AU599926B2 - Dipeptide derivative and synthesis and use thereof - Google Patents

Description

599,926

AUSTRALIA

Patents Ack COMLETE SPECIFICATION

(ORIGINAL)

Class Int. Class Application Number: cl Lk Lodged: Complete Specification Lodged: Accepted: Published: Priority Related Art: Thj~ ocumet cocains th; amlendyments made zd.j Section 49 and is correct for printing, APPLICANT'S REF.: FP/S-38-125 Name(s) of Applicant(s): XS,,intory Limited Addresi(es) of Applicant(s): 1-40, Dojixuahama 2-chome Kita-ku, Osaka-shi, Osaka Japan o Actual Inventor(s): 0 Ot o Takaharu TANAKA Masayuki SAITO- Noaki HIGUCHI Masaki HASHIMOTO Address for Service is: PHILL IPS, ORMONDE AND FITZPATRICK Patent and Trade Mark Attorneys 367 Collins Street Melbourne, Australia, 3000 Complete Specification for the invention enititled: DIPEPTIDE DERIVATIVE AND SYNTHESIS AND USE THEREOF The following statement is a full description of this invention, including the best method of performing it known to applicant(s), P'19,3/84 -la- DIPEPTIDE DERIVATIVE AND SYNTHESIS AND USE THEREOF Background of the Invention: The present invention relates to a novel dipeptide derivative that exhibits enzyme inhibiting activity against prolyl endopeptidase (EC, 3. 4. 21. 26).

More particularly, the compounds of the invention not only exhibit enzyme inhibiting activities against prolyl endopeptidase, but also are effective for the improvement and remedy of symptoms caused by an organic disorder in the brain.

The present invention also relates to the synthesis and use of such a derivative.

The term "organic disorder in the brain" here means various symptoms caused by ischemic disorders such as sequela of cerebral infarction, sequela of cerebral hemorrhage and sequela of cerebral arteriosclerosis, and various organic disorders cuased by presbyophrenia, presenile dementia, amnesia, sequela of external wounds of the head, o 20 and sequela of cerebral operation.

:o o Prolyl endopeptidase is known to inactivate neuro- So transmitters such as Substance P, thyrotropin-releasing oo hormone (TRH) and neurotensin or vasopressin speculatively associated with memory, Tsuru and Yoshimoto of the Depart- 25 ment of Pharmaceutical Sciences, Nagasaki University, found that compounds capable of inhibiting the prolyl endopep- 0 0o tidase activity were effective for preventing experimental n amnesia caused in rats by scopolamine, and inferred that prolyl endopeptidase inhibitors have some relation to the 30 fixation of memory. This suggests the potential use of oprolyl endopeptidase activity inhibitors as anti-amnesic agents for preventing and curing amnesia.

o Actually a novel dipeptide derivative according to o the present invention which is represented by the formula as shown later was confirmed to be effective as an antiamnesic agent by animal experiments.

A biain cell retains an intracellular environment completely different from the surrounding envoironment (extracellular liquid) and lives while maintaining this difference, which requires energy to be continually produced and supplied to the cell. Most of the energy necessary for nerve cells of the brain is supplied by oxygen and glucose which are constantly supplied from the blood, since these energy sources are not stored in the brain.

If there is a disorder in the brain, and the supply of oxygen and glucose is interrupted, then energy metabolic disorders will sequentially progress and the cells will lose their functions after a time, finally causing organic collapse, and failure of normal functions.

The prevent this, brain blood vessels themselves have developped mechanisms for controlling the bloodstream so as to safely supply the energy sources for the brain tissue and maintain the external environment of the cranial nerve cells such as to be constant.

When disorders of a blood vessel in the brain are internally treated, various kinds of brain circulation improving agents, brain vasolidators, brain metabolism improving agents and the like are used. In the present state of the art, however, these agents can improve subjective symptoms but can hardly improve the symptoms of nerves.

The present inventors found compounds having antiprolyl endopeptidase activity, and disclosed them in Japanese Patent Laid-Open No. 37764/1986. Tsuru et al. also discloses compounds having said activity in Japanese Patent Laid-Open No. 188317/1985.

Summary of the Invention: The present inventors have made various studies to find a compound having anti-prolyl endopeptidase activity and anti-amnesic activity which are considered associated with the improvement and remedy of symptoms caused by various disorders in the brain described above. Further, to find a novel compound of sufficiently low toxicity, they 35 synthesized compounds which have structures similar to natural substances by combination of fatty acidswith amino acids or peptide compounds which are highly safe as natural compounds.

)o 001~ O 0i 0 0 0 0 -3- As a result, it has been found that a novel compound according to the present invention which is represented by the general formula has anti-prolyl endopeptidase activity. This compound exhibits the strongest anti-prolyl endopeptitase activity among the anti-prolyl endopeptidase compounds heretofore dicovered (Tadashi yoshimoto, Agr.

Biol. chem. Assoc. (Japan), Synopses of Lectures, pp 752 to 754, (1983), and Japanese Patent Applicaiton No.

160994/1984, filed on Jul,! 31, 1984).

The present inventors have also found that a novel dipeptide derivative represented by the general formula (1) having anti-prolyl endopeptidase activity has anti-amnesic effects on laboratory animals.

The present invention has been accomplished on the basis of these findings.

A novel dipeptide derivative according to the present invention which is represented by the formula is effective for the improvement and remedy of mental symptoms caused by organic disorders in the brain and, especially, for improvement and remedy of amnesia. The compounds according to the present invention represented by the formula differ greatly from the known piracetam derivatives based anti-amnesic agents in that the Sformer contains a proline group. Because of this feature, e. 25 the compounds of the formula present extremely low toxicity levels in organisms.

Detailed Description of the Invention: The novel dipeptide derivatives of the present invention are represented by the formula

R

1 (CH2) CO-N-CH-CO-N (1) (CH2) n

-R

S 30 (wherein m is an integer of 3 to 8; n is 0 or an integer of 1 to 6; R 1 is hydrogen atom; R 2 is hydrogen atom, a branched alkyl group having 3 to 5 carbon atoms, phenyl grovp, 1 2 hydroxyphenyl group or methylthio group, or R an., i 2 S'/t together represent a single bond between carbon atom and

J

-4- 3 '5 nitgogen atom; and R 3 4 nowr alky l-str grou.- hyd-roeKy- CAAO; 2 Mef_*yl- 4 -guoPa a ag provided that when n is 0, R represents one of the above-described groups except hydrogen atom).

When R 2 is the branched alkyl group, it can be, for examle, isopropyl, sec-butyl, tert-butyl, 1-methylbutyl, 1ethylpropyl or 1,1-dimethylpropyl, among which isopropyl is particularly preferred.

methoxycarbonyl group, ethoxyca group, propoxycarbonyl group, isopropox c group, butoxycarbonyl group, sec- Se ny-l-g-up-ad- t- a y btox'grcup.

