CA1335493C - Process for the preparation of tripeptides - Google Patents
Process for the preparation of tripeptidesInfo
- Publication number
- CA1335493C CA1335493C CA000614545A CA614545A CA1335493C CA 1335493 C CA1335493 C CA 1335493C CA 000614545 A CA000614545 A CA 000614545A CA 614545 A CA614545 A CA 614545A CA 1335493 C CA1335493 C CA 1335493C
- Authority
- CA
- Canada
- Prior art keywords
- denotes
- compound
- trp
- tyr
- ser
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Lifetime
Links
- 238000000034 method Methods 0.000 title claims abstract description 29
- 238000002360 preparation method Methods 0.000 title claims abstract description 7
- 150000001875 compounds Chemical class 0.000 claims abstract description 13
- 238000006243 chemical reaction Methods 0.000 claims abstract description 12
- 238000007327 hydrogenolysis reaction Methods 0.000 claims abstract description 8
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 claims abstract description 5
- 229910052739 hydrogen Inorganic materials 0.000 claims abstract description 5
- 239000001257 hydrogen Substances 0.000 claims abstract description 5
- 125000000217 alkyl group Chemical group 0.000 claims abstract description 3
- 230000002255 enzymatic effect Effects 0.000 claims abstract description 3
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 24
- PAQZWJGSJMLPMG-UHFFFAOYSA-N 2,4,6-tripropyl-1,3,5,2$l^{5},4$l^{5},6$l^{5}-trioxatriphosphinane 2,4,6-trioxide Chemical compound CCCP1(=O)OP(=O)(CCC)OP(=O)(CCC)O1 PAQZWJGSJMLPMG-UHFFFAOYSA-N 0.000 claims description 12
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 11
- 102000004142 Trypsin Human genes 0.000 claims description 6
- 108090000631 Trypsin Proteins 0.000 claims description 6
- 125000001584 benzyloxycarbonyl group Chemical group C(=O)(OCC1=CC=CC=C1)* 0.000 claims description 6
- 239000012588 trypsin Substances 0.000 claims description 6
- 239000000203 mixture Substances 0.000 claims description 5
- UYWQUFXKFGHYNT-UHFFFAOYSA-N phenylmethyl ester of formic acid Natural products O=COCC1=CC=CC=C1 UYWQUFXKFGHYNT-UHFFFAOYSA-N 0.000 claims description 5
- 150000001412 amines Chemical class 0.000 claims description 4
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 claims description 4
- 125000003118 aryl group Chemical group 0.000 claims description 3
- 125000006239 protecting group Chemical group 0.000 claims description 3
- 125000004432 carbon atom Chemical group C* 0.000 claims description 2
- 229960002376 chymotrypsin Drugs 0.000 claims description 2
- 108090000765 processed proteins & peptides Proteins 0.000 abstract description 6
- 239000000543 intermediate Substances 0.000 abstract description 4
- 230000015572 biosynthetic process Effects 0.000 abstract description 3
- 238000003786 synthesis reaction Methods 0.000 abstract description 3
- 230000000975 bioactive effect Effects 0.000 abstract description 2
- 102000004196 processed proteins & peptides Human genes 0.000 abstract description 2
- 230000008030 elimination Effects 0.000 abstract 2
- 238000003379 elimination reaction Methods 0.000 abstract 2
- 150000001413 amino acids Chemical class 0.000 abstract 1
- 238000007792 addition Methods 0.000 description 12
- 239000012071 phase Substances 0.000 description 12
- 239000008346 aqueous phase Substances 0.000 description 10
- 150000002148 esters Chemical class 0.000 description 10
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 9
- 239000000243 solution Substances 0.000 description 9
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 6
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 description 6
- UIIMBOGNXHQVGW-UHFFFAOYSA-M Sodium bicarbonate Chemical compound [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 description 6
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 5
- KDLHZDBZIXYQEI-UHFFFAOYSA-N Palladium Chemical compound [Pd] KDLHZDBZIXYQEI-UHFFFAOYSA-N 0.000 description 5
- HVCNXQOWACZAFN-UHFFFAOYSA-N 4-ethylmorpholine Chemical compound CCN1CCOCC1 HVCNXQOWACZAFN-UHFFFAOYSA-N 0.000 description 4
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
- 239000002244 precipitate Substances 0.000 description 4
