain and 6.6 mg of L-cysteine were dissolved in a solution of 1.8 g of sodium carbonate and 0.52 g of sodium hydrogen carbonate in 20 ml of water, 20 g of wheat gluten in limited amounts was added to the solution and mixed therewith. The previously prepared reaction solutions of L-lysine and L-methionine ethyl esters were all added to the protein system of wheat gluten, and the system was kept at 37° C. for 24 hours to effect plastein reaction. The rates of incorporation of L-lysine and L-methionine were 62.7% and 81.6%, respectively. At the end of the reaction, the reaction solution was diluted with water to a volume of 300 ml and adjusted to pH 7 with hydrochloric acid before it was subjected to repeated concentration operations for desalting and purification using a reverse osmosis membrane equipped small experimental apparatus (equipped with a DRS-10 flat membrane, manufactured by Daicell Chemical K.K.) under an operating pressure of 15 kg/cm2. The concentrate resulting from the reverse osmosis treatment was again diluted with water to a volume of 400 ml and adjusted to pH 2.0 with phosphoric acid. After 0.4 g of pepsin was added to the solution, the hydrolytic reaction was carried out for 3 hours. Thereafter, 0.4 g of orientase was added and the hydrolytic reaction was continued for an additional 30 hours. The reaction solution was heated to 90° C. for 10 minutes and then centrifuged under 2000G to remove the solids. The supernatant was freeze dried to yield 18.3 g of a low-molecular weight peptide mixture. This product had an average molecular weight of 350 and contained 8.9% of free amino acids, and the gel filtration showed that it contained 86.2% of compounds with a molecular weight of 700 or lower. Table 2 shows the amino acid composition of the starting protein, wheat gluten and the low-molecular weight peptide mixture produced therefrom. As seen from Table 2, the low-molecular weight peptide mixture had the increased contents of L-lysine and L-methionine as compared with the starting protein. was added to the solution mixed all added to the protein system of wheat gluten plastein reaction At the end of the reaction was subjected to repeated concentration operations purification equipped small experimental apparatus ( equipped with a DRS-10 flat membrane The concentrate resulting from the reverse osmosis treatment After 0.4 g of pepsin was added to the solution the hydrolytic reaction was carried out for 3 hours Thereafter, 0.4 g of orientase was added the hydrolytic reaction was continued for an additional 30 hours to remove the solids dried

Conditions: temp:90.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: In 2.5 liters of water containing 85 g of sodium carbonate and 26 g of sodium hydrogen carbonate, were dissolved 33 g of papain and 3.3 g of L cysteine as a papain activator. With stirring, 1 kg of a commercial soybean protein (manufactured and sold by Fuji Seiyu K.K. under trade name "Fujipro R") in limited amounts was added to the solution and thoroughly mixed and agitated. Separately, a suspension of 50 g of L-methionine in 320 ml of ethyl alcohol was gradually added to 35 ml of conc. sulfuric acid to completely dissolve the methionine. The solution was heated under reflux at 90° C. for 5 hours, obtaining a reaction solution of L-methionine ethyl ester in an yield of 92.3%. The reaction solution was diluted with ethyl alcohol to a volume of 500 ml. A 370-ml portion of the diluted reaction solution was added to the separately prepared protein system. After thorough mixing and agitation, the solution was kept at 37° C. for 24 hours to effect plastein reaction. The yield of L methionine incorporated in the thus produced protein-like material was 88.1% based on the amount of L-methionine ethyl ester added at the start of the reaction. The reaction solution was then diluted with water to a volume of 20 liters and adjusted to pH 6-7 with hydrochloric acid before it was subjected to repeated concentration operations for desalting and purification using a reverse osmosis membrane (tubular type, manufactured by Daicell Chemical K.K. under tradename DRS-10) under an operating pressure of 15 kg/cm2. The concentrate resulting from the reverse osmosis treatment was again diluted with water to a volume of 20 liters and adjusted to pH 3 with phosphoric acid. Then, 20 g of orientase (manufactured by Hankyu Kyoei Bussan K.K.) and 20 g of molsin (manufactured by Fujisawa Chemicals K.K.) were added to the solution, which was kept at 40° C. for 40 hours to effect hydrolysis. During the reaction, the solution was controlled to pH 3 with a 6N phosphoric acid. At the end of the reaction, the solution was heated to 90° C. for 10 minutes to deactivate the proteases. The solids were removed by filtration, and the filtrate was spray dried to yield 685 g of a dry powder, which was a low-molecular weight peptide mixture having L-methionine replenished in the form of an acid residue. The low-molecular weight peptide mixture was found to have an average molecular weight of 320. A sample of the powder was subjected to gel filtration using Cephadex G-10 to find that 87.3% of the sample had a molecular weight of 700 or lower. It was also found that the content of free amino acids was 9.8%. Table 1 shows the amino acid composition of the starting soybean protein and low-molecular weight peptide mixture produced therefrom. 1 kg of a commercial soybean protein (manufactured and sold by Fuji Seiyu K.K. under trade name "Fujipro R") in limited amounts was added to the solution thoroughly mixed agitated The solution was heated

Conditions: temp:15.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: 554 g of N-formyl-p-fluoro-L-phenylalanyl-m-di-(2-chloroethyl)amino-L-phenylalanine are dissolved in 4.5 liters of dimethyl formamide (DMF). The solution is heated slowly for the dissolution step and then cooled to 15° C. While stirring add: 172 g of N-hydroxysuccinimide+225 g of dicyclohexylcarbodiimide+1,235 g-moles of L-methionine ethyl-ester dissolved in DMF. After 16 hours of stirring at room temperature, the precipitated dicylcohexylurea is removed and to the stirred filtrate 18 liters of a mixture of water and ice are added. The white precipitate is filtered, washed with water and completely dried under vacuum. Yield: 710 g (97.2%); m.p.=180°-182° C.; [α]D20 =-7.0°±1° (c=2, DMF). The solution is heated slowly for the dissolution step add After 16 hours of stirring at room temperature the precipitated dicylcohexylurea is removed and to the stirred filtrate 18 liters of a mixture of water and ice are added The white precipitate is filtered washed with water completely dried under vacuum [α]D20 =-7.0°±1° (c=2, DMF)