FR2501678A1 - NEW PROCESS FOR OBTAINING METHYL ASPARTYL PHENYL ALANINATE - Google Patents

Abstract

New process for the synthesis of methyl L-//c-aspartyl phenyl alaninate comprising the esterification of phenylalaninate by methanol into methyl ester, the condensation of the latter with a mixed carbonic-L-aspartic anhydride in alkaline medium and the decarboxylation of the methyl N-carboxy L-//c-aspartyl phenyl alaninate formed in an acid medium. The product thus obtained may be used as a sweetener in the pharmaceutical or food industry.

Description

The present invention relates to a new process for the synthesis of a polypeptide.

More specifically, it relates to the synthesis of a dipeptide ester by a new route, significantly improved compared to those already described in the literature.

It specifically relates to a new process for the synthesis of the dipeptideester, 1 -OL- aspartyl phenyl methyl alaninate or Aspartame. At the present time, a large number of processes for the synthesis of aspartame are known, which constitutes the most pleasant synthetic sweetener in terms of taste and the most non-toxic among all the known sweetening products (cf. Dupaigne Fruits vol. 32 (1977) 117).

The method according to the invention differs from those previously known by its extreme ease in implementation, the economy of the reagents used and the possibility of using an essentially aqueous medium for the various stages of this synthesis.

The process according to the invention is characterized by the fact that the l-aspartic acid is reacted with excess phosgene to form a mixed aspartyl carbonic anhydride which is condensed with methyl phenyl alaninate in basic medium to obtain a disel of the acid N - carboxy 1 -o (- aspartyl phenyl alaninate of methyl, decomposes the latter in acid medium and obtains the 1- ot- aspartyl phenyl alaninate of methyl.

The process according to the invention can also be defined by the following procedures, currently preferred 1 / the condensation of phosgene with aspartic acid is carried out in an inert oxygenated solvent, such as for example a linear or cyclic ester such as dioxane, letrahydrofuran, or dimethoxy ethane.

2 / the condensation is carried out by heating to a temperature between 40 and 1000, preferably around 600.

3 / the condensation simultaneously produces hydrochloride of aspartic acid insoluble in the reaction medium, which can thus be easily separated and used for a new operation and mixed aspartyl carbonic anhydride.

4 / the condensation of the mixed aspartyl carbonic anhydride and of methyl phenyl alaninate is carried out in the presence of an alkali metal hydroxydeda, such as sodium hydroxide, potassium hydroxide, or lithine. The condensation is further improved by adding carbonate to the alkali metal which has the effect of buffering the pH and of avoiding too large variations in pH '.

5 / the condensation is carried out at a pH of between 10 and 11.

6 / the condensation is carried out at a temperature between - 5 and + 150.

7 / the hydrolysis of the N - carboxylic derivative disel is carried out using a mineral ecide in squous medium.

 s / hydrolysis is carried out using a dilute hydrochloric acid solution.

9 / the hydrolysis of the N - carboxylic derivative is carried out at a pH between 2 and 4 and preferably between 3 and 3.5.

The three stages of the synthesis are therefore carried out using extremely simple and inexpensive reagents. Each of these stages can be easily industrialized and allow continuous operation.

The methyl phenyl alaninate is obtained co-modally by the conventional methods of esterification starting from the phenyl alanine.

A preferred method consists in subjecting the phenyl alanine to the action of methanol in the presence of a strong excess of concentrated sulfuric acid or to the action of methyl sulfate and then to pouring the reaction mixture into an alkaline solution which leaves the pnenyl alanine - which has not reacted - insoluble and which breaks down the methyl sulphate which has formed into the salt of methylsulfouric acid.

Unprocessed phenyl alanine, recovered after esterification can be used for a new esterification reaction.

The mixture soluble in basic medium is extracted with an organic solvent immiscible with water. After evaporation of the solvent, exclusively methyl phenyl alaninate is thus obtained.

The methyl t-aspartyl phenyl alaninate obtained according to the process of the invention is thus completely devoid of ss isomer or of optical isomer d which could have come from a possible racemization. These impurities are therefore eliminated and the product thus obtained manifests a very high sweetness and sweetness without leaving a bitter or salty aftertaste. The product according to the invention can then be presented in the form of tablets or powders with an inert excipient. The usual dosage is 1 mg per unit dose. The following examples illustrate the invention. They do not limit it in any way.

EXAMPLE I
Esterification of phenyl alanine.