- 239000000047 product Substances 0.000 description 4
- 102000004190 Enzymes Human genes 0.000 description 3
- 108090000790 Enzymes Proteins 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- YXFVVABEGXRONW-UHFFFAOYSA-N Toluene Chemical compound CC1=CC=CC=C1 YXFVVABEGXRONW-UHFFFAOYSA-N 0.000 description 3
- 229940088598 enzyme Drugs 0.000 description 3
- 239000011541 reaction mixture Substances 0.000 description 3
- 235000017557 sodium bicarbonate Nutrition 0.000 description 3
- 229910000030 sodium bicarbonate Inorganic materials 0.000 description 3
- AHYFYYVVAXRMKB-KRWDZBQOSA-N (2s)-3-(1h-indol-3-yl)-2-(phenylmethoxycarbonylamino)propanoic acid Chemical compound N([C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)O)C(=O)OCC1=CC=CC=C1 AHYFYYVVAXRMKB-KRWDZBQOSA-N 0.000 description 2
- JTNCEQNHURODLX-UHFFFAOYSA-N 2-phenylethanimidamide Chemical compound NC(=N)CC1=CC=CC=C1 JTNCEQNHURODLX-UHFFFAOYSA-N 0.000 description 2
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 2
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 2
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 2
- KFZMGEQAYNKOFK-UHFFFAOYSA-N Isopropanol Chemical compound CC(C)O KFZMGEQAYNKOFK-UHFFFAOYSA-N 0.000 description 2
- LRHPLDYGYMQRHN-UHFFFAOYSA-N N-Butanol Chemical compound CCCCO LRHPLDYGYMQRHN-UHFFFAOYSA-N 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 230000008878 coupling Effects 0.000 description 2
- 238000010168 coupling process Methods 0.000 description 2
- 238000005859 coupling reaction Methods 0.000 description 2
- 238000004128 high performance liquid chromatography Methods 0.000 description 2
- 229960000443 hydrochloric acid Drugs 0.000 description 2
- 235000011167 hydrochloric acid Nutrition 0.000 description 2
- 229910000343 potassium bisulfate Inorganic materials 0.000 description 2
- 229960001407 sodium bicarbonate Drugs 0.000 description 2
- 239000011780 sodium chloride Substances 0.000 description 2
- 239000002904 solvent Substances 0.000 description 2
- 239000007858 starting material Substances 0.000 description 2
- GNIDSOFZAKMQAO-VIFPVBQESA-N (2s)-3-hydroxy-2-(phenylmethoxycarbonylamino)propanoic acid Chemical compound OC[C@@H](C(O)=O)NC(=O)OCC1=CC=CC=C1 GNIDSOFZAKMQAO-VIFPVBQESA-N 0.000 description 1
- VPNIQGRFZCTBEZ-SPTGULJVSA-N 3-n-[(2s,3r)-4-(cyclopropylamino)-3-hydroxy-1-phenylbutan-2-yl]-5-[methyl(methylsulfonyl)amino]-1-n-[(1r)-1-phenylethyl]benzene-1,3-dicarboxamide Chemical compound C([C@H](NC(=O)C=1C=C(C=C(C=1)C(=O)N[C@H](C)C=1C=CC=CC=1)N(C)S(C)(=O)=O)[C@H](O)CNC1CC1)C1=CC=CC=C1 VPNIQGRFZCTBEZ-SPTGULJVSA-N 0.000 description 1
- DKPFZGUDAPQIHT-UHFFFAOYSA-N Butyl acetate Natural products CCCCOC(C)=O DKPFZGUDAPQIHT-UHFFFAOYSA-N 0.000 description 1
- 102400000932 Gonadoliberin-1 Human genes 0.000 description 1
- 101500026183 Homo sapiens Gonadoliberin-1 Proteins 0.000 description 1
- FXHOOIRPVKKKFG-UHFFFAOYSA-N N,N-Dimethylacetamide Chemical compound CN(C)C(C)=O FXHOOIRPVKKKFG-UHFFFAOYSA-N 0.000 description 1
- SECXISVLQFMRJM-UHFFFAOYSA-N N-Methylpyrrolidone Chemical compound CN1CCCC1=O SECXISVLQFMRJM-UHFFFAOYSA-N 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 229960000583 acetic acid Drugs 0.000 description 1
- 239000006227 byproduct Substances 0.000 description 1
- 239000003054 catalyst Substances 0.000 description 1
- 239000013078 crystal Substances 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 125000001495 ethyl group Chemical group [H]C([H])([H])C([H])([H])* 0.000 description 1
- 239000012362 glacial acetic acid Substances 0.000 description 1
- XLXSAKCOAKORKW-AQJXLSMYSA-N gonadorelin Chemical compound C([C@@H](C(=O)NCC(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N1[C@@H](CCC1)C(=O)NCC(N)=O)NC(=O)[C@H](CO)NC(=O)[C@H](CC=1C2=CC=CC=C2NC=1)NC(=O)[C@H](CC=1N=CNC=1)NC(=O)[C@H]1NC(=O)CC1)C1=CC=C(O)C=C1 XLXSAKCOAKORKW-AQJXLSMYSA-N 0.000 description 1
- 229960001442 gonadorelin Drugs 0.000 description 1
- GNOIPBMMFNIUFM-UHFFFAOYSA-N hexamethylphosphoric triamide Chemical compound CN(C)P(=O)(N(C)C)N(C)C GNOIPBMMFNIUFM-UHFFFAOYSA-N 0.000 description 1
- FUZZWVXGSFPDMH-UHFFFAOYSA-N hexanoic acid Chemical compound CCCCCC(O)=O FUZZWVXGSFPDMH-UHFFFAOYSA-N 0.000 description 1
