* All experimental methods are cited from the reference, please refer to the original source for details. We do not guarantee the accuracy of the content in the reference.
Stage #1: With hexachloroethane; triphenylphosphine In tetrahydrofuran at 0℃; for 1.5 h; Stage #2: With potassium hydroxide In tetrahydrofuran; dichloromethane; water at 0℃; for 2 h;
Dissolve 15 mmol of triphenylphosphine with 10 ml of tetrahydrofuran, drop it into a mixed system composed of 10 mmol of carbobenzoxy-alanine (Z-Ala), 20 mmol of hexachloroethane and 10 ml of tetrahydrofuran. After reacting at 0° C. for 1.5 hours, drop it into a liquid mixture containing 18 mmol of glutamine, 20 ml of water and 40 ml of dichloromethane. While reacting, regulate pH to 13 with potassium hydroxide, the reaction temperature is 0° C., the reaction time after dropping is 2 hours. And then acidify it to regulate pH=2.0 with dilute sulfuric acid. The aqueous phase, after concentrated, reacts with hydrogen in methanol at room temperature for 15 hours. The product L-Ala-L-Gln with a yield of 65percent is obtained.
48%
Stage #1: With hexachloroethane; triphenylphosphine In toluene at 0℃; for 3 h; Stage #2: With sodium hydroxide In water; toluene at 15℃; for 1.5 h;
In a round bottom flask, add in 10 mmol each of carbobenzoxy-alanine (Z-Ala), triphenylphosphine and hexachloroethane respectively, and then 30 ml of toluene. After reacting at 0° C. for 3 h., drop it into a liquid mixture containing 25 mmol of glutamine and 20 ml of water. While reacting, regulate pH to 12 with sodium hydroxide, the reaction temperature is 15° C., the reaction time after dropping is 1.5 hours. And then acidify it to pH=2.5 with dilute hydrochloric acid. The aqueous phase, after concentrated, reacts with hydrogen gas in methanol at room temperature for 15 hours. The product L-Ala-L-Gln with a yield of 48percent is obtained.
45%
Stage #1: With hexachloroethane; triphenylphosphine In acetonitrile at 5℃; for 1 h; Stage #2: With sodium hydroxide; potassium carbonate In water; acetonitrile at 5℃; for 2 h;
Dissolve 30 mmol of hexachloroethane with 10 ml of acetonitrile, drop it into a mixed system composed of 10 mmol of carbobenzoxy alanine (Z-Ala), 20 ml of triphenylphosphine and 10 ml of acetonitrile. After reacting at 5° C. for 1.0 h., drop it into 20 ml of water containing 10 mmol of glutamine. While reacting, regulate pH to 10 with 20 mmol of sodium hydroxide and then potassium carbonate successively, the reaction temperature is 5° C., and the reaction time after dropping is 2 hours. And then acidify it to regulate pH to 3.0 with concentrated hydrochloric acid. The aqueous phase, after concentrated, reacts with trifluoroacetic acid at room temperature for 40 hours. After the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln is obtained with a yield of 45percent by recrystallizing the solids with 1,4-dioxane-water.
Stage #1: With hexachloroethane; triphenylphosphine In toluene at 5℃; for 2 h; Stage #2: With sodium hydroxide In water; toluene; Petroleum ether at 10℃; for 1.5 h;
Dissolve 10 mmol of triphenylphosphine with 10 ml of toluene, drop it into a mixed system composed of 10 mmol of tert-butylcarbonylalanine (Boc-Ala), 10 mmol of hexachloroethane and 20 ml of toluene. After reacting at 5° C. for 2 hours, drop it into a liquid mixture containing 15 mmol of glutamine, 20 ml of water and 60 ml of petrolium ether. While reacting, regulate pH to 12 with sodium hydroxide, the reaction temperature is 10° C., the reaction time after dropping is 1.5 hours. And then acidify it to regulate pH=1.5 with dilute sulfuric acid. The aqueous phase, after concentrated, reacts with hydrogen chloride/1,4-dioxane at room temperature for 5 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln with a yield of 52percent is obtained by recrystallizing the solids with 1,4-dioxane-water.
