DK155732B - ANALOGY PROCEDURE FOR THE PREPARATION OF AMINO ACID DERIVATIVES - Google Patents

Description

DK 155732B

The present invention relates to an analogous process for the preparation of novel amino acid derivatives of the general formula R1-NH-CH-CO-A1

5 I

(CHo) I

I 2 η 1

CO-N-CH- (CHO) -B

I '2 2 t

R R

11, wherein A, B, R, R, R, n and t have the meanings specified in the preamble of claim 1, or physiologically acceptable salts or optically active antipodes thereof.

These compounds have valuable biological or pharmacological effects.

Among the compounds of the invention, due to its biological action, it is particularly noted that γ-L-glutamyltaurine having the formula H₂N-CH-COOH

CH2 XXIV

C0-NH-CH2-CH2-SO20H

and which has a broad therapeutic and preventive spectrum of action against morbid changes that can be traced back to damage by "AGAS" (the aerobiospheric genetic adaptation system).

To illustrate the concept of "AGAS", the following are the main tissues and organs that form this system.

(a) All biological interfaces that are in contact with the external air such as the biosphere (skin and skin formation, corneal and conjunctival eyes, oral and pharyngeal cavities, respiratory tract and lung); b) skeleton and joints (tubular and spongy bones, ball joints, synovial membranes, skeletal muscle); C) the organs involved in the regulation of the ionic household (transepithelial transport system: intestinal tracts and renal duct); 2

DK 155732B

d) the tooth kit needed for comminuting the food (rooted in tandaveolans); (e) hearing, smell and voice organs.

Thus, the compounds of this invention exert a favorable biological or therapeutic effect on the organs or tissues of the AGAS system enumerated herein.

In addition, the compounds of the present invention act upon the following functions associated with the AGAS system: radiation protection, favoring wound healing, common activating effect on mesenchyme, protection against the ever-growing infection and contamination risk of skin and mucosa ( the lysozyme production of the moist mucosa, activation of the gut epithelium in the respiratory tract), increased protection against the infections caused by viruses and fungi.

The compounds of the present invention are effective against the ever-increasing stress effects associated with mainland life (e.g., meteorological influence, large differences in day and night temperature, increased risk of injury) as they stabilize. adaptation syndrome and at the same time mitigates the peripheral tissue damage of glucocorticoids (such as damage to the connective tissue, injury to bone-trix constituents). Development of immuno-homeostasis (increasing body recognition of which cells are suitable and which are not).

The compounds prepared by the process of the invention exert their effect, both immediately and partly through the regulation of vitamin A metabolism, in the production of vitamin A metabolites of a stronger polar nature. This effect is comparable to the effect of the parathormone on the 25-hydroxycholecalciferol-1oclhydroxylase enzyme in the renal tract. The directions of action of the compounds are as follows:

DK 155732B

Δ) Vitamin A grade effects: a) Pharmacological and biochemical effects:

Increasing effect on chondroitin sulfate synthesis; beneficial effect on the wound healing or on the experimentally impaired wound healing in rats and dogs by cortisone administration; potentiating effect on the effect of vitamin A in rats and chickens in experimentally induced hypo- or hypervitaminoses; attenuating effect on the ulceration-related stress effects in rats; beneficial effect on the degranulation of mastocytes; increasing effect on the production of lysozyme; effect on household traceability (silicon, zinc, copper, manganese, fluorine); promoting effect on epithelial formation; promoting effect on the alkaline phosphatase activity; effect on the granuloma bag formation (Granulomsackbildung) induced by local action of vitamin A; the extremely flat course of the dose-effect curve or the change of effect sign at large doses; activating effect on the Golgi apparatus; beneficial effect on the formation of mucus or goblet cells; increasing effect on the concentration of vitamin A.

B) Clinical-therapeutic indications: keratoconjunctivis sicca; Sjogren's syndrome; rhino-laryngo-pharingitis sicca; ozæna; chronic bronchitis; sinobronchitis; mucoviscidosis; constitutional lung disease in young children; periodontal disease; skin and mucosal susceptibility to viruses and fungi; cortisone antagonistic action; beneficial effect on healing of operative and mucosal wounds; erosio colli; pruritus-like disorders; reducing the sense of smell and taste.