- 239000000601 hypothalamic hormone Substances 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 229910000402 monopotassium phosphate Inorganic materials 0.000 description 1
- 235000019796 monopotassium phosphate Nutrition 0.000 description 1
- 238000005580 one pot reaction Methods 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 description 1
- 239000002798 polar solvent Substances 0.000 description 1
- 239000012429 reaction media Substances 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
- C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
- C07K5/08—Tripeptides
- C07K5/0821—Tripeptides with the first amino acid being heterocyclic, e.g. His, Pro, Trp
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/06—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length using protecting groups or activating agents
- C07K1/08—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length using protecting groups or activating agents using activating agents
- C07K1/082—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length using protecting groups or activating agents using activating agents containing phosphorus
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P20/00—Technologies relating to chemical industry
- Y02P20/50—Improvements relating to the production of bulk chemicals
- Y02P20/55—Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Genetics & Genomics (AREA)
- Biochemistry (AREA)
- Biophysics (AREA)
- General Health & Medical Sciences (AREA)
- Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Molecular Biology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Life Sciences & Earth Sciences (AREA)
- Analytical Chemistry (AREA)
- Peptides Or Proteins (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
A process for the preparation of tripeptides A process for the preparation of tripeptides of the general formula I
U - A - B - C - OH I
in which U denotes hydrogen or a urethane protective group and A, B and C denote amino acids, by reaction of a compound of the general formula II
U' - B - OH II
in which U' is a urethane protective group which can be eliminated by hydrogenolysis, with a compound of the general formula III
H - C - OR III
in which R denotes alkyl, by the PPA method, elimination of U', reaction of the resulting compound with a compound of the general formula IV
H - B - C - OR IV
by the PPA method and subsequent enzymatic elimination of R. The tripeptides are intermediates used in the synthesis of bioactive peptides.
U - A - B - C - OH I
in which U denotes hydrogen or a urethane protective group and A, B and C denote amino acids, by reaction of a compound of the general formula II
U' - B - OH II
in which U' is a urethane protective group which can be eliminated by hydrogenolysis, with a compound of the general formula III
H - C - OR III
in which R denotes alkyl, by the PPA method, elimination of U', reaction of the resulting compound with a compound of the general formula IV
H - B - C - OR IV
by the PPA method and subsequent enzymatic elimination of R. The tripeptides are intermediates used in the synthesis of bioactive peptides.
Description
De~cription 13 3 S ~ g 3 A process for the preparation of tripeptides Tripeptides are important intermediates in the synthesis of bioactive peptides such as, for example, the hypo-thalamus hormone gonadorelin and its analogs. For this purpose the tripeptides must be available in the most ~traightforward manner possible, on the one hand in good yields and, on the other hand, in high purity. The processes hitherto disclosed for the preparation of tripeptides do not meet these requirements in an optimal manner and are associated with disadvantages, some of which are serious. Thus, for example, even the products prepared by the process described in EP-A 156,280 are contaminated with byproducts which become disadvanta-geously evident in the subsequent synthetic steps. Thus the object of the present invention is to provide a process for the preparation of tripeptides which does not have the said disadvantages and provides, in a straight-forward manner, products of high purity in good yields.