45%
Stage #1: With hexachloroethane; triphenylphosphine In tetrahydrofuran at 0℃; for 1.5 h; Stage #2: With potassium hydroxide; sodium carbonate In tetrahydrofuran; dichloromethane; water at 8℃; for 2 h;
Dissolve 20 mmol of hexachloroethane with 10 ml of tetrahydrofuran, drop it into a mixed system composed of 10 mmol of tert-butylcarbonylalnine (Boc-Ala), mmol of triphenylphosphine and 20 ml of tetrahydrofuran. After reacting at 0° C. for 1.5 hours, drop it into a liquid mixture containing 20 mmol of glutamine, 20 ml of water and 15 ml of dichloromethane. While reacting, regulate pH to 10 with 20 mmol of potassium hydroxide and then with sodium carbonate successively, the reaction temperature is 8° C., the reaction time after dropping is 2 hours. And then acidify it to regulate pH=2.0 with concentrated hydrochloric acid. The aqueous phase, after concentrated, reacts with methylsulfonic acid at room temperature for 20 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln with a yield of 45percent is obtained by recrystallizing the solids with methanol-water.
40%
Stage #1: With hexachloroethane; triphenylphosphine In 1,2-dichloro-ethane at 10℃; for 0.333333 h; Stage #2: With potassium hydroxide In cyclohexane; water; 1,2-dichloro-ethane at 20℃; for 0.5 h;
In a round bottom flask, add in 10 mmol of tert-butylcarbonylalanine (Boc-Ala), 15 mmol of triphenyl phosphine and 20 mmol of hexachloroethane, and then add in 20 ml of 1,2-dichloroethane. After reacting at 10° C. for 20 min, drop it into a liquid mixture containing 30 mmol glutamine, 20 ml of water and 20 ml of cyclohexane. While reacting, regulate pH to 11 with potassium hydroxide, the reaction temperature is 20° C., the reaction time after dropping is 30 min. And then acidify it to regulate pH=1.5 with dilute nitric acid. The aqueous phase, after concentrated, reacts with trifluoroacetic acid at room temperature for 15 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln with a yield of 40percent is obtained by recrystallizing the solids with tetrahydrofuran-water.
Stage #1: With hexachloroethane; triphenylphosphine In tetrahydrofuran at 10℃; for 0.333333 h; Stage #2: With sodium hydroxide In tetrahydrofuran; cyclohexane; water at 25℃; for 0.5 h;
Dissolve 30 mmol of triphenylphosphine with 30 ml of tetrafuran, drop it into a mixed system composed of 10 mmol of N-(O,O-dimethyl) phosphoalanine, 30 mmol of hexaethane and 10 ml of tetrahydrofuran. After reacting at 10° C. for 20 min., drop it into a liquid mixture containing 30 mmol of glutamine, 20 ml of water and 20 ml of cyclohexane. While reacting, regulates pH to 12 with sodium hydroxide, the reaction temperature is 25° C., the reaction time after dropping is 30 min. And then regulate pH to 1.5 by acidifying it with dilute nitric acid. The aqueous phase, after concentrated, reacts with 20percent hydrogen bromide/glacial acetic acid at room temperature for 5 hours. As the reaction is finished, drop in 50 ml of ether, solids deposit, the product L-Ala-L-Gln is obtained with a yield of 50percent by recrystallizing with methanol-water.