One preferred group among the compounds of the present invention of the formula is expressed by the formula:

H

(CH2) CO-N--CH--CO--N

(CH

2 )n-R R

R

(whrein, m, n and R 3 have the same meanings as given above; and R is hydrogen atom, a branched alkyl group having 3 to carbon atoms, phenyl group, hydroxyphenyl group or methyl- R4 thio group, provided that when n is 0, R represents one of said groups except hydrogen atom).

i R4 In this embodiment, R when n is 0 is preferably a branched alkyl group having 3 to 5 carbon atoms.

Another preferred group of compounds of the invention are expressed by the formula:

(CH

2 )m -CO-N 3 (wherein m and R 3 have the same meanings as given above).

The following compounds of the formula are particularly preferred because of their high anti-prolyl endopeptidase activities. The following compounds will _PLt_

IV

~~A

sometimes be referred to by the respective numbers indicated in the parentheses shown hereinunder.

H O 0 O CCf Y (SUAM 1333) (SUAM 1234) (SUAM 1345)

SHO

CH3 Yi

CHO

CHO

(SUAM 1231) ~1

N

N0 0!

N

CHO

0

N

o N

CHO

0 0 N (SUAM 1336) (SUAM 1339) (SUAM 1342) Q a~ a~ H 0 N

N

0 03

CHH

H Y, N N 0 CH CH 3

CHO

(SUAM 1110) (SUAM 1117)

H

N

(SUAM 1115) CH 3 -7- The compounds according to the present inveniton are prepared by a general peptide synthesis method, but they are more conveniently synthesized by a method according to the present inveniton which will be explained in the following.

Explanation of the abbreviations Z benzyloxycarbonyl group Boc tert-butyloxycarbonyl group Pro proline residue -Pro-OMe: -N,

COOCH

3 -Pro-CH OH: -N 2

CH

2

OH

-Pro-CHO: -N

CHO

Ala alanine residue Val valine residue Leu luecine residue norLue norleucine residue Phe phenylalanine residue 0o 0 Met methionine residue OMe methylester residue WSCD N-ethyl-N',N'-dimethylaminopropyl carbodiimide TEA triethylamine The compounds of the formula of the present invention may be synthesized by the following procedures: If the compounds have the formula (la) wherein R in formula is a lower alkyl ester group of -COOB: 1

(CH

2 CH-CO-N (la)

COOB

1 2 (wherein m, n, R and R respectively, represent the same meanings as in the formula and B represents a lower -8alkyl group), they may be prepared by reacting dipeptide represented by the formula

R

1 HN-CH-CO-N (2)

(CH

2 )n-R C

COOB

1 2 (wherein n, R R and B, respectively, represent the same meanings as in the formula and carboxylic acid, acid halide or acid anhydride represented by the formula in the presence of a base: 0_(CH 2 )m-CO-A (3) (wherein m represents the same meaning as in the formula and A represetns hydroxyl group, a halogen atom, or the following group -0--CO-(CH 2 m (wherein m represents the same meaning as the above).

This synthesis utilized the conventional acylating reaction of an amino group, and reagents such as a base s, differ depending upon which of carboxylic acid, acid halide o' and acid anhydride is used as the starting compound. For 15 example, if acid halide is used, trialkylamine such as triethylamine is preferable as a base, but the aqueous solution of alkali metal hydroxide, alkali metal carbonate or the like may be used.

When acid anhydride is used, the above-described aqueous solution of alkali metal hydroxide, sodium hydroxide and potassium hydroxide, or alkali metal carbonate such as sodium carbonate and potassium carbonate is used as a base.

When carboxylic acid is used, a condensing agent such as WSCD and dicyclohexyl carbodiimide is used in an organic solvent which is inert in the reaction.

If the compounds have the formula (lb); -9-

R

1 (CH CO-N- CH--CO--N 2 n-Rl ib)

(CH

2 )n-R (Cb) (wherein m, n, R and R are the same as above), they are readily obtained by treating the compound (la) with a reducing agent. For example, an alcohol form (lb) is obtained at a high yield by adding methanol dropwise to a suspension of the compound of the formula (la) and sodium borohydride in tertiary butyl alcohol or tetrahydrofuran.

An alcohol form (Ib) is also obtained by adding a reducing agent such as sodium borohydride, lithium borohydride, zinc borohydride, and potassium borohydride to the alcohol solution of the compound of the formula (la).

The prolinal derivative having the formula (lc): 1 S (CH CH--CO-N (lc)

(CH

2 )nR

CHO

1 2 (wherein m, n, R and R are the same as above) is obtained by treating the compound of the formula with an oxidizing agent in an organic solvent.

The solvent used in this reaction may be anything that does not participate in the reaction, but dimethyl sulfoxide is the most preferable. As an oxydizing agent, a complex of sulfur trioxide-pyridine complex, dimethyl sulfoxide, dimethyl sulfoxide-dicyclohexyl carboimide-phosphoric acid, silver oxide and manganese dioxide may be mentioned.

When the effects of these compounds for preventing decomposition of Z-glycyl-prolyl-B-naphtylamide by prolyl endopeptidase were examined, they exhibited very high anti- 4 prolyl endopeptidase activities as will be shown in the S 25 later-described example 4, and no inhibitory activity to S proteinases such as papain, bromelain, trypsin, chymotrypsin, thermolysin and pepsin.

These compounds obtained in this way are novel and have anti-amnesic effects.

The present invention will be explained in more detail with reference to the examples.

Preparation H-Phe-Pro-OMe Z-Phe-OH (1 equivalent), H-Pro-OMe hydrochloride (1 equivalent) and TEA (1 equivalent) were dissolved in dry methylene chloride, and WSCD (1 equivalent) was added dropwide under cooling with ice. After being stirred for hours at re ,m temperature, the reaction mixture was washed successively with IN HC1, water, saturated aqueous sodium bicarbonate, water and brine, and was dried over anhydrous magnesium sulfate. The solvent was distilled off under vacuum, and the resulting residue was purified by column chromatography on silica gel. Z-Phc-Pro-OMe (1 equivalent) obtained was dissolved in ethanol (1 equivalent) and boron trifluoride etherate complex (1 equivalent) and palladium on carbon (trace) were added to remove the Z group by catalytic reduction in an atmosphere of hydrogen. The solvent was distilled off under vacuum to obtain the end compound.