Accordingly, the invention relates to a process for thepreparation of tripeptides of the general formula I
U - A - B - C - OH
in which U denotes hydrogen or a urethane protective group A denotes a natural ~-amino acid or derivatives thereof B denotes a natural ~-amino acid or derivatives thereof and C denotes an aromatic ~-amino acid, which comprises reacting a compound of the general formula II
U' - B - OH
~ II
Accordingly, the invention relates to a process for thepreparation of tripeptides of the general formula I
U - A - B - C - OH
in which U denotes hydrogen or a urethane protective group A denotes a natural ~-amino acid or derivatives thereof B denotes a natural ~-amino acid or derivatives thereof and C denotes an aromatic ~-amino acid, which comprises reacting a compound of the general formula II
U' - B - OH
~ II
- 2 - 1 33~93 in which U' is a urethane protective group which can be eliminated by hydrogenolysis, and B has the above-mentioned meaning, with a compound of the general formula III
H - C - OR
III
in which R represents alkyl having 1 to 4 carbon atoms, and C has the abovementioned meaning, in the presence of propylphosphonic anhydride, eliminating the protective group U' by hydrogenolysis, reacting the resulting compound of the general formula IV
H - B - C - OR IV
with a compound of the general formula V
U - A - OH V
in the presence of propylphosphonic anhydride, and finally eliminating R enzymatically.
The urethane protective groups representing U are prefer-ably the urethane protective groups customary in peptide chemistry, as are described, for example, in Kontakte Merck 3/79, page 14.
The benzyloxycarbonyl and the tert.-butyloxycarbonyl groups are particularly preferred.
A urethane protective group U' which can be eliminated by hydrogenolysis is preferably the benzyloxycarbonyl group.
Natural ~-amino acids or their derivatives representing A and/or B are preferably Gly, Ala, Ser, Thr, Val, Leu, Ile, Glu, Gln, p-Glu, Tyr, Phe, Trp and His. Ser, Thr, Trp and Phe are particularly preferred.
An aromatic Q-amino acid representing C is preferably Tyr or Phe.
_ 3 _ 133549~
R in the general formula IV preferably denotes methyl.
A process in which U and U' denote benzyloxycarbonyl, A
denotes Trp, B denotes Ser, C denotes Tyr and R denotes methyl is very particularly preferred.
S The formation of a peptide linkage in the presence of propylphosphonic anhydride is known as the PPA method (Angew. Chem. Int. Ed. 19, 133 (1980)). This reaction i8 preferably carried out in polar solvents ~uch as, for example, dimethylacetamide, dimethylformamide, dimethyl sulfoxide, phosphoric tris(dimethylamide), N-methyl-pyrrolidone or water. However, chloroform, methylene chloride or ethyl acetste are also employed.
It is also po~sible in an advantageous manner to use mixtures of the said solvents with water. An ethyl acetate/water mixture is particularly preferred. The ~ynthesis can be carried out between -10C and room temperature. It is preferable to start at about 0C and subse~uently to raise to room temperature.
The elim~nation of the U' protective group by hydrogeno-lysis is advantageou~ly carried out in a known manner with hydrogen on a Pd/C catalyst.
The enzymatic e~terolysis in the last reaction step is prefersbly carried out with trypsin and/or ~-chymotrypsin (Hoppe-Seylers Zeit~chrift f. physiol. Chemie, 336, 248 (1964)). Trypsin is particularly preferred. Where ap-propriate, enzymes which are immobilized by known methods on ~ support are also used, such as de~cribed, for example, in Canadian Patent 1,265,083. In this case, the enzymes are advantageously employed in amounts of 0.01 to 20%
by weight relative to the amount of substrate. An amount of 2~ by weight of enzyme is particularly preferred.
Examples of solvents which can be employed sre water, dimethylformamide, methanol, ethanol, isopropanol, butanol, ethyl acetate, butyl acetate, toluene or J~
methylene chloride.
An ethyl acetate/water mixture is preferred. The tempera-tures are advantageously between O and 60C. A temperature ranqe from 20 to 35C is preferred. The pH of the reaction medium is preferably in the range between 4 and 10, particularly preferably between 4 and 8.
The process according to the invention can be carried out in ~uch a way that each intermediate is isolated. How-ever, it i~ preferably carried out in a one-pot process, that i8 to say without isolation of the intermediates.