45%
Stage #1: With hexachloroethane; triphenylphosphine In dichloromethane; toluene at 0℃; for 3 h; Stage #2: With potassium hydroxide; potassium hydrogencarbonate In dichloromethane; water; toluene; Petroleum ether at 0℃; for 1.5 h;
EXAMPLE 1 Dissolve 20 mmol of hexachloroethane with 10 mmol of dichloromethane, drop it into a mixed system composed of 10 mmol of N-(O,O-dimethyl) phosphoalanine, 20 mmol of triphenylphosphine and 20 mmol of toluene. After reacting at 0° C. for 3 hours, drop the reaction mixture into a stirring mixture containing 25 mmol of glutamine, 20 ml of water and 60 ml of petrolium ether, regulate pH to 10 with 20 mmol of potassium hydroxide and then potassium bicarbonate successively, reaction temperature is 0° C., the reaction time after dropping is 1.5 hours, stirring and the condition of pH=10 are maintained during the course of the reaction, and then it is acidified to pH=2.5 with concentrated hydrochloric acid. The aqueous phase, after concentrated, reacts with methylsulfonic acid at room temperature for 20 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln with a yield of 65percent is obtained by recrystallizing the solids with isopropanol-water. [α]D=10.55, C=2, m.p.=214-215.5. EXAMPLE 2 Dissolve 20 mmol of hexachloroethane with 10 ml of dichlormethane, drop it into a mixed system composed of 10 mmol of N-(O,O-dimethyl) phosphoalanine, 20 mmol of triohenylphosphine and 20 ml of toluene. After reacting at 0° C. for 3 hours, drop it into a liquid mixture containing 25 mmol of glutamine, 20 ml of water and 60 ml of petrolium ether. While reacting, regulate pH to 10 with 20 mmol of potassium hydroxide and then with potassium carbonate successively, the reaction temperature is 0° C. the reaction time after dropping is 1.5 hours. Then acidify it to regulate pH=2.5 with concentrated hydrochloric acid. The aqueous phase, after concentrated, reacts with methylsulfonic acid at room temperature for 20 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln is obtained with a yield of 45percent by recrystallizing the solids with isopropanol-water.
40%
Stage #1: With hexachloroethane; triphenylphosphine In toluene at 5℃; for 1 h; Stage #2: With sodium carbonate In ethanol; water; toluene at 5℃; for 0.166667 h;
In a round bottom flask, add in 10 mmol of N-(O,O-dimethyl) phosphoalanine, 20 mmol of triphenylphosphine and 30 mmol of hexachloroethane respectively, and then add in 20 ml of toluene. After reacting at 5° C. for 1 hour, add it into a liquid mixture containing 10 mmol of glutamine, 20 ml of water and 5 ml of ethanol. While reacting, regulate pH to 9.5 with sodium carbonate, the reaction temperature is 5° C., the reaction time is 10 min. And then regulate pH to 1.0 by acidifying it with phosphoric acid. The aqueous phase, after concentrated, reacts with trifluoroacetic acid at room temperature for 15 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln with a yield of 40percent is obtained by recrystallizing the solids with methanol-water.
Stage #1: With hexachloroethane; triphenylphosphine In dichloromethane at 0℃; for 0.666667 h; Stage #2: With potassium hydroxide In dichloromethane; cyclohexane; water at 20℃; for 0.5 h;
Dissolve 30 mmol of hexaethane with 20 ml of dichloromethane, drop it into a mixed system composed of 10 mmol of N-(O,O-diethyl) phosphoalanine, 30 mmol of triphenylphosphine and 10 ml of dichloromethane. After reacting at 0° C. for 40 min., drop it into a liquid mixture containing 30 mmol of glutamine, 20 ml of water and 10 ml of cyclohexane. While reacting, regulate pH to 13 with potassium hydroxide, the reaction temperature is 20° C., the reaction time after dropping is 30 min. And then acidify it to regulate pH=1.5 with dilute nitric acid. The aqueous phase, after concentrated, react with trifluoroacetic acid at room temperature for 10 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln is obtained with a yield of 60percent by recrystallizing the solids with 1,4-dioxane-water.
