WO1998046629A1

WO1998046629A1 - Hexahydroaspartame amides and method for the preparation thereof

Info

Publication number: WO1998046629A1
Application number: PCT/NL1998/000189
Authority: WO
Inventors: Gerardus Karel Maria Verzijl; Quirinus Bernardus Broxterman
Original assignee: DSM NV
Current assignee: Koninklijke DSM NV
Priority date: 1997-04-14
Filing date: 1998-04-03
Publication date: 1998-10-22
Anticipated expiration: 1999-10-14
Also published as: AU6750498A; NL1005808C2

Abstract

The invention relates to hexahydroaspartame amides, or salts thereof, of formula (1) in which R represents H or a protective group, in particular a benzyloxycarbonyl group or a t-butyloxy-carbonyl group, and to a method for preparing a hexahydroaspartame amide, in which an ester of an N-protected L-aspartyl-L-phenylalanine is amidated to give the corresponding amide and the amide obtained is then subjected to catalytic hydrogenation wherein the phenyl ring is converted into a cyclohexyl group, and to a hydrogenolysis, wherein the L-aspartyl-protecting group is removed. The hydrogenation and the hydrogenolysis can be carried out simultaneously or after one another, with the option of first carrying out the hydrogenation followed by hydrogenolysis, or the converse. If required, the hexahydroaspartame amide can then be provided with an N-protecting group, for example a BOC group. Such hexahydroaspartame amides are potentially particularly suitable for use in the preparation of pharmaceuticals, in particular (pseudo)peptides with antithrombotic action.

Description

HEXAHYDROASPARTAME AMIDES AND METHOD FOR THE PREPARATION THEREOF

The invention relates to hexahydroaspartame amides, of the formula 1

or salts thereof in which R represents H or an N- protecting group, with the proviso that the HCl-salt is excluded when R represent H.

Such hexahydroaspartame amides are particularly suitable as an intermediate in the preparation of (pseudo)peptides with antithrombotic action, for example as described in US-A-5332726. The preparation of (pseudo)peptides, as described in this patent, involves a complicated route via peptides being linked together. The novel compounds according to the invention provide for a much simpler and less expensive. synthesis route.

The invention also relates to the preparation of hexahydroaspartame amides and salts thereof. The fact is that it proved possible, starting from an inexpensive base material, the methyl ester of L-N-protected aspartyl-L-phenylalanine (N-protected

APM) , in particular the methyl ester of (Z-L-aspartyl) - L-phenylalanine (Z-APM) , to obtain the desired hexahydroaspartame amide in a simple manner, via amidation of the N-protected APM, followed by removal of the protective group on the amino group of the aspartic acid unit and hydrogenation of the phenyl ring .

Possible candidates for the N-protecting group in principle include all those protective groups which can be used in peptide chemistry, for example the protective groups described in T.W. Green, Protective groups in Organic Synthesis, John Wiley & Son, New York, 1981. Examples of frequently used protective groups include the t-butyloxycarbonyl group (BOC) , the formyl group, the fluorenylmethoxycarbonyl group (FMOC) , and the benzyloxycarbonyl group (Z) . In the method according to the invention,

N-protected APM, preferably Z-APM, is first subjected to an amidation with the aid of NH₃ to give the corresponding amide (N-protected AP-NH₂) . It was found that in contrast to unprotected APM, the amidation of N-protected APM can be carried out readily and with a high yield.

All those solvents can be used as the solvent in the amidation which are inert in the reaction and in which the N-protected APM, NH₃ and the ammonium salt of N-protected APM are reasonably soluble. Examples of solvents often used in practice include water and alcohols. Optionally, the solution can be maintained under NH₃ pressure, to increase the NH₃ concentration. The amidation of N-protected APM is preferentially carried out in an aqueous solution saturated with ammonia. The temperature at which the amidation is carried out is not critical and is preferably between 5 and 35°C, in particular between 15 and 25°C. The most important factors in determining the temperature to be employed are the occurrence of side reactions and the effect of the temperature on the solubility of the various components present in the reaction mixture.

The N-protected AP-NH₂ obtained can then have its protective group removed, in a manner known in principle, and the phenyl group can be hydrogenated. To this end, Z-APM-NH₂ for example, is subjected to a hydrogenation in the presence of H₂, so that the phenyl group becomes saturated and the protective group is removed. Both reactions can be carried out simultaneously; the removal of the protective group and the saturation of the phenyl ring can alternatively be carried out in two separate steps .

The removal of the Z group can, in principle, involve the use of the known hydrogenolysis catalysts, for example Pt, Rh or Co, in particular Pd, possibly on a support, for example a carbon or alumina support, or Ni, possibly on a support, for example on alumina or silica. Preference is given to the use of a Pd/C, in particular 2-10 wt% Pd on C. In principle, all those known solvents can be used as the solvent which are inert in the hydrogenolysis, for example alcohols, in particular methanol; water; ketones, in particular methyl isobutyl ketone (MIBK) ; esters, in particular ethyl acetate; carboxylic acids, in particular acetic acid; and mixtures of solvents, in particular CH₃OH/H₂0, MIBK/H₂0 and CH₃OH/acetic acid. By the addition of carboxylic acids, for example acetic acid, amines can be converted into salts which are more soluble in polar systems. Preference is given to the use of a methanol/water, a methanol/acetic acid or a methanol/water/acetic acid mixture.