30 B) Vitamin A-grade effects (a) Pharmacological and biochemical effects: effects on blood glucose levels with a transient decrease; increasing effect on phosphaturia, lowering effect on phosphate level t 35 in serum; radiation protective effect; diminishing effect on 4

DK 155732B

the time needed to reach the goal of maze trials in inactivated animals; diminishing effect on experimentally induced fluorine and cadmium toxicosis; increasing effect on the cyclic adenosine monophosphate depletion of the kidneys; attenuating effect 5 on the symptoms of experimentally induced lathyrism; decrease in histamine sensitivity; enhancing effect on liver enzyme tyrosine aminotransferase activity.

b) Therapeutic indications: weak radiation damage; vitiligo; muscle hypotonia; psychoenergetic effect; beneficial effect on involutional and gerontological conditions as well as on the mnestic functions; keloid inclinations; spondylosis ankylo-poetics; diseases of the movement organs due to wear and tear; scle-rotic fundus; amyloidosis; morphea; fibrocytic mastopathy.

In the veterinary medicine, the compounds prepared according to the invention have similar fields of application as in human medicine, ie. eg skin damage (peeling), wound healing and bone fractures.

The process according to the invention is characterized by the characterizing part of claim 1. - ·

In the process of the invention, the compounds of general formula I are prepared by forming an acid amide bond. Hereby, at the amino group or at the amino and α-carboxyl group, one or more protecting groups optionally substituted derivative of an α-aminodicarboxylic acid over its ω-carboxylic group is linked with, for example, 2-aminoethanesulfonic acid, 3-aminopropanesulfonic acid or 2-phosphonethanolamine. Various protecting groups can be used in the formation of the acid amide bond. The mode of coupling is the active ester method (monograph on structure and selective removal of protecting groups and coupling methods: E. Schroder, K. Liibke: The peptides, Volume 1: Methods of peptide synthesis, Academic Press, 1965).

The process according to the invention is illustrated in the following by means of some examples.

35 5

DK 155732B

Example 1 5.42 g (1 mmol) of carbobenzyloxy-L-glutamic acid (a-benzyl) -γ-p-nitrophenyl ester (Chem. Der. 96, 204 »1963) are dissolved in 50 ml of pyridine. The solution is cooled to 0 ° C and within half an hour a solution of 1.25 g (10 mmol) of taurine in 20 ml of water is added dropwise. Then 3 »08 ml (22 mmol) of triethylamine is added to the mixture and cooling and stirring are interrupted. The mixture is left at room temperature for 72 hours and then evaporated under vacuum. The residue is dissolved in 50 ml of water and IN hydrochloric acid is added to the yellow solution until it is decolorized. To remove the p-nitrophenol, shake the solution ten times with 50 ml ether each time. The aqueous phase is evaporated under vacuum. 6.9 g of carbene benzyloxy-γ- (α-benzyl) -L-glut amyltaurine-triethylammonium salt are obtained.

Relative mobility to cysteic acid by paper electrophoresis at pH 6.5 is 0.5.

Example 2 1.082 g (2.2 mmol) of carbobenzyloxy-L-glutamic acid (α-benzyl) -γ-p-nitrophenyl ester are dissolved in 6 ml of a mixture of pyridine and water in a 2: 1 ratio. To the solution is first added 278 mg (2.25 mmol) of homotaurin and then 0.59 ml (4.2 mmol) of triethylamine. The yellow solution is left at room temperature for 72 hours and then evaporated under vacuum. The oily residue is dissolved in water, neutralized with hydrochloric acid and then extracted to remove p-nitrophenol in a continuous ether extractor for eight hours. The aqueous phase 30 is evaporated under vacuum. 1.68 g of carbobenzyloxy-γ- (α-benzyl) -L-glutamyl homotaurine are obtained. Relative mobility to cysteic acid by paper electrophoresis at pH 6.5 is 0.49.