40%
Stage #1: With hexachloroethane; triphenylphosphine In acetonitrile at 15℃; for 2.5 h; Stage #2: With sodium carbonate In water; toluene; acetonitrile at 30℃; for 1 h;
In a round bottom flask, add in 10 mmol of N-(O,O-diethyl) phosphoalanine, 10 mmol of triphenylphosphine and 15 mmol of hexachloroethane respectively, and then add in 20 ml of acetonitrile. After reacting at 15° C. for 2.5 hour, drop it into a liquid mixture composed of 30 mmol of glutamine, 20 ml of water and 30 ml of toluene. While reacting, regulate pH to 8.5 with sodium carbonate, the reaction temperature is 30° C., the reaction time after dropping is 1 hour. And then acidify it to regulate pH=2 with dilute sulfuric acid. The aqueous phase, after concentrated, reacts with saturated hydrogen bromide/1,4-dioxane at room temperature for 5 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln is obtained with a yield of 40percent by recrystallizing the solids with tetrahydrofuran-water.
Stage #1: With hexachloroethane; triphenylphosphine In 1,2-dichloro-ethane at 20℃; for 1.5 h; Stage #2: With potassium hydroxide In water; 1,2-dichloro-ethane at 10℃; for 2 h;
Dissolve 15 mmol of hexachloroethane with 10 ml of 1,2-dichloroethane, add it into a mixed system composed of 10 mmol of N-(O,O-diisopropyl) phosphoalanine, 15 mmol of triphenylphosphine, and 10 ml of 1,2-dichloroethane. After reacting at 20° C. for 1.5 hours, drop it into 20 ml of water containing 20 mmol of glutamine. While reacting, regulate pH to 13 with potassium hydroxide, the reaction temperature is 10° C., the reaction time after dropping is 2 hours. And then regulate pH to 1.5 by acidifying it with dilute nitric acid. The Aqueous phase, after concentrated, reacts with trifluoroacetic acid at room temperature for 10 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln with a yield of 52percent is obtained by recrystallizing the solids with isopropanol-water.
45%
Stage #1: With hexachloroethane; triphenylphosphine In dichloromethane at 10℃; for 3 h; Stage #2: With sodium hydroxide; sodium carbonate In dichloromethane; water; ethyl acetate at 10℃; for 1 h;
Dissolve 15 mmol of triphenylphosphine with 20 ml of dichloromethane, drop it into a mixed system composed of 10 mmol of N-(O,O-diisopropyl) phosphoalanine, 20 mmol of hexachloroethane and 10 mmol of dichloromethane. After reacting at 10° C. for 3 hours, add it into a liquid mixture containing 10 mmol of glutamine, 20 ml of water and 20 ml of ethylacetate. While reacting, regulate pH to 9 with 10 mmol of sodium hydroxide and then with sodium carbonate successively, the reaction temperature is 10° C., and the reaction time after dropping is 1 hour. And then acidify it to regulate pH=2.0 with dilute sulfuric acid. The aqueous phase, after concentrated, reacts with 20percent hydrogen bromide/glacial acetic acid at room temperature for 5 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln is obtained with a yield of 45percent by recrystallizing the solids with methyl alcohol-water.
Preferred examples of glutamine derivatives where the group. R2 is an amino acid residue are the dipeptides: L- alpha-alanylglutamine (2) L- beta-alanylglutamine (3) L- asparaginylglutamine (4) L- cysteinylglutamine (6) L- 3,4-dihydroxyphenylalanylglutamine (7) L- cystinylglutamine (8) L- glutaminylglutamine (9) L- glutamylglutamine (10) L- methionylglutamine, (11), and ...