In place of Z-Phe-OH, a) Z-Met-OH, b) Z-norLeu-OH, c) Z-Ala-OH, d) Z-Val-OH, and e) Z-Leu-OH were used as the starting compounds and treated by the procedures described above to obtain H-Met-Pro-OMe, H-norLeu-Pro-OMe, H-Ala-Pro-OMe, H-Val-Pro-OMe and H-Leu-Pro-OMe, respectively. All of these compounds were in t~h form of an oil.

H-Pro-Pro-OMe Z-Pro-OH (1 equivalent), H-Pro-OMe hydrochloride (1 equivalent) and TEA (1 equivalent) were dissolved in dry methylene chloride, and WSCD (1 equivalent) was added dropwise under cooling with ice. After being stirred for hours at room temperature, the reaction mixture was washed successively with IN HC1, water, aqueous sodium bicarbonate, water and brine, and was dried over anhydrous magnesium sulfate. The solvent was distilled off under vacuum, and the resulting residue was purified by column chromatography on silica gel. The Z-Pro-Pro-OMe obtained was dissolved (1 equivalent) in ethanol, and boron trifluoride etherate i -11complex (1 equivalent) and palladium on carbon (trace) were added to iemove the Z group by catalytic reduction in an atmosphere of hydrogen. The solvent was distilled off under vacuum to obtain the end compound as an oil.

Example 1 N-(4-phenyl)butanoyl-Phe-Pro-OMe (SUAM 1232) H-Phe-Pro-OMe (1 equivalent) and TEA (1 equivalent) were dissolved in dry tetrahydrofuran, and 4-phenylbutanoyl chloride (1 equivalent) was added dropwise under cooling with ice. The mixture was stirred for 6 hours at room temperature, and the hydrochloride of TEA separated out was removed by filtration. The solvent was distilled off under vacuum and the resulting residue was dissolved in a small amount of ether. The ethereal solution was washed successively with IN HC1, brine, saturated aqueous sodium bicarbonate and brine, and thereafter was dried over anhydrous magnesium sulfate. The solvent was removed under vacuum and the residue was purified by column chromatography on silica gel to obtain the required compound (an oily compound).

In place of H-Phe-Pro-OMe, a) H-Met-Pro-OMe, b) HnorLeu-Pro-OMe, c) H-Ala-Pro-OMe, d) H-Val-Pro-OMe, and e) H-Leu-Pro-OMe were used as the starting compounds and treated by the procedures described above to obtain N- (4-phenyl)butanoyl-Met-pro-OMe (SUAM 1343), N-(4phenyl)butanoyl-norLeu-Pro-OMe (SUAM 1235), N-(4phenyl)butanoyl-Ala-Pro-OMe (SUAM 1098), N-(4phenyl)butanoyl-Val-Pro-OMe (SUAM 1112) and N-(4pehnyl)butanoyl-Leu-Pro-OMe (SUAM 1113), respectively. All of these compounds were oily.

N-(4-phenyl)butanoyl-Pro-Pro-OMe (SUAM 1229) H-Pro-Pro-OMe (1 equivalent) and TEA (1 equivalent) were dissolved in dry tetrahydrofuran, and 4-pehnylbutanoyl chloride (1 equivalent) was added dropwise under cooling with ice. The mixture were stirred for 6 hours at coom temperature, and the hydrochloride of TEA sep-rete ut removed by filtration. The solvint, was dJtiX8 vacuum. The resulting residue was dissolve amount of ether, was washed successively with

I

-12brine, saturated aqueous sodium bicarbonate and brine, and thereafter was dried over anhydrous magnesium sulfate.

After being condensed under a reduced pressure, the remaining product was purified by column chromatography on silica gel to obtain the required compound (an oily compound).

In place of 4-phenylbutanoyl chloride, a) 2phenylacetyl chloride, b) 3-phenylpropionyl chloride and c) 5-phenylpentanoyl chloride were used as the starting compounds and treated by the procedures described above to obtain N-(2-phenyl)acetyl-Pro-Pro-OMe (SUAM 1334), b') N-(3-phenyl)propionyl-Pro-Pro-OMe (SUAM 1337) and phenyl)pentanoyl-Pro-Pro-OMe (SUAM 1340), respectively. All of these compounds were oily.

Examp.e N-(4-phenyl)butanoyl-Phe-Pro-CH 2 OH (SUAM 1233) N-(4-phenyl)butanoyl-Phe-Pro-OMe (SUAM 1232) (1.3 g) obtained in Example 1 and sodium borohydride (460 mg) were dissolved in tertiary butyl alcohol (20 ml) and heated to C with stirring. Absolute methanol (3.4 ml) was added dropwise under reflux, and thereafter the mixture was refluxed for 2 hours with stirring. After reaction, the heated mixture was cooled to room temperature and several milliliters of water was added under cooling with ice to inactivate unreacted sodium borohydride. After methanol and tertiary butyl alcohol were distilled off under vacuum, the residue was extracted with ethyl acetate. The extract was washed with brine and dried over anhydrous magnesium sulfate, The solvent was distilled off under vacuum and the resulting residue was purified by column chromatography on silica gel to obtain the end compound as an oil (1.2 g).

In place of N-(4-phenyl)butanoyl-Ph(-Pro-OMe, a) N- (4-phenyl)butanoyl-Met-Pro-OMe, b) 'T 4-phenyl)butanoylnorLeu-Pro-OMe, c) N-(4-phenyl)butanoyiXAla-Pro-OMe, d) N- (4-phenyl)butanoyl-Val-Pro-OMe and e) N-(4-phenyl)butanoyl- Leu-Pro-OMe were used as the starting compounds and treated by the procedures described above to obtain N-(4phenyl)butanoyl-Met-Pro-CH 2 OH (SUAM 1344), N-(4- -6 111 I I I Y trf I I i -13phenyl)butanoyl-norLeu-Pro-CH 2 OH (SUAM 1236), N-(4phenyl)butanoyl-Ala-Pro-CH 2 OH (SUAM 1109), N-(4phenyl)butanoyl-Val-Pro-CH 2 OH (SUAM 1116) and N-(4pehnyl)butanoyl-Leu-Pro-CH 2 OH (SUAM 1114), respectively.

All of these compounds were oily.