35 6

DK 155732B

Example 3 1.083 g (2.2 mmol) of carbobenzyloxy-L-glutamic acid (α-benzyl) γ-p-nitrophenyl ester is reacted in the manner described in Example 2 with 278 mg (2 mmol) of N-methyltaurine. 1.59 g of carbo-benzyloxy-γ- (α-benzyl) -L-glutamyl-N-methyltaurine are obtained. Its relative mobility to cysteic acid by paper electrophoresis at pH 6.5 is 0.49.

Example 4 2.87 g (6.6 mmol) of carbobenzyloxy-L-glutamic acid (α-benzyl) -yp-nitrophenyl ester are dissolved in 20 ml of pyridine and a solution of 1.25 g (6 mmol) of L is added. -cysteic acid monohydrate in a mixture of 17 ml of water and 17 ml of pyridine. After the addition of 2.6 ml of (18.6 mmol) of triethylamine, the reaction mixture is left at room temperature for 72 hours. The solution is then evaporated under vacuum at 30 ° C. The residue is dissolved in 20 ml of water, acidified with concentrated hydrochloric acid and then shaken 15 times with 10 ml of ether each time. The aqueous phase is evaporated under vacuum at 35 ° C. Carbobenzyloxy-γ- (α-benzyl) -L-glutamyl-L-cysteic acid is obtained. The relative mobility of the compound in relation to cysteic acid by paper electrophoresis at pH 2.0 is 1.03.

2b Example 5 1.083 g (2.2 mmol) of carbobenzyloxy-L-glutamic acid (a-benzyl) -γ-p-nitrophenyl ester are dissolved in 6 ml of a prepared mixture of pyridine and water in a 2: 1 ratio. To the solution is first added 30 282 mg (2 mmol) of colamine phosphate (U.S. Patent No. 2,730,542) and then 0.87 ml (6.2 mmol) of triethylamine. The reaction mixture is left at room temperature for 72 hours and then evaporated under vacuum. Further work-up is carried out in the manner described in connection with carbobenzyloxy-γ- (α-benzyl) -L-glutamylhomotaurine (Example 2). 1.25 g of carbobenzyloxy-γ- (α-benzyl) -L-glutamyl-cholamine phosphate are obtained. The relative mobility of the product to cysteic acid by paper electrophoresis at pH 2.0 is 0.9,

DK 155732B

7

Example 6 526 mg (1.1 mmol) of carbobenzyloxy-L-aspartic acid (oc-benzyl) - 3-p-nitrophenyl ester (Chem. Ber. 97, 1789, 1964) is dissolved in 5 ml of pyridine. The solution is cooled to 0 ° C, then a solution of 125 mg (1 mmol) of taurine in 2 ml of water is added in small portions followed by 0.28 ml (2 mmol) of triethylamine. The reaction mixture is left at room temperature for 48 hours and then evaporated under vacuum. The residue is dissolved in 5 ml of water and to the initially yellow solution is added 1N 10 hydrochloric acid until the solution is decolorized. To remove p-nitrophenol, the solution is shaken ten times with 5 ml of ether each time. The aqueous phase is evaporated under vacuum. 478 mg of carbobenzyloxy-α- (α-benzyl) -L-aspartyl taurine are obtained. The relative mobility of this compound relative to cysteic acid 15 by paper electrophoresis at pH 6.5 is 0.51.

Example 7 526 mg (1.1 mmol) of carbobenzyloxy-L-aspartic acid (α-2Q benzyl) -β-p-nitrophenyl ester and 139 mg (1 mmol) of homotaurine are reacted in the manner described in Example 6. Carbobenzyloxy-3- (a-benzyl) -If-aspartyl homotaurine is obtained. Its relative mobility to cysteic acid by paper electrophoresis at pH 6.5 is 0.5.