Into a 200 ml three-necked flask was charged 5.0 g (14.2 mmol) of Z-L-Ala-L-Gln, and the peptide was dissolved in 100 ml of a mixture of methanol and water (methanol:water =7:3) and stirred with a magnetic stirrer. After the air in the flask had been replaced with nitrogen, 0.8 g of 2% by weight of palladium-carbon was added thereto, and the protective group was eliminated in a stream of hydrogen. After 2 hours, the completion of the reaction was confirmed by thin layer chromatography, and then the palladium-carbon was filtered off. The filtrate was concentrated to 20 ml under reduced pressure, and 40 ml of isopropyl alcohol was added thereto. The resulting mixture was stirred for 30 minutes, during which time crystals precipitated. The crystals were collected by filtration, washed with isopropyl alcohol and then dried under reduced pressure to give 2.8 g (12.9 mmol) (90%) of L-Ala-L-Gln. m.p.: 205-207 C. (decomp.) [alpha]hd D20: +10.5 (C=2.0, H2 O) IR gammamaxKBr cm-1: 3400, 3350, 1640, 1630, 1600, 1520, 1380, 1150
EXAMPLE 16 0.1 mole L-alanyl-L-glutamine was dissolved in 50 ml 2N sodium hydroxide solution. 0.11 mole propionic acid anhydride were added drop-by-drop at 0 C. [in 18 ml tetrahydrofuran and 27.5 ml 4N sodium hydroxide solution] in such a manner that the pH of the reaction mixture was constantly at approximately 11. After the end of the addition, the mixture was agitated for 1 hour, then acidified with concentrated hydrochloric acid to pH 1 and the volume of the solution concentrated by one half by evaporation. The N-propionyl-L-alanyl-L-glutamine crystallized out, was filtered off, washed with water and dried. Yield: 19.41 g (71% of theory) Melting point: 78-82 C. 1 H-NMR (D2 O): 0.98 (t, 3H), 1.28 (d, 3H), 1.89 (m, 1H), 2.10 (m, 1H), 2.15 (m, 2H), 2.24 (q, 2H), 4.17 (q, 1H), 4.27 (dd, 1H)
2(a). Preparation of H-Ala-Gln-OH 3.50 Grams of Z-Ala-Gln-OH was dissolved in 50 ml of water and 30 ml of methanol, and the mixture was catalytically reduced by using palladium as a catalyst to obtain H-Ala-Gln-OH.
Dissolve 15 mmol of triphenylphosphine with 10 ml of tetrahydrofuran, drop it into a mixed system composed of 10 mmol of carbobenzoxy-alanine (Z-Ala), 20 mmol of hexachloroethane and 10 ml of tetrahydrofuran. After reacting at 0 C. for 1.5 hours, drop it into a liquid mixture containing 18 mmol of glutamine, 20 ml of water and 40 ml of dichloromethane. While reacting, regulate pH to 13 with potassium hydroxide, the reaction temperature is 0 C., the reaction time after dropping is 2 hours. And then acidify it to regulate pH=2.0 with dilute sulfuric acid. The aqueous phase, after concentrated, reacts with hydrogen in methanol at room temperature for 15 hours. The product L-Ala-L-Gln with a yield of 65% is obtained.
48%
In a round bottom flask, add in 10 mmol each of carbobenzoxy-alanine (Z-Ala), triphenylphosphine and hexachloroethane respectively, and then 30 ml of toluene. After reacting at 0 C. for 3 h., drop it into a liquid mixture containing 25 mmol of glutamine and 20 ml of water. While reacting, regulate pH to 12 with sodium hydroxide, the reaction temperature is 15 C., the reaction time after dropping is 1.5 hours. And then acidify it to pH=2.5 with dilute hydrochloric acid. The aqueous phase, after concentrated, reacts with hydrogen gas in methanol at room temperature for 15 hours. The product L-Ala-L-Gln with a yield of 48% is obtained.