N-(4-phenyl)butanoyl-Pro-Pro-CH 2 OH (SUAM 1230) A mixture of the N-(4-phenyl)butanoyl-Pro-Pro-OMe (SUAM 1229) (1.3 g) obtained in Example 1 and sodium borohydride (510 mg) was dissolved in tertiary butyl alcohol (20 ml) and heated to 80 C with stirring. Absolute methanol (3.8 ml) was added dropwise under reflux, and thereafter the mixtue was refluxed for 2 hours with stirring. After reaction, the heated mixture was cooled to room temperature and several mililiters of water was added under cooling with ice to inactivate unreacted sodium borohydride. After methanol and tertiary butyl alcohol were distilled off under vacuum, the residue was subjected to extraction with ethyl acetate. The extract was washed with brine and dried over anhydrous magnesium sulfate. The solvent was distilled off under vacuum and the resulting crude prOduct was purified by column chromatography on silica gel to obtain the end compound as an oil (1.2 g).

o In place of N-(4-phenyl)butanoyl-Pro-Pro-OMe, a) N- (2-pehnyl)acetyl-Pro-Pro-OMe, b) N-(3-phenyl)propionyl-Pro- Pro-OMe and c) M-(5-pehnyl)pentanoyl-Pro-Pro-OMe were used as the starting compounds and treated by the procedures described above to obtain a')-N-(2-phenyl(acetyl-Pro-Pro-

CH

2 0H (SUAM 13359, N-(3-phenyl)propionyl-Pro-Prop-CH 2

OH

(frAM 1338) and N-(5-phenyl)pentanoyl-Pro-Pro-CH 2

OH

(SUAM 1341), respectively. All of these compounds were oily.

Example 3 N-(4-phenyl)butanoyl-Phe-Pro-CHO (SUAM 1234) A mixture of N-(4-phenyl)butanoyl-Phe-Pro-CH2OH (SUAM 1233) (1.2 g) obtained in Example 2 and TEA (1.6 ml) was dissolved in anhydrous dimethyl sulfoxide (8 ml), and to the stirred solution, a solution of sulfur trioxide-pyridine complex (1.9 g) in dimethyl sulfoxide (8 ml) was added.

-27- Example 6 -14- After stirring the mixture at room temperature for about minutes, the reaction solution was poured into iced water (100 ml) and subjected to extraction with ethyl acetate.

The extract was washed successively with 10% aqueous citric acid, brine, saturated aqueous sodium bicarbonate and brine, and dried over anhydrous magnesium sulfate. After distilling off the solvent under vacuum, the resulting crude product was purifed by column chromatography on silica gel to obtain the end compound as an oil (800 mg).

In place of N-(4-phenyl)butanoyl-Phe-Pro-CH2 OH, a) N- (4-phenyl)butanoyl-Met-Pro-CH 2 OH, b) N-(4-phenyl)butanoylnorLeu-Pro-CH2OH, c) N-(4-phenyl)butanoyl-Ala-Pro-CH 2 OH, d) N-(4-phenyl)butanoyl-Val-Pro-CH20H and e) N-(4phenyl)butanoyl-Leu-CH 2 OH were used as the starting compounds and treated by the procedures described above to obtain N-(4-phenyl)butanoyl-Met-Pro-CHO (SUAM 1345), b') N-(4-phenyl)butanoyl-norLeu-Pro-CHO (SUAM 1333), N-(4phenyl)butanoyl-Ala-Pro-CHO (SUAM 1110), N-(4phenyl)butanoyl-Val-Pro-CHO (SUAM 1117) and N-(4phenyl)butanoyl-Leu-Pro-CHO (SUAM 1115), respectively. All of these compounds were oily.

N-(4-phenyl)butanoyl-Pro-Pro-CHO (SUAM 1231) A mixture of N-(4-phenyl)butanoyl-Pro-Pro-CH 2 OH (SUAM 1230) (1.3 g) obtained in Example 2 and TEA (2.0 ml) was dissolved in anhydrous dimethyl sulfoxide (8 ml), and to the stirred solution, a solution of sulfur trioxide-pyridine oo complex (2.3 g) in dimethyl sulfoxide (8 ml) was added.

After stirring the mixture at a room temperature for about minutes, the reaction solution was poured into iced water S 30 (100 ml) and subjected to extraction with ethyl acetate.

The extract was washed successively with 10% aqueous citric acid, brine, saturated aqueous sodium bicarbonate and brine, and dried over anhydrous magnesium sulfate. After S distilling off the solvent under vacuum, the resulting crude product was purified by column chromatography on silica gel to obtain the end compound as an oil (700 mg).

I

In place of N-(4-phenyl)butanoyl-Pro-Pro-CH 2 OH, a) N- (2-phenyl)acetyl-Pro-Pro-CH 2 OH (SUAM 1335), b) N-(3phenyl)propionyl-Pro-Pro-CH 2 OH (SUAM 1338) and c) phenyl)pentanoyl-Pro-Pro-CH 2 OH (SUAM 1341) were used as the starting compounds and treated by the procedures described above to obtain N-(2-phenyl)acetyl-Pro-Pro-CHO (SUAM 1336), N-(3-phenyl)propionyl-Pro-Pro-CHO (SUAM 1339) and N-(5-phenyl)pentanoyl-Pro-Pro-CHO (SUAM 1342), respectively. The analytical data for the compounds obtained in Examples 1 to 3 are listed in Table 1.

All the compounds shown in Table 1 are oily and soluble in carbon tetrachloride, ether, chloroform, methylene chloride, ethyl acetate and methanol.

When the effects of these compounds for inhibiting decomposition of Z-glycyl-prolyl-B-naphthylamide by prolv 1 endopeptidase were examined, they exhibited very high ai,..

prolyl endopeptidase activities as will be shown in the later-described Example 4, and no inhibitory activity to proteinase such as papain, bromelain, trypsin, chymotrypsin, thermolysin and pepsin.

These compounds obtained in this way are novel and have anti-amnesic effects as shown in Example 0 0 i 8: a, £?0 Table 1 ExapleSUAM No. Property IR Spectrum Proton NMR Spectrum[a IM No. (Film, ai-) in.CC 3 (ppm) from TMS D (/Z 3300, 1740, 1-70-2.20(8H,m), 2.54(2H,m), 11630, 1530, 3.00(2H,m), 3.43-3.67(2H,m), 1232 oily 1450, 1190, 3.73(3H,S), 4.44(lH,m), 1170, 740, 4.96(lH,m), 6.16(lH,d,J=7.8Hz), 700 7.13(511,s), 7.24(5H,s) 3300, 2940, 1.62-.2.29(10H,m), 2.05(3H,s), 1 2920, 2870, 2.29-2.72(4H,m), 3.30-3.93 1343 oily 1740, 1620 3.63(3H,s), 4.38(lH,m), 1520, 1440, 4.94(lH,m), 6.77(lH,d,J=7.8Hz), 12 7.14 3270, 960, a 270, 2960, 0.92(3H,m), l.16-1.60(6H,m), 1 2870, 1740, 1.77-2.44(8H,m), 2.64(2H,m), 1235 oily 1660, 1530, 3.52(2H,m), 3.66(3H,s), 1440, 110, 447(1H,m), 4.-7 (lH,m), 170, 30 7.16(5H,s), 7.5 (1H,d,J=9.OHz) 3300- 2950, 1.35(3H,d,J=h7Hz 2 2870, 1740, 1.90-2.22(8H,m):, 2.62(2H,m), [a]D 1 1620, 1530, 3.51-3.78(2H,M)', 3.66(3H,S), 1098 oily 1450, 1190, 4.46(1H,in), 4.7 f6(1H,m), -71.80 346 1170, 740, 6.85(1H,brd,J=9Hz), (c=0.17, 700 7.16(5H,s) in CHC1 3 Table 1 (cont'd) Example No.