25

Example 8

Carbobenzyloxy-L-aspartic acid (α-benzyl) -β-p-nitrophenyl ester and colamine phosphate are reacted with one another in the manner described in Example 5. Carbobenzyloxy- [3- (cx-benzyl) -L-aspartylcolamine phosphate is obtained. Its relative mobility to cysteic acid by paper electrophoresis at pH 2.0 is 0.9.

35 8

DK 155732B

The biological effect of the compounds of the invention according to the invention will be elucidated in two biological test reports, A and B. In these test reports, the biological effect of four different types of 5 compounds of general formula I. The compounds tested have all unprotected N -terminal and C-terminal groups, 1 1 ie. R is hydrogen and A is hydroxy. Compounds of general formula I wherein R is different from hydrogen and / or where 1A is different from hydroxy have the same biological effect 10 as the corresponding unprotected amino acid derivatives, one having to expect the protecting groups to be cleaved in the organism.

Biological Test Report A.

15

Examination of γ-L-glutamyltaurine as in this report is called litoralon.

In the following Tables 1-6, the experimental results are shown as mean values + standard deviation with the number of determinations in parentheses. Determination of whether there is a significant difference between control trials and trials in treated animals was done with Student's t-test.

Table 1 25 Effect of Litoraion on vitamin A concentration in serum.

Group of litoralone vitamin A concentration in serum _ug / day_mg% _ I. Control - 28.3 + 0.7 (20) 30 II. 0.1 41.9 + 1.0X (20) III. 0.3 32.9 + 1.1XX (20) IV. 1.0 27.4 + 0.6 (20) x P <0.001 25 xx P <0.01

DK 155732B

9 20 Sprague Dawley rats (10 males and 10 females) weighing 180-200 g were orally treated with the daily doses of litoralone in Table 1 in the form of an aqueous solution for a period of 8 days. On the 9th test day, the animals were sacrificed by decapitation and the blood was collected. The vitamin A concentration in the serum was determined by Neeld and Pearson's method.

Table 2

Effect of Litoralon and Vitamin A on Granule Formation Caused by Implanted Cotton Balls

Group Dose Dry weight off. x. ,, Ί granuloma

Vitamin A litoralon local local oral y 15 mg ug yg / day_ I. Control - - - 52 + 1.0 (24) II. Control +

Solvent - - - 54 + 3.1 (8) III. 2 - - 64 + 2.5 (8) 20 “IV. 2 0.1 - 65 + 2.7 (8) V. - - 0.1 73 + 2.9 (8) VI. 2 - 0.1 90 + 4.1 (8) x Hoffmann La Roche 25

The difference is significant: between groups II and III at P <0.05, between II and V at P <0.001, and between V and VI at P <0.01. The granule formation was determined according to Lee et al with male Sprague-Dawley rats weighing 110-120 g. The tampons were removed from the dorsolateral subcutaneous implants after 10 days and weighed after drying to constant weight at 65 ° C.

35

DK 155732B

10

cm cm cm ω in I

O Η Η Ό Ό (U OJ

- - - Hi h 4J <Λ

g O C W

cm r- ~ oo Λ> i U ft

con • tf'd '<r ^ w fi W ^ & U

ft o o oo o to 0) Cl) 0 O - - »- -» + i Λ fcr> -P Cn H o oo oo Η 3.Π3-ΡΟ + 1 +1 +1 «. ° d m <u 1 1 iw ft ro h d rfj T3 s 3 dl 01

3 Μ o ω Λ U

0) d © oo © W + H d

oo co r · »o r ~ o CO 4J d H

λ; Η O «S 'CM HH φ · © 2 · Η Ή d

thaw "". - »κ» tr> ft O

o CM Ο HO CM O · & (D <c S-l CO 4-1 fccj + | +1 +1 tfl H CQ Ό 0) C (0 c ω

03 0 M &> 0 -η H

ft O g X> X

O co tn

CD H CT * 4-10 - T3 C

Pi ^ CM CM ΙΟ Η · Μ * HO 'H ft Η Ή 00 Ο CD Ο 00 Ο Ο Η 0 Ο1 (¾¾ - - «. -»% «. Η tT» 43 dooooooo ti O d 3 ® + 1 +1 +1 <5 o © v Ό »Xi ©