The temperature at which the hydrogenolysis takes place is not particularly critical and is preferably between 20 and 100°C, in particular between 40 and 60°C. The H₂ pressure at which the hydrogenolysis reaction is carried out is equally noncritical and preferably varies from atmospheric pressure to 1 MPa. The hydrogenation of the phenyl ring to a cyclohexyl group can in principle involve the use of the same catalysts as with the hydrogenolysis of the Z group, for example a Pd, Raney-Ni, supported (for example on Raney-Ni Al₂0₃) Ni, Rh, Ru or Pt catalyst, in particular a supported 2-10 wt% Rh, supported 2-10 wt% Pd or supported 2-10 wt% Pt catalyst, the support used preferably being carbon or alumina. The solvent used can in principle be chosen from the same solvents as in the hydrogenolysis reaction.

The H₂ pressure at which the hydrogenation of the phenyl group is carried out is not particularly critical and varies as a function of the catalyst.

Preferably, if a Pt, Rh or Ru catalyst is employed, a pressure between atmospheric pressure and 5 MPa, in particular between atmospheric pressure and 1 MPa is used, and if an Ni or Pd catalyst is employed, a pressure between atmospheric pressure and 20 MPa is used. The temperature is noncritical too, and is preferably between 25 and 100°C.

When the hydrogenation of the phenyl ring and the hydrogenolysis are carried out simultaneously the pressure applied, if a 2-10 wt% Pd/C catalyst is employed, preferably varies from atmospheric pressure to 10 MPa at a temperature between 20 and 100°C. When a RaNi or a supported Ni catalyst is employed an H₂ pressure of between 5 and 20 MPa is preferably used at a temperature between 40 and 100 °C. In the case of the simultaneous hydrogenation of the phenyl group and hydrogenolysis another very suitable option is to employ a combination of catalysts, for example a combined Rh or Pt and Pd catalyst, preferably supported.

The hexahydroaspartame amide obtained can, if required, then be provided with a protective group on the amino group of the asparagine unit, use being made of standard methods which are frequently employed in peptide synthesis and are known to those skilled in the art. Thus, for example, a BOC protective group can be attached by bringing the hexahydroaspartame amide into contact with (BOC)₂0.

In a particular embodiment of the method according to the invention, successive process steps are carried out without intervening purification and drying of the products obtained intermediately. It was found that, using such a method, it is also possible to achieve high yields . Salts of hexahydroaspartame amides of the formula 1 are to be understood, within the scope of the present invention, as salts, for example, alkali metal salts or alkaline earth metal salts of hexahydroaspartame amides, for example Na, K, or (tetraalkyl) ammonium salts of the compounds according to formula 1 as well as acid salts of the compounds according to formula 1 where R is equal to H, for example salts of mineral acids, in particular hydrochloric acid, phosphoric acid, nitric acid or sulphuric acid. The invention will be explained in more detail with reference to the examples, without being limited thereto, however.

Example I Amidation of Z-APM to Z-AP-KH,

140 g (0.33 mol) of Z-APM were suspended in 360 ml of saturated NH₃ in water and stirred for 2 hours at 20°C. The mixture was diluted with 840 ml of H₂0. The solution was acidified to pH 2, while being cooled to 20°C, with 340 ml (400 g) of concentrated HC1. The product was filtered off and, while wet, taken up again in 1200 ml of methanol. To this solution, 19.4 g of saturated NH₃ in water were added, followed by dissolution at 70°C with some additional water. The solution was acidified with concentrated HC1 and cooled to room temperature. The product was filtered off and dried in air. Yield 122.4 g (0.03 mol), 91% of purified Z-AP-NH₂. Hydrogenolysis of Z-AP-NH to AP-NH.

50 g (0.121 mol) of Z-AP-NH₂ and 4.1 g of 5% Pd/C were suspended in a mixture of 700 ml of methanol and 300 ml of acetic acid (HOAc) . H₂ was passed through the suspension for 2 hours. 5 g of concentrated HC1 were added to the homogeneous solution and the methanol together with some of the acetic acid was evaporated under reduced pressure. Some of the acetic acid was removed azeotropically with acetone. The crystalline solid was taken up in acetone and filtered off and dried. Yield: 31.7 g (83%) of Z-AP-NH₂. The filtrate was boiled down and the oil was treated with acetone. The crystalline solid was filtered off and dried. Yield: 2.3 g. Total yield: 34.0 g (0.108 mol) (89%).

Hydrogenation of AP-NH to hexahydro-AP-NH

10.0 g (0.0317 mol) of NMR-pure AP-NH₂ were dissolved and 5.0 g of 5% Pt/C were suspended in 200 ml of 1 N HC1. The solution was stirred for 6 hours at 4 bar (0.4 MPa) of H₂, being monitored by means of TLC for complete conversion of the starting material. The catalyst was filtered off via a fluted filter.