SUAM No. Property IR Spectrum (Film, am- 1 Proton NMR Spectrum in CDC1 3 6 (ppm) from TMS

EI-MS

(m/z) 3300, 2950, 0.94, 1.03(3H each, both d, 26 2870, 1740, J=:7Hz), 1.80-2.37(9H,m), 2.63 [a] 1 oily 1620, 1530, 3.53-3.78(2H,), 3.67 374 111 1430, 1190, 4.48(1H,m), 4.61(1H,dd, -65.00 1170, 750, J=7Hz, 9Hz), 6.40(lH,brd, (c=1.71, 690 J=9Hz), 7.18(5H,s) in CHC1 3 3300, 2950, 0.95, 1.10(3H each, both d, [a]26 2870, 1730, J=6Hz), 1.60(3H,m), 1.84-2.20 D 1 1113 oily 1620, 1530, 2.63(2H,m), 3.55-3.73 -57.80 388 1440, 1190, 3.68(3H,s), 4.48(1H,m), (c=1.56, 1170, 740, 4.88(1H,m), 6.27,(lH,brd,J=9Hz), in CHC1 3 690 7.18(5H,s) 2950, 2870, 1 1740, 1640, 1.86-2.36(12H,m), 2.63(2H,m), 1229 oily 1430, 1320, 3.34-3.88(4Hm), 3.66(3H,s), 1190, 1170, 4.60(2H,m), 7.18(5H,s) 740, 700 2970, 2940, 1.71-2.46(8H,m), 12870, 1730, 3.38-3.93(4H,m), 3.67(3H,s), 1334 oily 1630, 1420, 3-67(2H,s), 4.57(2H,m), 1190, 1170, 7.22(5Hs) 740, 690 Table 1 (cont'd) Example SUA No. Property IR SpcrmProton NMR Spectrum [a I-MS No. (Film, am) in CDCl 3 6 (ppm) from TMSTC E mz 2970, 2940, 1.75-2.32(8H,m), 1 2870, 1730, 1337 oily 1630, 1430, 3.22-3.96(4H,m), 1190, 1170, 3.66(3H,s), 4.57(2H,m), j740, 700 7.18(5H,s) 2940, 2870, 1.53-1.85(4H,m), 11730, 1630, l.90-2.31(10H,m), 1340 oily 1420, 1190, 2.43-2.80(2H,m), 1170, 740, 3.32-3.94(4H,m), 3.65(3H,s), 700 4.53(2H,m), 7.14(5H,s) 3370, 3280, l.50-2.31(8H,m), 2.62(2H,m), 2 2930, 2830, 3.02(2H,m), 3.34-3.72(4H,n), 1233 oily 1620, 1530, 4.14(2H,m), 4.94(1H,m), 1440, 1040, 6.64(lH,d,J=-7.8Hz), 740, 690 7.17(5H,S), 7.23(5H,s'.

3380, 3280, 1.62 '2.31(10Ii,m), 2.03(3H-,s), 2 2950, 2870, 2.31-2.87(4fl,m), 1344 oily 1610, 1440, 3.18-3.74(4H,m), 4.06(lH,m), 1040, 750, 3.97-5.15(2H,m), 7.13(5H,s) 690 3400, 3280, 0.88(3Hm), l.l0-l.62(6H,m), 2960, 2870, l.68-2.32(8H,m), 2.62(2H,m), 2 1650, 1620, 3.35-3.66(4H,n), 3.98(lH,m), 1236 oily 1540, 1450, 4.44-4.90(2H,m), 1050, 750, 6.88(lH,d,J=7.8Hz), 700 7.18(5H,S) Table 1 (cont'd) Example SUAM No. Property IR Spectrum Proton NMR Spectrum []EI-MS No. (Film, am- 1 in CDC1 3 iS (ppm) from TMS 24 D j(m/z) 3330, 3300, l.34(3H,d,J=7Hz), [CL24 2 2940, 2870, 1.84-2.21(8H,m), 2.63(2H,M),D 1109 oily 1620, 1530, 3.56(4H,m), 4.18(lH,M), -62.00 316 (Mt) 1450, 1050, 4.49-4.86(2H,mn), 6.72(1iH,brd, (c=0.56, 740, 690 J=9Hz), 7.17(5H,s) in CHCl 3 33o0 30, .95, 1.00(3H each, both d, [a 23 2950, 2860, J=7Hz), 1.75-2.22(9H,m), 2.65 [JD 21116 Oily 1610, 13,(2H,m), 3.58(4H,in), 4.15(1H,M), _32.40 346 1440, 1040, 4.54(1H,t,J=4Hz), 4.61(1H,dd, (c=1.02, 740, 690 J=7Hz, 9Hz), 6.33(1H,brd, in CHC1 3 J=9Hz), 7.18(5H,s) 0,94, 0-98(3H each, both d, 2 3380, 3280, J=-6Hz), 1.56(3H,m), [a] 24 2 2950, 2860, 1.75-2.38(8H,m), 2.63(2H,m), D361 1114 oily 1620, 1540, 3.58(4H,m), 4.18(1H,m), -33.10 1440, 1050, 4.48(lH,t,J=4Hz), 4.86(1H,m), (c=1.59, 740, 690 6.28(1H,brd,J=9Hz), in CHC1 3 7.17 3400 (br), 2940, 2820, l.80-2-43(12H,m), 2.67(2H,m), 2 13 Oiy 1720, 1620, 3.33-3.73(6H,m), 4.10(lH,m), 13oiy 1450, 1320, 4.63(1?H,m), 5.08(1H,m), 1235, 1040, 7.18(5H,s) 740, 700