^ IH CT

ft tn <(0 <h H

0 W d Φ

. r ^ cMco'04-iD <DLP

CD CM H CD OH <DO C 4-1 H

e (0 cn o cn o m o> o · H tn 03 U .....- ε O CD ft • Η HOHOHO φ O Ό>> i 3 P - C <D η

S__Id (D ε w Ό -H

4J A4 U © S <> d Λ crf Ή ί> ι »C ^ Η M4 Q U ·! 4 CO CM H CD d ft

CM Γ— CM CD CM in CM d · Λ tj! CD

nrfl ft coo cm o ho tT tn £ d 'd

+) cj * * ~ «· -« · Η σι Ό. K

r-IG OOOOOO BOH * H

CD 03 +1 +1 +1 in 03 ο d CD

n g Η H co CT

B H CD I - 4-1 H

ft H O 4J h d -Q. ·

m H'H'dCDCHH

Q. Η H J4 dP Ο · Η d * 1 ^ * H d 00 0 © row CM H CO OH 4-1 ε CT H d 4-1

03 <ro r- o cto tT ίτ 3 ft <D <D

<h HO ** · »». vv OroCH »d tS

(0 OOOOOO> I ft ε 0 c d) +1 +1 +1 H to H 0 4-1 d C η ε · Η Ό «CD d

So cd m cd ε ε o, CD tT 4-1 Λ 4-1 + j c o ft © -H d 0) TJ + »

-H H CM * = r ft H d CD> (D

h 2 cm in co in h 3 0 k -Η> Ω

Kc * p, j ^^^ v η M * d 'H Cn -H 2 co o tn o co o tn φ ft d d co ·

> +1 +1 +1 ^ ε 0 © d <u \ O

• a ε n -p 3 © µ ft ^ µ ε n Η © o © © © d ocfl H 4-1 d ε + * ft "3 · CO CD CM O (D 4-1 Ή - 03 4- *

»• cm» -cm «-Η ε ο ε - ft © <D

tr> d • M '- cm »<^» H d E-> U Λ ^ e U HO HO HO © d IH · - © H +1 +1 +1 β 5 «a ^^ y c___ © 43 ·» - du «y -f- Η · Η 01 H tntn H> 1 2 d © · © • η ε ε © © ft o 4-) λ! λ> Otn 44 Η <3 Η Ο 0) ο Η 44 03 & Ρ3 d Η 3 £

01 HH MdM ftMO

c O ScnOQCTOOXl © O M 2 ft - h I 6 + * * 4.

H 44 ft <| O © · ΗΤ3 44Μ d © d <rø dOHdO © c. Q. Offl · tT H O Ή 44 O ft «-54 © ΛίΑίΟε 4J 3 · HH H CO 03 Ή © • Η Η · Η H id ft H» H tT H 44 j O Η Η H + 03ί> 44θΧθ3 1 1

DK 155732B

tn 0) Λ ^ r *. (O Φ

VD l £> Ο Ό P

^ · §

UP

tn XX m tn id X X <l> Ό σι H rH ri in «tf β Ρ Λ« tf H m H m Η Φ tu

• Honono O-HIS

o ·. - - - - -> p

O O o O O O (Otfl-H

+ i +1 +1 ϋ s

tn rH

G) Q) -rH

C rH ω tn XX p id XX Φ Ό · O) O ΰφ πν mn Ρ Φ Ρ β C \ | O O rH O rH id g φ Φ • rH O Cl O ri O P 01> O »» »·« · * »" * Η p 0 Φ

CMOOOOOO Ρ Φ U P

+ | +1 + | W a -Θ.