16 g 52 of phenyl alanine are suspended in 200 ml of previously cooled methanol and 29 g 40 of sulfuric acid are added very gradually with vigorous stirring. The addition takes about 1 hour and a half and the temperature of the medium is maintained at around 25. during the whole addition by immersion in a water bath.

However, the stirring continues and the mixture is heated at reflux for 3 hours, then the reaction mixture is poured into 400 ml of cold water, the pH of which has been previously adjusted and is kept at 5.5 by addition of sodium hydroxide. The addition is done very slowly to avoid too rapid heating. Then left in contact for one hour with gentle stirring.

The insoluble material is then separated by filtration. The insoluble material consisting essentially of phenyl alanine which has not reacted is drained, washed with cold water and then dried under vacuum.

The filtrate is then exhausted 3 times with 25 ml of methylene chloride.

The separate methylene phases are combined, washed with water (the methyl ester of l phenyl alanine is alkaline and soluble in water. Therefore washing with water is always alkaline by loss of base ester), then dried over sodium sulfate.

They are filtered and evaporated to dryness. 18g65 are thus collected, ie a yield close to 95%.

The product is sufficiently pure to be used without additional purification for the next stage of the synthesis.

EXAMPLE II
Other phenylalanine esterification process
165 g of l-phenylalanine (1 mole) and 191 ml (1 mole) of methyl sulfate diluted in 600 ml of methanol are loaded into a flask equipped for heating under reflux. Heating at reflux with stirring for 6 hours. The transformation of phenyafltne is complete. (TLC on G 60 Merck silica plate - solvent
CHCl364, MeOH 30, H20 4, H COOH 2)
Revelation nirthydrine at 800C
Concentration under vacuum. Obtaining 499 g of an oil which is neutralized at 0 ° C. with 5N sodium hydroxide (213 ml) and extracted with trichloroethane, in three batches. The extracts in trichloroethane are dried over anhydrous sodium sulphate, then concentrated under vacuum. An oil (1929) del is obtained. methyl phenylalaninate which can be stored for one week at OOC. This oil can be crystallized in the form of sulphate by dissolving it in acne, then by slowly adding, while cooling, a 15% solution of sulfuric acid in acetone, prepared immediately. A clear, slightly pink solution is obtained which quickly becomes cloudy and abundantly crystallizes.

Filtration of the crystals, washing with acetone, drying under vacuum or in a ventilated oven at 450C - 191.92 g of methyl phenylalaninate sulfate (white crystalline product). Are thus obtained.

EXAMPLE III
Stage A
1-aspartylcarbonic anhydride.

40 g of l-aspartic acid are dissolved in 250 ml of dioxane at ordinary temperature. Once the dissolution is obtained, the solution is brought to 600, and there is made
@@ remove phosgene until the medium is saturated for 15 hours. Gn observes
gradual precipitation of aspertiquye acid hydrochloride. We filter the
solution after cooling, then the clear filtra is brought to dryness by evspo-
solvent vacuum ration, 13g1 of N - carboxy anhydride are thus collected
mixed which is purified by recrystallization from dichloro 1,2 ethane by hot
and cold.

Stage B
N-carboxy sodium disel 1 - aspartyl phenyl methyl alaninate.

19, tg of 1-aspartylcarbonic mixed anhydride are suspended in 60 ml
of water and while maintaining mechanical agitation, simultaneously adding a
suspension of 15 g of methyl phenyl alanine in 45 ml of water, then a
scude solution to adjust the pH of the medium to 10%, keep stirring
for 2 minutes at the temperature of 0. We get the sodium salt or we
isolates @er filtration
Stage C
l - α - aspartyl phenyl methyl alaninate.

We say 17g5 of sodium disel of N - carboxy 1 - aspartyl phenyl alaninate of
methyl in 100 ml of water, heating the medium to around 40. We then add a
solution N of hydrochloric acid until a pH of 3 to 3.5 is obtained. We
keep stirring the reaction medium until cessation of release
gaseous.

The solution of 1 - aspartyl is then allowed to cool to room temperature.
methyl phenyl alaninate and the pH of the solution is brought to the value of
6 approx.

The methyl 1 - α - aspartyl phenyl alaninate precipitates. We maintain the sus
board in a cooler for 12 hours, then separate the precipitate by filtration,
wring out the insoluble product, wash it with water and then dry it under vacuum. Cn
thus obtains 1192 of l - α -aspartyl phenyl alaninate which one could
rified by recrystallization from isopropyl ether.