I

Table 1 (cont'd) Example SUMN.Poet R Spectrum Proton NMR Spectrum El-MS N. SNo. o Prpty (Film, aim-) in CDCl 3 6 (ppm) from TmS LQJD (m/z) 2 3400, 2960, 1.65-2.82(8H,m), 1335 143 280, 1040, 3 31-4.33(7H,m), 3.68(2H,s), 7430, 690, 4.49-5.12(2H,m), 7.22(5H,s) 3400, 2970, 1. 66-2.30 (8H,m) 2 13 Oiy 2870, 1620, 2.42-3.13(4H,m), 13oiy 1430, 1050, 3.28-4.40(7H,m), 740, 690 4.46-5.25(2H,m), 7.20(5H,s) 3400, 2930, l.48-2.40(14H,m), 2 14 oiy 2870, 1620, 2.40-2.78(2H,m), 14 oiy 1430, 1050, 3.16-4.32(7H,m), 740, 690 4.35-5.19(2H,mi), 7.l0(5H,s) 3300, 2930, 1.71-2.02(6H,m), 2.15(2H,m), 2870, 2850, 2.55(2H,m), 3.06(2H,m), (a 1234 oily 1630, 1540, 3.28-3.86(2H,m), 4.20(1H,m), 130, 540,5.02(lH,m), 7.18(5H,s), i1440.1 1060, 7.25(5H,s), 9.l9(lH,d,J--2.0Hz) 74,700 3300 2920, 1.67-2.32(lOH,m), 3.l0(3H,s), 2880, 2720, 2.32-2.83(4H,m), 31345 oily 1730, 1620, 3.40-3.90(2H,m), 1530, 1450, 4.03-5.26(2H,m), 7.19(5H,s), 750, 700 9.38 (lH,d,J=2..OHz) Table 1 (cont'd) Example SMNo. [Property IR Spectrum Proton NMP Spectrum []El-MS No. 51A 1 Y (Film, am- 1 in ODC1 3 6 (ppm) from TMS D (m/z) 3300, 2960, 0.88(3H,in), 1.10-i-.60(6H,m), 3 2870, 2730, l.68-2.38C8H,m), 2.13(2H,m), 1333 oily 1730, 1620, 3.25-3.82(2H,mil, 4.53-4.98 1540, 1440, 6.83(1H,d,J=7.8Hz), 740, 700 7.16(5H,s), 9.35(1H,d,J=2.OHz) 3300, 2970, 1.40(3H,d,J6GHz), 1.88-2.28 [ca] 24 3 2870, 2720, 2.65(2H,m), 3.56-3.80

D

1110 Oily 1720 1620, 4.52(1H,in), 4.78(1H,m), -62.0' 316 1530, 1450, 6.35(lH,brd,J=9Hz), 7.21(5H,s), (c=0.56, 740, 690 9.49 (lH,d,J=2Hz) in CHC1 3 330, 260,0.95, 1.02(3H1 each, both d, [a 31 3300, 2960, J=7Hz), 1.32-2.27(9H,m), 2.61D (a 1117 28y 730, 2720, 3.52-3.83(21,m), 4.441 -65.00 345+l (a)1730, 1620, 4.62(1H,dd,J=7Hz, 9Hz), 0c(l.00, 153, 140, 6.63(lH,brd,J=9Hz), 7.18(5H,s), inCHl3) 740, 690 9.46(lH,d,J=1.5Hz)inC13 328, 250,0.95, 1.01(3H each, both d, ri( 31 32860, 2950, J=6Hz), 1.57(3H,m), 1.85-2.41

D

31115 Oily 1720, 1620, 2.62(2H,m), 3.45-3.83 44.20 358 1530, 1450' (2H,rn), 4.48(1H,m), 4.82(1H,m), (c=0.33, 740~~ 69 6.40(1H,brd,J=-9Hz),inC13 740 690 7.18(5H,s), 9.45(11,d,J=2Hz)in C 13 Table 1 (cont'd) Example IR Spectrum Proton NMR SpectrumE-S Mo. SUAM No. Property (F im, am-1) Iin CDCl 3 6 (ppm) from TMS (OID IM/z 2970, 2870, 3 1730, 1630, l.88-2.43(12H,m), 2.63(2H,m), 1231 oily 1440, 1320, 3.28-3.82'41,M), 4.57(2H,m), 2720, 740, 7.13(SH,s), 9.49(lH,d,j=l.0Hz) 700 2970, 2880, l.72-2.43C8H,m), 3 13 oiy 1720, 1640, 3.27-3.95(4H,m), 3 .67(2H,s), 13 oiy 1420, 740 4.63(2H,rn), 7.22(5H,s), 690 9.42(1H,d,j=l.OHz) 2970, 2870, 1.66-2.32(8H,mi), 3 2800, 2700, 2.42-3.13(4H,m), 1339 oily 1720, 1630, 3.30-3.98(4H,m), 1430, 740, 4-53(2H 7.19(5H,s), 690 19. 43 (IlId,d, J=l.0Hz) oily 1342 2970, 2800, 1720, 1430, 700 2880, 2710, 1640, 740, l.51,-2.46(14H,M), 2.4 6-2. 83 (2H,m), 3. 20-3. 92 (4H,m) 4.52(2H,m), 7.14(5H,S), 9.39 (lH,d,J=2.OHz) L -23- Example 4 Measurement of Anti-prolyl Endopeptidase Activity The method of Yoshimoto and Tsuru Yoshimoto and D. Tsuru, Agr. Biol. Chem. 42,2417,1978) was used to measure the anti-prolyl endopeptidase activities of several compounds according to the present invention. A mixture of 0.0025M Z-glycyl-proline-8-naphthylamide (0.25 ml), 0.1M phosphate buffer (pH 7.0; 0.99 ml) and a solution of a particular anti-prolyl endopeptidase compound (0.01 ml) was incubated in a test tube at 37 0 C for 3 minutes. Thereafter 0.1 m of a solution of prolyl endopeptiuse (0.2 U/ml) was added and the mixture was heated at 35 0 C for 10 minutes.

After the reaction, 2.0 ml of Triton x-100 in 1M acetate buffer (pH 4.0) was added to the reaction mixture until the final concentration of the surfactant was 10%. The mixture was left at a room temperature for 15 minutes and the absorbance at 410 nm was measured.

A sample for a blind test was prepared by using the buffer solely in place of the anti-prolyl endopeptidase comound and its absorbance was alsotmeasured. The percent inhibition of prolyl endopeptidase was calculated by the formula: [(b-a)/b]xl00, and the amount of a specific compound needed to achieve 50% inhibition (IC 50 was determined. The results are shown in Table 2.

Table 2 Compound IC 50 (ng/test tube) SUAM 1231 0.2 SUAM 1234 0.3 SUAM 1333 0.4 SUAM 1345 0.7 SUAM 1336 SUAM 1339 0.4 n SUAM 1342 0.2 SUAM 1115 0.3 SUAM 1117 0.2 -24- Example Measurement of Preventive Effect against Experimental Amnesia caused in Rats by Scopolamine (Intraperitoneal Administration) Several of the anti-prolyl endopeptidase compounds according to the present invention were checked for their ability to prevent the inhibition of long-term memory fixation by scopolamine. Solutions of physiological saline which contained 20 pg/kg of selected compounds of the present inveniton were administered intraperitoneally once a day to Wister male rats (100 to 120 One hour after the administration, electric shocks were applied to the rats so that they would acquire passive avoidance learning. Immediately thereafter, scopolamine was administered intraperitoneally to each rat in an amount of 3 mg per kg of body weight.