The starting compounds of the general formulae II, III
and V are known and can be obtained by the customary methods.
The process according to the invention surpri~ingly provides products of high chemical and optical purity, which can be employed- without difficulty in further syntheses. The yields are likewise excellent and are between 40 and 50 % based on the amount of the compound of the general formula III employed.
It has to be regarded as particularly surprising that the process according to the invention is distinctly su-perior, in terms both of purity and of yield, to the proces~ of Canadian Patent 1,278,650, which ha~ only three stages.
B~ample Z-Trp-Ser-Tyr-OH
a) 350 ml of water are placed in a 2 1 stirred apparatus, and 47.8 g (0.200 mol) of Z-Ser-OH, 46.4 g (O.200 mol) of H-Tyr-O~ex~Cl and 150 g of sodium chloride are introduced. Also added are 700 ml of ethyl acetate and, after everything has dissolved, the pH of the mixture is ad~usted to 5.0 by addition of about 25 ml of N-ethylmorpholine. Durinq the addition of about 220 ml (0.42 mol) of PPA solution .,; .
~ 5 ~ 1335~93 (w(PPA) in % = 50) in about 30 minutes at a maximum of 30C (cool somewhat at the end), about 110 ml (0.86 mol) of N-ethylmorpholine are added via a pH-stat pump at pH 5Ø The PPA addition is terminated when a precipitate forms in the reaction mixture.
The precipitate is redissolved by subsequent add-ition of 350 ml of water. The aqueous phase i8 separated off in a separating funnel and then the ester phase is washed with 700 ml of potassium bisulfate solution (w(RHSO4) in % = 10) and 700 ml of sodium bicarbonate solution (w(NaHCO3) in % = 5). The aqueous phase from the reaction and the wash phases are discarded.
b) About 700 ml of ester phase from the 1st coupling, 200 ml of water and 3.3 g of palladium on carbon w(Pd) in % = 2.5 are placed in a 2 1 stirred ap-paratus and a stream of hydrogen is passed in at 25-30C. During the reaction the pH is maintA i n~ at 4.0 with a pH-stat pump and addition of about 160 ml (0.16 mol) of hydrochloric acid c(HCl) = 1 mol/l.
After the reaction is complete, when no more hydro-chloric acid is consumed, (about 30 minutes) the reaction mixture is filtered through a suction funnel, and the aqueous phase is separated from the ester phase in a separating funnel. The ester phase is discarded.
c) About 430 ml of aqueous phase from the hydrogeno-lysis and 700 ml of ethyl acetate are placed in a 2 1 stirred apparatus and 50.7 g (0.15 mol) of Z-Trp-OH and 125 g of sodium chloride are sdded.
After everything has dissolved, the pH i8 ad~usted to 5.0 with about 19 ml of N-ethylmorpholine. During the addition of about 220 ml (0.42 mol) of PPA
solution (w(PPA) in % = 50) in about 30 minutes at a maximum of 30C (cool somewhat at the end), about 110 ml (0.86 mol) of N-ethylmorpholine are added via a pH-stat pump at pH 5Ø The PPA addition is terminated when a precipitate forms in the reaction mixture. The precipitate is redissolved by subsequent addition of 350 ml of water. The aqueous phase is separated off in a separating funnel, and then the ester phase is washed with 700 ml of potassium bisulfate solution (w(RHSO4) in % = 10) and several times with 700 ml portions of sodium bicar-bonate solution (w(N-~CO~) in ~ = 5) until Z-Trp-OH
has been completely removed (according to TLC
analysis). The aqueous phase from the reaction and the wash phases are discarded.
d) About 700 ml of ester phase from the 2nd coupling and 700 ml of water are placed in a 2 1 stirred apparatus and heated to 35-40C, and 1 g of trypsin is initially added. The reaction starts immediately and, during it, the pH is maintAineA constant at pH 7.0 with about 110 ml (0.11 mol) of sodium hydroxide solution (c(NaOH) = 1 mol/l). The reaction lasts about 7 hours and, during this, the rate is increased now and again by further addition of 0.5 g of trypsin. It is complete when trypsin addition now brings about only a slight increase in the rate of absorption of sodium hydroxide solution, or TLC
analysis shows hardly any starting material remain-ing. The reaction solution is clarified through a suction funnel, and the ester phase is separated from the aqueous phase in a separating funnel. The ester phase is discarded.