45%
Dissolve 30 mmol of hexachloroethane with 10 ml of acetonitrile, drop it into a mixed system composed of 10 mmol of carbobenzoxy alanine (Z-Ala), 20 ml of triphenylphosphine and 10 ml of acetonitrile. After reacting at 5 C. for 1.0 h., drop it into 20 ml of water containing 10 mmol of glutamine. While reacting, regulate pH to 10 with 20 mmol of sodium hydroxide and then potassium carbonate successively, the reaction temperature is 5 C., and the reaction time after dropping is 2 hours. And then acidify it to regulate pH to 3.0 with concentrated hydrochloric acid. The aqueous phase, after concentrated, reacts with trifluoroacetic acid at room temperature for 40 hours. After the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln is obtained with a yield of 45% by recrystallizing the solids with 1,4-dioxane-water.
Dissolve 10 mmol of triphenylphosphine with 10 ml of toluene, drop it into a mixed system composed of 10 mmol of tert-butylcarbonylalanine (Boc-Ala), 10 mmol of hexachloroethane and 20 ml of toluene. After reacting at 5 C. for 2 hours, drop it into a liquid mixture containing 15 mmol of glutamine, 20 ml of water and 60 ml of petrolium ether. While reacting, regulate pH to 12 with sodium hydroxide, the reaction temperature is 10 C., the reaction time after dropping is 1.5 hours. And then acidify it to regulate pH=1.5 with dilute sulfuric acid. The aqueous phase, after concentrated, reacts with hydrogen chloride/1,4-dioxane at room temperature for 5 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln with a yield of 52% is obtained by recrystallizing the solids with 1,4-dioxane-water.
45%
Dissolve 20 mmol of hexachloroethane with 10 ml of tetrahydrofuran, drop it into a mixed system composed of 10 mmol of tert-butylcarbonylalnine (Boc-Ala), mmol of triphenylphosphine and 20 ml of tetrahydrofuran. After reacting at 0 C. for 1.5 hours, drop it into a liquid mixture containing 20 mmol of glutamine, 20 ml of water and 15 ml of dichloromethane. While reacting, regulate pH to 10 with 20 mmol of potassium hydroxide and then with sodium carbonate successively, the reaction temperature is 8 C., the reaction time after dropping is 2 hours. And then acidify it to regulate pH=2.0 with concentrated hydrochloric acid. The aqueous phase, after concentrated, reacts with methylsulfonic acid at room temperature for 20 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln with a yield of 45% is obtained by recrystallizing the solids with methanol-water.
40%
In a round bottom flask, add in 10 mmol of tert-butylcarbonylalanine (Boc-Ala), 15 mmol of triphenyl phosphine and 20 mmol of hexachloroethane, and then add in 20 ml of 1,2-dichloroethane. After reacting at 10 C. for 20 min, drop it into a liquid mixture containing 30 mmol glutamine, 20 ml of water and 20 ml of cyclohexane. While reacting, regulate pH to 11 with potassium hydroxide, the reaction temperature is 20 C., the reaction time after dropping is 30 min. And then acidify it to regulate pH=1.5 with dilute nitric acid. The aqueous phase, after concentrated, reacts with trifluoroacetic acid at room temperature for 15 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln with a yield of 40% is obtained by recrystallizing the solids with tetrahydrofuran-water.
Dissolve 30 mmol of triphenylphosphine with 30 ml of tetrafuran, drop it into a mixed system composed of 10 mmol of N-(O,O-dimethyl) phosphoalanine, 30 mmol of hexaethane and 10 ml of tetrahydrofuran. After reacting at 10 C. for 20 min., drop it into a liquid mixture containing 30 mmol of glutamine, 20 ml of water and 20 ml of cyclohexane. While reacting, regulates pH to 12 with sodium hydroxide, the reaction temperature is 25 C., the reaction time after dropping is 30 min. And then regulate pH to 1.5 by acidifying it with dilute nitric acid. The aqueous phase, after concentrated, reacts with 20% hydrogen bromide/glacial acetic acid at room temperature for 5 hours. As the reaction is finished, drop in 50 ml of ether, solids deposit, the product L-Ala-L-Gln is obtained with a yield of 50% by recrystallizing with methanol-water.