The results of the test were assessed both 24 hours and 48 hours after the administration of scopolamine. The number of amnesic rats and of sound rats was counted for each of the control group (rats to which the test compounds were not administered but to which only scopolamine and physiological saline were administered intraperitoneally) and the treated group (rats to which both the test comound and scopolamine were administered). The results are shown in Table 3. With respect to SUAMs 1110, 1115 and 1117, administration of 1 mg/kg did not manifest the desired effect but when 0.1 mg/kg and 0.025 mg/kg of each was administered, very strong effects were produced. SUAMs 1231 and 1333 also exhibited a strong effect at 0.020 mg/kg.

Thus these compounds exhibited very strong anti-amnesic effects in a low dose without producing any apparent change in the rats when the dose was raised up to more than 100 mg/kg.

Table 3 :Amnesia test with rats Cintraperitoneal administration) leIarning Avoidance time Pharmacological (sec.) effects Drug of Initial 1b. of Dern Sample administered rats a-oid- avoid- Merul. of ecafe erin etdtne acs ime age ftr anei eret afterim lerig tstdge nesAtr Ate mei time during (sec.) 24 hrs. 48 brs. rats/lb. ofamei (sec.) learning rats testedamei 1) physiological saline 2) physiological saline~ 3) SIAM 1110 (1 mg/kg, i. p.) 4) SUZAM 1110 (250 pag/kg, i.p-) SUA.M 1-117 (1mg/lkg, i p-) 6) SU7VM 1117 (250 pg/kg, i-p.) 7) SEJAM 1117 (100 PS/kg, i-p.) physiological saline scopolamine (3 mg/kq, i.p.

scopolamine (3 mg/kg,ip) scopolamine (3 mg/kg, i.p-) scopolamine (3 mg/kg, i-p.) scopolamine (3 mg/kg, i. p) scopolamine mg/kg, 9 10 6.2 2.7 230.0 90. 6 272.8 300.0 161.6 232.0 300-0 209.3 110.4 271.2 300.0 125.0 194.4 300.0 3/9 8/10 1/10 0/10 6/10 4/10 0/10 lil~ II I _I Table 3 (cont'd) Sample Drug administered after learning No. of rats tested Initial avoidance time (sec.) Learning Nz. of avoidances during learning Avoidance time (sec.) learning time (sec.) AftEr 24 hrs.

Pharmacological effects After 48 hrs.

No. of amnesic rats/No. of rats tested Percentage amnesia I. 8) SUAM 1117 pg/kg, i.p.) 9) SUAM 1115 (250 }pg/kg, i.p.) SiUTAM 1115 (100 pg/kg, i.p.) 11) SL.AM 1115 )g/kg, i.p.) 12) SUAM 1231 pg/kg, i.p.) 13) SUAM 1333 pg/ku, i.p.) scopolamine (3 mg/kg, i.p,) scop-lamine (3 mg/kg, i.p.) scopolamine (3 mg/kg, i.p.) scopolamine (3 mg/kg, i.p.) scopolamine (3 mg/kg, i.p.) scopolamine (3 mg/kg, i-p.) 1.9 1.5 2.5 2.7 3.1 1.7 100 100 100 100 00 100 300.0 193.2 300.0 300.0 300.0 300.0 289.5 217.1 300.0 300.0 1/10 3,10 0/10 0/10 0/10 0/10 0 0 0 0

L

-27- Example 6 Anti-hypoxia effect Male mice of ddY-strain weighing 25 to 34 g were allotted to groups consisting of 7 to 10 animals each. Each compound to be tested was suspended in physiological saline with 2 to 3 drops of Tween 80 R added thereto, and administered intraperitoneally at the rate of 0.i ml per 10 g of body weight.

Each mouse was put into a 1 k desiccator 30 minutes after the administration, and the pressure in the desiccator was reduced to 180 mmHg by means of a vacuum pump. The period from the start of pressure reduction to the respiratory arrest of the mouse was assumed to be the survival time (minute) of the mouse.

If the mouse survived more than 15 minutes, its survival time was recorded as being 15 minutes. A student's t-test was adopted for the significance analysis.

The results are shown in Table 4.

S 0 a 0 a a a a b a a aaa 00~ c, Table 4 1,,b of urvval/sedSurvival time Chemical Dose (min. ofsMevlusdan r S Drug structure (mrg/kg i. _____Ratio Control Drug Control Drug 1 SLTA-1234 0/10 0/10 2.32 0.11 2.25 0.15 2.88 0.17* 2.45 0.19 1.24 1.09 i P'0. -29- Example 7 Acute Toxicity Test in Mice The compounds of the present invention were checked for their acute toxicity in CDF-1 strain male mice (body weight: 27.2 to 30.1 g) purchased from Awazu Laboratory Animals Co., Ltd.

Test samples were prepared by dissolving the respective compounds in DMSO, and a portion (0.1 ml) of the so conditioned test sample was administered intraperitoneally to each of the mice used. Each of the treated groups consisted of five mice. After 24 hours and 48 hours had passed from the time of administration, respectively, the mice were observed. The average amount of each test compound administered in this Example is shown in Table Table Compound No. Average-Dose (mq/kg) No. 1 SUAM 1231 634.6 No. 2 SUAM 1333 707.7 Each of the groups tested remained sound and showed no sign of intoxication at the times of observation 24 hours and 48 hours after the administration of the doses shown in S Table The present invention includes pharmaceutical compositions which contain compounds according to the present invention which are effective for the improvement of symptoms caused by organic disorders of the brain together with adjuvants which are acceptable from the pharmaceutical Spoint of view.

These active ingredients and pharmaceutical compositions are administered orally in suitable solid forms such Sas capsules, tablets and powders, or liquid forms such as elixir, syrup and suspensions. They may also be administered parenter illy, in the form of injection or suppository.

As an example of suitable excipients for solid drugs which are contained in drug components, a carrier in solid powder form may be cited such as lactose, saccharose, mannitol, sorbitol, cellulose and glycine.

I

As an example of suitable lubricants, silicon dioxide, tali, magnesium stearate and polyethylene glycol may be cited, while starch, gelatin, tragacanth, methyl cellulose, sodium carboxymethyl cellulose and polyvinyl pyrrolidone may be cited as binders. Starch or agar may be used as a disintegrator.