The aqueous phase is initially extracted by shaking twice at pH 5.8-6.0, by addition and dissolution of 4.0 g of potassium dihydrogen phosphate, with 700 ml of ethyl acetate each time. The ester phases are discarded. The aqueous phase is then extracted by shAking three times at pH 5.0, ad~usted by addition of about 5 ml of glacial acetic acid, with 700 ml of ethyl acetate each time. The aqueous phase is discarded. The ester phases contain the tripeptide ~ 7 ~ 1335493 which, on evaparation to dryness in vacuo, remains in the form of loosely packed crystals. The product is dried in a vacuum oven at 40C.
Weight: 51.2 g Yield: 42.0 % based on H-Tyr-OM~Y~Cl Purity: 98.2 % (determined with HPLC LiChrosorb Si 60/peptide buffer) Comparison ExEmple Z-Trp-Ser-Tyr-OH was prepared by the process specified in EP-A 156,280.
Yield: 30 %
Purity: 78.8 % (determined with HPLC LiChrosorb Si 60/peptide buffer)
H - C - OR
III
in which R represents alkyl having 1 to 4 carbon atoms, and C has the abovementioned meaning, in the presence of propylphosphonic anhydride, eliminating the protective group U' by hydrogenolysis, reacting the resulting compound of the general formula IV
H - B - C - OR IV
with a compound of the general formula V
U - A - OH V
in the presence of propylphosphonic anhydride, and finally eliminating R enzymatically.
The urethane protective groups representing U are prefer-ably the urethane protective groups customary in peptide chemistry, as are described, for example, in Kontakte Merck 3/79, page 14.
The benzyloxycarbonyl and the tert.-butyloxycarbonyl groups are particularly preferred.
A urethane protective group U' which can be eliminated by hydrogenolysis is preferably the benzyloxycarbonyl group.
Natural ~-amino acids or their derivatives representing A and/or B are preferably Gly, Ala, Ser, Thr, Val, Leu, Ile, Glu, Gln, p-Glu, Tyr, Phe, Trp and His. Ser, Thr, Trp and Phe are particularly preferred.
An aromatic Q-amino acid representing C is preferably Tyr or Phe.
_ 3 _ 133549~
R in the general formula IV preferably denotes methyl.
A process in which U and U' denote benzyloxycarbonyl, A
denotes Trp, B denotes Ser, C denotes Tyr and R denotes methyl is very particularly preferred.
S The formation of a peptide linkage in the presence of propylphosphonic anhydride is known as the PPA method (Angew. Chem. Int. Ed. 19, 133 (1980)). This reaction i8 preferably carried out in polar solvents ~uch as, for example, dimethylacetamide, dimethylformamide, dimethyl sulfoxide, phosphoric tris(dimethylamide), N-methyl-pyrrolidone or water. However, chloroform, methylene chloride or ethyl acetste are also employed.
It is also po~sible in an advantageous manner to use mixtures of the said solvents with water. An ethyl acetate/water mixture is particularly preferred. The ~ynthesis can be carried out between -10C and room temperature. It is preferable to start at about 0C and subse~uently to raise to room temperature.
The elim~nation of the U' protective group by hydrogeno-lysis is advantageou~ly carried out in a known manner with hydrogen on a Pd/C catalyst.
The enzymatic e~terolysis in the last reaction step is prefersbly carried out with trypsin and/or ~-chymotrypsin (Hoppe-Seylers Zeit~chrift f. physiol. Chemie, 336, 248 (1964)). Trypsin is particularly preferred. Where ap-propriate, enzymes which are immobilized by known methods on ~ support are also used, such as de~cribed, for example, in Canadian Patent 1,265,083. In this case, the enzymes are advantageously employed in amounts of 0.01 to 20%
by weight relative to the amount of substrate. An amount of 2~ by weight of enzyme is particularly preferred.
Examples of solvents which can be employed sre water, dimethylformamide, methanol, ethanol, isopropanol, butanol, ethyl acetate, butyl acetate, toluene or J~
methylene chloride.
An ethyl acetate/water mixture is preferred. The tempera-tures are advantageously between O and 60C. A temperature ranqe from 20 to 35C is preferred. The pH of the reaction medium is preferably in the range between 4 and 10, particularly preferably between 4 and 8.
The process according to the invention can be carried out in ~uch a way that each intermediate is isolated. How-ever, it i~ preferably carried out in a one-pot process, that i8 to say without isolation of the intermediates.