φ · Ρ φ

Ρ β tn w id Η Φ tn χ Η C

tn 0 β tn Φ ιΰ XX to φ in Όη'ΗΓΜνοΗΐη Η, β U> ι rHHCOOOrH ο Ό • rH Ο Η Ο ΓΗ Ο - Φ Φ r—. η ****** ο χ3 β id UOOOOOO Φ> Ρ Ο +1 +1 +1 V η · Ρ Ρ rH> tn Λ ft id β -Ρ 13 Φ Φ tn Ό β tn \ φ · Ρ «tf Φ tn> -Ρ

g tn «ud Η U

w ιΰ fltn h ^ 1 H «i Φ a Φ Ρ T3 HH LD o r * O Ρ Λ

g rH O rH O rH O fi M id P

^ j3 ****** * id φ -Ρ φ • HP 'OO oo oo Ai P> rH U +1 +1 +1 -ri P p ·

φ ρ · Η Ρ Ό. P

X) (d <-H -Ρ Ρ Φ a

<d φ -Ρ β Ό D 'O

EH> w 5 0

• Η · Η Ή rH

a W Φ Λ g tn X G -ρ ρ υ Ό (Λ IN οω ΡΦγΗγΗ

• Η> d <! Ό ΙΛΗ hO φ tn Id S

H Ο Ο rH Ο Η Ο Ό.Ρ • η. ^ ^ ~ t n οι ο t n

i-η tn o o o o o o id P

ri +1 + 1 + 1 Ό Ρ P | d

01 (p φ P

•> P

οι η · · Η 4J rH rH tn 13

qj o 13 P

irj β O β

o φ Φ - · Η N

h s H 'V ° ^ t! η -π ro m in vd nr- H o> (d P oo mo oo oo id v Φ Λ

0 (3 rH O O O rH O * * Ρ Η P

(¾. * * * * V «. O O P {I, ρ β

O O O O O O O O

tn + | + | +1 v v Φ τι β d β Φ O Φ • p Hl Al p> H tn d Φ id h v * ·· ·· Ρ Φ ρ ©.

u dtnoioiidPOP

• ri β Old ββΡβΡ-Η

*> OtnHO (didPid-P

h id idX. At X P a: PI β οι h idO P tn -ρ -ρ οι -ρ φ β o P \ O Ρ- Ρ Ρ Φ P · u

O P 0 tn Ρ · ρ · ρ (¾ · ρ tn O

Η ΡΡ ^ -ΡΟβββΑίΡ id Φ β · Ρ PlrH tn tn tn Φ P Qj O PI tn · ρ · Ρ · ρ ci g o a «· ω to in i _ ρ d. η mu • ρρ · Η η X Ρ-Φ Η Η Η XX Φ Ν> 12 DK 1557328

Table 5

The effect of Litoralon on the metamorphosis of Rana dalmatina

Group Body length Tail length 5 _mm_mm_

Control 45.9 +0/2 31.4 + 0.2 (60)

Treated animals 40.2 + 0.2X 26.7 + 0.2X (60) x Significance: P <0.001 in η

The experiment was conducted with larvae of Rana dalmatina with a age of 30 days, a body length of 20-25 mm, and with already visible projecting hind legs. Over a period of 30 days, the test animals were placed in tap water containing 0.5 µg / ml litoralon for two hours each day. The control group was placed in 15 tap water without the addition of litoralone. The tails were measured on the 30th day.

Table 6 2Q Litoralone's effect on blood sugar levels

Group I. Study II. Survey _mg% _mg% _

Control 94 + 3.6 (10) 94 + 4.09 (10)

Treated animals 82.4 + 3.8 (10) 81 + 1.32 (10) 25

Significance at I. study: P <0.05 Significance at II. study: P <0.01

Male CGY white body rats 160-180 g were used for the experiment. The animals were kept on a standard diet. 2Q blood samples were taken on the 5th day of trial after 18 hours of fasting. The blood glucose assays were performed according to the method of E. Hultman.

Litoralon was given orally at daily doses of 1 µg / kg.

35 13

DK 155732B

Biological Test Report B.

Investigation of various compounds of formula I.