The compounds according to the present invention are orally administered in a daily dose of 10 to 4,000 mg, preferably, 100 to 1,000 mg per adult. Alternatively, they may be administered parenterally in a dose of 1 to 2,000 mg, preferably, 50 to 500 mg. It is understood that the dosage will differ depending upon the symptoms of the disease, the age and weight of a patient, the stage of the symptoms and the form of administration.

Formulation 1 active substance 10 parts lactose 75 parts heavy magnesium oxide 15 parts These components were uniformly mixed and tablets or capsules.

Formulation 2 formed into active substance 45 parts starch 15 parts lactose 40 parts These components were uniformly mixed and formed int powders or granules.

Formulation 3 active substance 1 part surfactant 5 parts physiological saline 94 parts These components were mixed under warming and were sterilized to obtain injections.

As described above, the compounds according to the present inveniton exhibit appreciable anti-prolyl endopeptidase activity and anti-amnesic effects. The acute toxicity test results show that the compounds cause no toxicity even when administered in such high doses as about 500 mg/kg/mouse. Because of this relatively wide margin of 0 S-31safety, as compared with their remarkable anti-prolyl endopeptidase activity, the compounds of the present invention hold promise as pharmaceuticals for preventing and curing amnesia.

o

Claims (8)

1. A dipeptide derivative of the general formula: 1 R (CH 2 -CO -N-CU--CO-NR m H n-R2 R3 (wherein m is an integer of 3 to 8; n is 0 or an integer of 1 to 6; R is hydrogen atom; R is hydrogen atom, a branched alkyl group having 3 to 5 carbon atoms, phenyl group, hydroxyphenyl group or methylthio group, or R 1 and R 2 together represent a single bond between carbon atom and nitrogen atom; and R 3 is CHO; provided that when n is 0, R represents one of said groups except hydrogen atom). S 2. A dipeptide derivative according to Claim 1, which is expressed by the general formula: 0 H (CH2 CO-N-CH- CO--N R3 (CH2 n-R 3 (wherein m is an integer of 3 to 8; n is 0 or an integer of °ooa 1 to 6; R is hydrogen atom, a branched alkyl group having 3 to 5 carbon atoms, phenyl group, hydroxyphenyl group or methylthio group, and R is CHO; provided that when n is 0, R represents one of said groups except hydrogen atom).

3. A dipeptide derivative according to Claim 1, which is expressed by the general formula: (CH 2 CO-N 0 N R 3 (wherein m is an integer of 3 to 8; and R3 is CHO).

4. A dipeptide derivative according to Claim 2, wherein, in said -(CH 2 )n-R 4 group, when n is an integer of 1 to 6 6, R is hydrogen atom, a branched alkyl group having 3 to L I~IIYBIYYalllllCII 33 carbon atoms, phenyl group, hydroxyphenyl group or methylthio group; and when n is 0, R 4 is a branched alkyl group having 3 to 5 carbon atoms. A process for producing a dipeptide derivative of the general formula: R R (CH2)m-CO-N-CH- CO-N 2 (CH 2)n- R3 (wherein m is an integer of 3 to 8; n is 0 or an integer of 1 2 1 to 6; R is hydrogen atom; R is hydrogen atom, a branched alkyl group having 3 to 5 carbon atoms, phenyl o group, hydroxyphenyl group or methylthio group, or R and R 2 together represent a single bond between carbon atom and nitrogen atom; and R is CHO; provided that when n is 0, R represents one of said groups except hydrogen atom), said process comprising the steps of o0 i) reacting a carboxylic acid, acid halide or acid anhydride having the general formula: (CH) COA (wherein m has the same meaning as defined above and A is o hydroxyl group, a halogen atom or a group: -O--C-(CH 2 (wherein m has the same meaning as defined above)) with a peptide ester derivative of the general formula: iA -34- 1 CH-CO-N (CHH2) n-R COOB (wherein n, R 1 and R 2 are as defined above; and B is a lower alkyl group), in the presence of a base to obtain an acyl peptide ester of the general formula: R 1 1 N-CH-CO-N (CH 2m I 2_R 2 (CH2)n-R COOB (wherein m, n, R, R 2 and B, respectively, have the same meanings as defined above); and (ii) reducing the compound so produced to give a compound in which -COOB is reduced to -CH OH, and (iii) oxidizing the compound so produced to give a compound of the formula in which R 3 represents -CHO.

6. A process according to Claim 5, wherein said m dipeptide derivative has the general formula: 0 0 ~O H (CH2) m-CO-N -C-CO--N 2 (CI R4 (cH 2 n R3 (wherein m is an integer of 3 to 8; n is 0 or an integer of 1 to 6; R 4 is hydrogen atom, a branched alkyl group having 3 to 5 carbon atoms, phenyl group, hydroxyphenyl group or 3 methylthio group; and R is CHO; provided that when n is 0, R 4 represents one of said groups except hydrogen atom).

7. A process according to Claim 5, wherein said dipeptide derivative has the general formula: (CH 2 m-CO--N a 02 J o oa (wherein m is an integer of 3 to 8; and R 3 is CHO).

8. A process according to any of Claims 5 to 7, wherein said base is trialkylamine, aqueous alkali metal hydroxide, or alkali metal carbonate.

9. A prolyl endopeptide inhibitor containing a dipeptide derivative of the following formula as an active ingredient: R 1 (CH2) m- CO- N -CH-CO-N m (CH 2 n-R 2 p 3 (wherein m is an integer of 3 to 8; n is 0 or an integer of 1 2 1 to 6; R is hydrogen atom; R is hydrogen atom, a branched alkyl group having 3 to 5 carbon atoms, phenyl group, hydroxyphenyl group or methylthio group, or R and R 2 together represent a single bond between carbon atom and nitrogen atom; and R 3 is CHO; provided that when n is 0, R represents one of said groups except hydrogen atom), together with an acceptable carrier. An anti-amnesic agent containing as the active ingredient a dipeptide derivative of the general o°u formula: Sm .o -R oo (CH2) m- CO--N--CH--CO-N o° (CH 2) n-R 3 R 2 together represent a single bond between carbon atom R3 and nitrogen atom; and R 3 is CHO; provided that when n is 2 0, R represents one of said groups except hydrogen atom), S' together with a pharmaceutically acceptable carrier. 0o 311. A dipeptide derivative according to Claim 1 substantially as hereinbefore described with reference to any one of the Examples. man M II_ i -36-

12. A process according to Claim 5 substantially as hereinbefore described with reference to any one of the Examples. DATED: 19 March, 1990 PHILLIPS ORMONDE FITZPATRICK Attorneys for: SUNTORY LIMITED 0729v 0729v oaP u o c u o o ir r? D o D a or o r ns 6 c. D i) i; O O ii