45 - 65%
EXAMPLE 1 Dissolve 20 mmol of hexachloroethane with 10 mmol of dichloromethane, drop it into a mixed system composed of 10 mmol of N-(O,O-dimethyl) phosphoalanine, 20 mmol of triphenylphosphine and 20 mmol of toluene. After reacting at 0 C. for 3 hours, drop the reaction mixture into a stirring mixture containing 25 mmol of glutamine, 20 ml of water and 60 ml of petrolium ether, regulate pH to 10 with 20 mmol of potassium hydroxide and then potassium bicarbonate successively, reaction temperature is 0 C., the reaction time after dropping is 1.5 hours, stirring and the condition of pH=10 are maintained during the course of the reaction, and then it is acidified to pH=2.5 with concentrated hydrochloric acid. The aqueous phase, after concentrated, reacts with methylsulfonic acid at room temperature for 20 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln with a yield of 65% is obtained by recrystallizing the solids with isopropanol-water. [alpha]D=10.55, C=2, m.p.=214-215.5. EXAMPLE 2 Dissolve 20 mmol of hexachloroethane with 10 ml of dichlormethane, drop it into a mixed system composed of 10 mmol of N-(O,O-dimethyl) phosphoalanine, 20 mmol of triohenylphosphine and 20 ml of toluene. After reacting at 0 C. for 3 hours, drop it into a liquid mixture containing 25 mmol of glutamine, 20 ml of water and 60 ml of petrolium ether. While reacting, regulate pH to 10 with 20 mmol of potassium hydroxide and then with potassium carbonate successively, the reaction temperature is 0 C. the reaction time after dropping is 1.5 hours. Then acidify it to regulate pH=2.5 with concentrated hydrochloric acid. The aqueous phase, after concentrated, reacts with methylsulfonic acid at room temperature for 20 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln is obtained with a yield of 45% by recrystallizing the solids with isopropanol-water.
40%
In a round bottom flask, add in 10 mmol of N-(O,O-dimethyl) phosphoalanine, 20 mmol of triphenylphosphine and 30 mmol of hexachloroethane respectively, and then add in 20 ml of toluene. After reacting at 5 C. for 1 hour, add it into a liquid mixture containing 10 mmol of glutamine, 20 ml of water and 5 ml of ethanol. While reacting, regulate pH to 9.5 with sodium carbonate, the reaction temperature is 5 C., the reaction time is 10 min. And then regulate pH to 1.0 by acidifying it with phosphoric acid. The aqueous phase, after concentrated, reacts with trifluoroacetic acid at room temperature for 15 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln with a yield of 40% is obtained by recrystallizing the solids with methanol-water.
Dissolve 30 mmol of hexaethane with 20 ml of dichloromethane, drop it into a mixed system composed of 10 mmol of N-(O,O-diethyl) phosphoalanine, 30 mmol of triphenylphosphine and 10 ml of dichloromethane. After reacting at 0 C. for 40 min., drop it into a liquid mixture containing 30 mmol of glutamine, 20 ml of water and 10 ml of cyclohexane. While reacting, regulate pH to 13 with potassium hydroxide, the reaction temperature is 20 C., the reaction time after dropping is 30 min. And then acidify it to regulate pH=1.5 with dilute nitric acid. The aqueous phase, after concentrated, react with trifluoroacetic acid at room temperature for 10 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln is obtained with a yield of 60% by recrystallizing the solids with 1,4-dioxane-water.
40%
In a round bottom flask, add in 10 mmol of N-(O,O-diethyl) phosphoalanine, 10 mmol of triphenylphosphine and 15 mmol of hexachloroethane respectively, and then add in 20 ml of acetonitrile. After reacting at 15 C. for 2.5 hour, drop it into a liquid mixture composed of 30 mmol of glutamine, 20 ml of water and 30 ml of toluene. While reacting, regulate pH to 8.5 with sodium carbonate, the reaction temperature is 30 C., the reaction time after dropping is 1 hour. And then acidify it to regulate pH=2 with dilute sulfuric acid. The aqueous phase, after concentrated, reacts with saturated hydrogen bromide/1,4-dioxane at room temperature for 5 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln is obtained with a yield of 40% by recrystallizing the solids with tetrahydrofuran-water.