Conversion of hexahydro-AP-NH_a to N-BOC-hexahydro-AP-NH^ The solution from the above example was set to a pH of 9.3 with 4 N NaOH while being cooled to 0°C. This solution was admixed with 0.0348 mol (1.1 eq.) of (BOC)₂0 and 100 ml of THF. The mixture was stirred overnight and the THF was then evaporated at 20°C under reduced pressure and the remaining water layer was acidified with 3 N HC1 to a pH of 2. The precipitated product was filtered off, washed with 0.1 N HC1 and dried in air. Yield: 9.73 g (0.0252 mol). Overall yield from AP-NH₂ 80%. The product was characterized by means of NMR.

Example II

Overall synthesis of N-B0C-hexahydro-AP-NH₂ from Z-APM In this example, all the reaction steps were carried out consecutively: 76.54 g (0.1731 mol) of Z-APM (content 96.8%) were weighed into 190 ml of 25% strength NH₃ aqueous. After 2 hours, 200 ml of methanol were added, followed by stirring for 1 hour. Most of the NH₃ was stripped under reduced pressure together with the methanol. The reaction mixture was diluted with 400 ml of methanol and acidified to pH 7.9 with 57.3 g of concentrated HC1. The solid was dissolved with warming to 60 to 70°C. While being cooled to 40°C the mixture was further acidified with 43 g of concentrated HCl to a pH of 1.5. The product was filtered off at room temperature. The product was free from Z-APM and Z- (L-aspartyl) -L-phenylalanine (Z-AP) , which remained in the filtrate. The yield was 100 g of wet solid which was used as such in the deprotecting operation. The input of Z-AP-NH₂ for the deprotecting operation is based on a yield of 91% in the amidation. The wet solid and 5.4 g of 5% Pd/C were slurried in 940 ml of methanol and 400 ml of acetic acid. The deprotecting operation was complete after 2 hours (monitored via TLC) . 30 g of 41% strength hydrochloric methanol were added to the mixture. Then the catalyst was filtered off and the solution was boiled down to a final weight of 80 g. The acetic acid solution of AP-NH₂ HCl was taken up in 500 ml of 1 N HCl and hydrogenated with 10.3 g of 5% Pt/C at room temperature and 0.4 MPa of H₂ over a period of 24 h. The catalyst was filtered off and washed with 150 ml of 1 N HCl. The solution of hexahydro-AP-NH₂.HCl was neutralized, being cooled to T < 20°C with 71 g of 50% strength NaOH. The product was filtered off and dried in air. Yield 38.3 g. The dried solid was taken up in 400 ml of H₂0, and at 10°C the pH was set to 9 with 6.0 g of 50% strength NaOH. 27.0 g of (B0C)₂0 (1.1 eq.) and 100 ml of THF were added, followed by stirring for 17 hours at a constant pH of 9. The solution was acidified with concentrated HCl to pH 5.3 and the product was then filtered off. The filtrate was boiled down to a volume of 200 ml (THF and water) and acidified to pH 1 with concentrated HCl. Then the remaining product was filtered off. Both preparations were dried in air. Total 35.9 g. Yield: 88%, based on the hexahydro-AP-NH₂. Overall yield from Z-APM: 54%.

Claims

C L A I M S

1 . A hexahydroaspartame amide , of the formula 1

or a salt thereof in which R represents H or an N- protective group, with the proviso that the HCl- salt is excluded when R represents H.

2. Compound according to Claim 1, in which R is H, a benzyloxycarbonyl group or a t-butyloxycarbonyl group .

3. Method for preparing a hexahydroaspartame amide, characterized in that an ester of an N-protected L-aspartyl-L-phenylalanine is amidated to give the corresponding amide and the amide obtained is subsequently subjected to a treatment in which the protective group of the amino group of the aspartic acid unit is removed and the phenyl group is hydrogenated.

4. Method for preparing a hexahydroaspartame amide, characterized in that an ester of N- benzyloxycarbonyl-L-aspartyl-L-phenylalanine is amidated to give the corresponding amide and the amide obtained is then subjected to catalytic hydrogenation in which the protective group of the amino group of the aspartic acid unit is removed and the phenyl ring is converted into a cyclohexyl group .

5. Method according to Claims 3 or 4 , wherein the removal of the protective group and the hydrogenation of the phenyl group is carried out in two separate steps.

6. Method according to Claim 4 or 5, wherein the removal of the Z group is carried out in the presence of a Pd/C catalyst.

7. Method according to any one of Claims 3-6, wherein the hydrogenation of the phenyl ring is carried out in the presence of an Rh/C or a Pt/C catalyst.

8. Method according to any one of Claims 3-7, wherein the hexahydroaspartame amide obtained is then subjected to a process in which the amino group of the aspartic acid unit is protected.

9. Method according to Claim 8, wherein a t- butyloxycarbonyl group is used as protective group of the amino group of the aspartic acid unit.

10. Method according to any one of Claims 3-9, wherein the ester used is a methyl ester.

11. Use of a hexahydroaspartame amide according to Claim 1 or 2 in the preparation of pharmaceuticals, in particular of (pseudo) peptides with antithrombotic action.