1. β-Aspartyl-N-methyltaurine 5 a) Effect on blood sugar level Control 107 mg%

Treated animals 93 mg%

Significance: P <0.05

20-20 rats were used for the samples. Measurements were carried out -10 after 18 hours of fasting. The dose used was 1 µg / kg body weight for 4 days in the form of a solution by oral administration.

b) Increasing effect on serum vitamin A level

Oral dose Conical vitamin A, mg% 15 µg / 200 g body weight 0 (control) 8.5 5 11.0 1 11.5 0.3 12.5 20 0.1 16.1 0.05 14.8 0.01 12.5 0.005 10.5 2 ,. Significance: P <0.01

For the experiments, 20-20 Wistar male rats weighing 200-220 g were used.

Trial period: 6 days.

c) Effect on blood silicon level:

Silicon (mg / g blood) 0 hours 20 days_40 days_

Control group 0.110 + 0.004 0.120 + 0.010 0.154 + 0.015

Treatment group 15 µg / day 0.100 + 0.005 0.315 + 0.014 0.345 + 0.015

Treatment Group II 10 µg / day 0.107 + 0.009 0.370 + 0.119 0.360 + 0.017 xx Significance: P <0.001 14

DK 155732B

Results are significant from day 20 at level P <0.001. The experiments were performed on inbred male rabbits of body weight 2.5-3 kg. The active ingredient was administered orally in the daily doses shown in the table. The determination of silicon was carried out i-5 according to Gaubatz's method (Gaubatz E., Clin. Wschrft. 14, 1753, 1935) in 5 ml blood samples taken from the animals' ears.

Each group consisted of five animals.

d) Effect of β-aspartyl-N-methyl-taurine and vitamin A on the 10 granulogenic effect caused by the implantation of cotton wool:

Group Dose Weight of dry

Vitamin Ax 3-aspartyl-N- ret 9ranuloma

t. IRCT

methyl taurus local local oral 15 mg / day / day ____

Control I. - - - 54 + 1.7

Control II. + solvent - - - 55 + 3.4

Treatment Group III. 2 - - 66 + 12.5 2g Treatment Group IV. 2 0.1 - 67 + 2.6

Treatment group V. - - 0.1 79 + 2.8

Treatment Group VI. 2 - 0.1 96 + 4.4 x Hoffmann la Roche 25 The differences are significant as follows:

Between groups II and III P <0.05, between groups II and V P <0.001 and between groups V and VI P <0.01.

The determination of granuloma was performed on groups of five male Sprague-Dawley body weights 110-120 g by the method of Lee et al (Lee K.H., Fu Ch.Ch., Spencer M.R.

Tong T.G. and Poon R.J., Pharm. Sci., 62, 895, 1973). The dor- solaterally subcutaneously implanted tampons were removed after 10 days and measured after drying at 65 ° C to constant weight.

35 15

DK 155732B

2. β-Aspartyl homotaurine, a) Effect on blood sugar level

Control group 105 mg%

Treated group 94 mg% 5 Significance, number of test animals and test method were as given in section. 1 a) above.

b) Increasing effect on serum vitamin A level

Oral dose Vitamin A concentration

10 pg / 200 g body weight mg% 0 (control) 8.6 5 12.0 1 13.5 0.3 14.0 15 0.1 15.8 0.05 15.0 0.01 12.0 0.005_10 , 0_

Significance, number of test animals and test method were as indicated above under section. Ib).

c) Effect on blood silicon level:

Silicon (mg / g blood) 25 _0 hours _20 days_40 days__

Control group 0.104 + 0.009 0.134 + 0.015 0.157 + 0.020

Treatment group I. 5yg / day 0.094 + 0.007 0.309 + 0.014 0.340 + 0.014

Treatment Group II. 10 pg / day 0.109 + 0.010 0.372 + 0.120 0.363 + 0.018ΛΛ 30 Significance, number of test animals, arrangement and method of determination as indicated above under section. 1 c).