Dissolve 15 mmol of hexachloroethane with 10 ml of 1,2-dichloroethane, add it into a mixed system composed of 10 mmol of N-(O,O-diisopropyl) phosphoalanine, 15 mmol of triphenylphosphine, and 10 ml of 1,2-dichloroethane. After reacting at 20 C. for 1.5 hours, drop it into 20 ml of water containing 20 mmol of glutamine. While reacting, regulate pH to 13 with potassium hydroxide, the reaction temperature is 10 C., the reaction time after dropping is 2 hours. And then regulate pH to 1.5 by acidifying it with dilute nitric acid. The Aqueous phase, after concentrated, reacts with trifluoroacetic acid at room temperature for 10 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln with a yield of 52% is obtained by recrystallizing the solids with isopropanol-water.
45%
Dissolve 15 mmol of triphenylphosphine with 20 ml of dichloromethane, drop it into a mixed system composed of 10 mmol of N-(O,O-diisopropyl) phosphoalanine, 20 mmol of hexachloroethane and 10 mmol of dichloromethane. After reacting at 10 C. for 3 hours, add it into a liquid mixture containing 10 mmol of glutamine, 20 ml of water and 20 ml of ethylacetate. While reacting, regulate pH to 9 with 10 mmol of sodium hydroxide and then with sodium carbonate successively, the reaction temperature is 10 C., and the reaction time after dropping is 1 hour. And then acidify it to regulate pH=2.0 with dilute sulfuric acid. The aqueous phase, after concentrated, reacts with 20% hydrogen bromide/glacial acetic acid at room temperature for 5 hours. As the reaction is finished, add in 50 ml of ether, solids deposit, the product L-Ala-L-Gln is obtained with a yield of 45% by recrystallizing the solids with methyl alcohol-water.
EXAMPLE 3 Effect of Methanol Crystallization The crude crystals of L-alanyl-L-glutamine obtained in Example 2 (comprising 0.111% alanyl-alanyl-glutamine) were dissolved in water, and the resulting solution was fractionated using WA30 resin and then concentrated in the same manner as in Example 2. After the obtained concentrated solution was divided into two portions, crystallization of L-alanyl-L-glutamine was carried out.
With ammonium hydroxide; In water; at 60℃; under 1500.15 Torr;
Alanylglutamine is prepared according to Chinese Patent CN1680428A. Wherein, N-alpha-chloropropionyl)-L-glutamine is synthesized first. Then, 0.12 g of the N-(alpha-chloropropionyl)-L-glutamine crystals containing two diastereomeric isomers and 300 mL of 28% aqueous ammonia are mixed in a reaction vessel under room temperature. The reaction is carried out under 60 C. temperature and 2 kg/cm2 pressure. L-Alanyl-L-glutamine crude product is obtained after workup procedures. TLC shows an Rf value of 0.14, with acetate:n-butanol:water (1:4:1) eluent and ninhydrin stain. The above L-alanyl-L-glutamine crude product is dissolved in water, charged with activated carbon, and stirred under room temperature. Then the activated carbon is filtered out, mother crystals are added into the solution, and the system is settled for a while. Ethanol is added and the system is stirred for 2 hours under room temperature, followed by filtration to provide L-alanyl-L-glutamine solid. Final purification uses Sephadex G-10 column chromatography, with 0.5% acetic acid aqueous solution as eluent and ultraviolet (UV) 210 nm detection. Fractions with Rf value of 0.14 are collected, and concentrated to provide refined product, which has purity of 94.80% as determined by HPLC.
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