We thus obtain 8gl of pure Aspartame which we define by its physical constants
The product is in the form of an odorless crystalline powder, practically
colorless, soluble in water, ethanol, methanol and sparingly soluble in
chloroform.

[α] 20 = + 12 5 (4% sol in 15 M formic acid)
seaete UV (c = 1% ethanol 96%) # max 247 m, 252 m, 258 m, 264 m
Determination of water by the method of K. Fischer concentration less than 5X.

 Protometric titer:: greater than 98S in dry product.

Microanalysis: C14 H18 N2 5 = 294.31
CH
Calculated 57.13 6.16 9.52
Found 57.06 6.14 9.68
Example IV
Stage A
Anhydride N carboxy 1 aspartic
A suspension of 40 g of aspartic acid in 250 ml of dioxane is stirred at ordinary temperature. Then phosgene is bubbled there, until the medium is saturated. The suspension is then heated to 60C until complete transformation. The anhydride N - Aspartic carboxy dissolves while the suspension changes appearance, the aspartic acid which has not reacted, transforming into its hydrochloride.

The suspension is degassed with nitrogen, then concentrated under vacuum, collecting the phosgene in a trap at low temperature. The concentrate is taken up in 100 ml of dioxane and filtered under vacuum. The filtrate is brought to dryness by evaporation under vacuum. 13 g of N-carboxy anhydride are thus collected, which is purified by recrystallization from 1,2-dichloroethane by hot and cold.

Stage B
Disodium salt of N carboxy 1. methyl aspartylphenylalaninate.

15 g of methyl phenylalaninate are dissolved in 100 ml of water. This solution cooled to 0oC is supplemented with 20 g of disodium carbonate. The suspension obtained is stirred very vigorously by an Ultraturax turbine (10000 rpm) during the addition in one go of the N-carboxy anhydride of aspartic acid.

As soon as the latter has completely dissolved (1 to 2 minutes) while maintaining the pH between 10 and 10.5 the solution is poured into dilute hydrochloric acid, while maintaining the pH at 3.5. Methyl N-carboxy l-aspartyl l-phenylalaninate decarboxylates in minutes to methyl W-aspartyl l-phenylalaninate
The latter is crystallized by adjusting the pH to 6.0 by adding sodium hydroxide. The product is separated by filtration, washing with water and then dried under vacuum.

Example V
A modification of this more advantageous procedure for industry extrapolation is as follows:
A solution of 15% methyl 1-phenyl slaninate, cooled to 0 ° C., is passed continuously through a colloid mill (for example of the Moritz type).

An addition of sodium carbonate in an amount of 2C g per 100 ml of this solution is carried out continuously. This introduction is followed, just before entering the mill, with a continuous addition of H - carboxy 1 - aspartic anhydride. The pass in the colioid grinder lasts about 20 seconds, time necessary for the dissolution and total reaction of the anhydride N-carboxy l - aspartic. At the outlet of the mill, the solution is continuously acidified to pH 3.5, by regular addition of hydrochloric acid. The solution obtained is adjusted to pH 6.0 and the product crystallizes.

Claims (7)

1 / A process for obtaining an ester dipeptide characterized in that the 1 - aspartic acid is reacted with excess phosgene to form a mixed aspartyl carbonic anhydride which is condensed with methyl phenyl alaninate in basic medium, forms a disel of the acid N -carboxy 1 aspartyl phenyl methyl alaninate then decomposes the latter in acid medium and obtains 1 - d - aspartyl phenyl alaninate methyl. 2 / A method according to claim 10 wherein the condensation of phosgene with aspartic acid is carried out in an inert oxygenated solvent. 3 / A method according to claim 10 wherein the condensation of the mixed aspartyl carbon dioxide with methyl phenyl alaninate is carried out in the presence of an alkali metal hydroxide. 4 / A method according to one of claims 1 and 30 in which the condensation is carried out at a pH varying between 10 and 11.  5 / A method according to one of claims 10 and 30 wherein the condensation of the mixed anhydride is carried out at a temperature between - 50 and + 150. 6 / A process according to claim 10 in which the hydrolysis of the disel of the N-carboxylic derivative is carried out using a mineral acid in an aqueous medium. 7 / A process for obtaining a means necessary for the process according to claim 10 characterized in that to prepare the methyl phenyl alaninate, the phenyl alanine and the methanol are reacted cold in the presence of a large excess of sulfuric acid.