35 16

DK 155732B

(d) Effect of β-aspartyl homotaurin and vitamin A on the granulomatous effect of cotton implantation;

Dose Weight of dried

Vitamin Ax β-aspartyl granuloma 5 local homotaurin g mg local oral _µg yg / day_

Control group I - - - 52 + 1.5

Control group II - - - 53 + 3.2 + solvent 10 Treatment group III. 2 - - 64 + 12.3

Treatment Group IV. 2 0.1 - 65 + 2.4

Treatment group V. - - 0.1 77 + 2.6

Treatment group VI._2_ 0.1_94 + 4.2__ x Hoffmann la Roche 15

Significance, number of test animals, arrangement and method of determination were as indicated above under point (ld).

3. γ-Glutamyl-Colamine Phosphate.

(a) Effect on blood sugar levels 20

Control group 104 mg%

Treatment group 93 mg%

Significance, number of test animals, arrangement and dosage were as indicated above under point 1a).

25 b) Increasing effect on serum vitamin A level

Oral dose of Vitamin A concentration yg / 200 g body weight_mg% _________ 0 (control) 8.8 30 5 12'3 1 13.4 0.3 14.3 0.1 16.0 0.05 15.5 0.01 11.2 35 0.005_9j2_ 17

DK 155732B

Significance, number of test animals, arrangement and method of determination were as indicated above under point 1 (b).

c) Effect on blood silicon level: 5 Silicon (mg / g blood) _0 hours_20 days_40 days_

Control group 0.106 + 0.011 0.136 + 0.017 0.159 + 0.022

Treatment group xx I 5 µg / day 0.096 + 0.009 0.311 + 0.016 0.340 + 0.017 .. Treatment group vv xx 10 II 10 µg / day 0.111 + 0.011 0.374 + 0.121xx 0.365 + 0.019

Significance, number of test animals, arrangement and method of determination were as indicated above under point 1 (c).

(d) Effect of γ-glutamyl-cholamine phosphate and vitamin A on the 1 s granuloma-inducing effect of water implantation;

Dose Weight of dry straight granuloma

Vitamin Ax γ-glutamyl mg local colamine phosphate mg local oral 2Qg / day / day

Control group I. - - - 49 + 1.2

Control group II. + solvent - - - 50 + 2.9

Treatment Group III. 2 - 61 + 12.0

Treatment Group IV. 2 .0.1 - 62 + 2.1. Treatment Group V. - - 0.1 74 + 2.3

Treatment Group VI. 2 - 0.1 91 + 3.9 x Hoffmann la Roche

Significance, number of test animals, arrangement and determination: - method of measurement was as indicated under '1 d) above.

35

Claims (2)

1. Analogous process for preparing amino acid derivatives of the formula 5 R1-NH-CH-CO-A1 (CEL ·) II 2 n -i CO-N-CH- (CH9) -BI 2 * RR where 10. is hydroxy, C, _ ^ alkoxy or phenyl-C ^ ^ alkoxy; B1 is a group of the formula -SO20H or -OPO (OH) 2; R is hydrogen or alkyl of 1-4 carbon atoms; R is hydrogen, alkoxycarbonyl or phenyl-C 1-4 alkoxycarbonyl; 2 is hydrogen or carboxyl, n is 1 or 2 and t is 1 or 2, or physiologically acceptable salts or optically active antipodes thereof, characterized by reacting an optically active or racemic compound of the general formula r ' -nh-ch-coa (CH2) n II Wherein R, A and n are as defined above and 2A is p-nitrophenoxy, pentachlorophenoxy or alkoxycarboxyloxy having 2-4 carbon atoms in the alkoxy moiety, with an optically active or racemic compound with the general Formula NH-CH - (CEL ·). - B1 III R Å2 2 * 35 2 1 wherein R, R, B and t have the meanings given above, and if desired, the compound thus obtained is converted into a physiologically acceptable salt thereof or release it from a salt and / or prepare the obtained compound in optically active form by digestion of a obtained racemic product. Process according to claim 1, characterized in that a compound of the general formula II, preferably a N-carbobenzyloxyaminodicarboxylic acid oc-benzyl-ω-ρ-nitrophenyl ester, is reacted with a compound of the general formula III, preferably with taurine, N -methyltaurine, homo-taurine, colamine phosphate or cysteic acid, in the presence of aqueous pyridine. 15 20 25 30 35