The starting compounds of the general formulae II, III
and V are known and can be obtained by the customary methods.
The process according to the invention surpri~ingly provides products of high chemical and optical purity, which can be employed- without difficulty in further syntheses. The yields are likewise excellent and are between 40 and 50 % based on the amount of the compound of the general formula III employed.
It has to be regarded as particularly surprising that the process according to the invention is distinctly su-perior, in terms both of purity and of yield, to the proces~ of Canadian Patent 1,278,650, which ha~ only three stages.
B~ample Z-Trp-Ser-Tyr-OH
a) 350 ml of water are placed in a 2 1 stirred apparatus, and 47.8 g (0.200 mol) of Z-Ser-OH, 46.4 g (O.200 mol) of H-Tyr-O~ex~Cl and 150 g of sodium chloride are introduced. Also added are 700 ml of ethyl acetate and, after everything has dissolved, the pH of the mixture is ad~usted to 5.0 by addition of about 25 ml of N-ethylmorpholine. Durinq the addition of about 220 ml (0.42 mol) of PPA solution .,; .
~ 5 ~ 1335~93 (w(PPA) in % = 50) in about 30 minutes at a maximum of 30C (cool somewhat at the end), about 110 ml (0.86 mol) of N-ethylmorpholine are added via a pH-stat pump at pH 5Ø The PPA addition is terminated when a precipitate forms in the reaction mixture.
The precipitate is redissolved by subsequent add-ition of 350 ml of water. The aqueous phase i8 separated off in a separating funnel and then the ester phase is washed with 700 ml of potassium bisulfate solution (w(RHSO4) in % = 10) and 700 ml of sodium bicarbonate solution (w(NaHCO3) in % = 5). The aqueous phase from the reaction and the wash phases are discarded.
b) About 700 ml of ester phase from the 1st coupling, 200 ml of water and 3.3 g of palladium on carbon w(Pd) in % = 2.5 are placed in a 2 1 stirred ap-paratus and a stream of hydrogen is passed in at 25-30C. During the reaction the pH is maintA i n~ at 4.0 with a pH-stat pump and addition of about 160 ml (0.16 mol) of hydrochloric acid c(HCl) = 1 mol/l.
After the reaction is complete, when no more hydro-chloric acid is consumed, (about 30 minutes) the reaction mixture is filtered through a suction funnel, and the aqueous phase is separated from the ester phase in a separating funnel. The ester phase is discarded.
c) About 430 ml of aqueous phase from the hydrogeno-lysis and 700 ml of ethyl acetate are placed in a 2 1 stirred apparatus and 50.7 g (0.15 mol) of Z-Trp-OH and 125 g of sodium chloride are sdded.
After everything has dissolved, the pH i8 ad~usted to 5.0 with about 19 ml of N-ethylmorpholine. During the addition of about 220 ml (0.42 mol) of PPA
solution (w(PPA) in % = 50) in about 30 minutes at a maximum of 30C (cool somewhat at the end), about 110 ml (0.86 mol) of N-ethylmorpholine are added via a pH-stat pump at pH 5Ø The PPA addition is terminated when a precipitate forms in the reaction mixture. The precipitate is redissolved by subsequent addition of 350 ml of water. The aqueous phase is separated off in a separating funnel, and then the ester phase is washed with 700 ml of potassium bisulfate solution (w(RHSO4) in % = 10) and several times with 700 ml portions of sodium bicar-bonate solution (w(N-~CO~) in ~ = 5) until Z-Trp-OH
has been completely removed (according to TLC
analysis). The aqueous phase from the reaction and the wash phases are discarded.
d) About 700 ml of ester phase from the 2nd coupling and 700 ml of water are placed in a 2 1 stirred apparatus and heated to 35-40C, and 1 g of trypsin is initially added. The reaction starts immediately and, during it, the pH is maintAineA constant at pH 7.0 with about 110 ml (0.11 mol) of sodium hydroxide solution (c(NaOH) = 1 mol/l). The reaction lasts about 7 hours and, during this, the rate is increased now and again by further addition of 0.5 g of trypsin. It is complete when trypsin addition now brings about only a slight increase in the rate of absorption of sodium hydroxide solution, or TLC
analysis shows hardly any starting material remain-ing. The reaction solution is clarified through a suction funnel, and the ester phase is separated from the aqueous phase in a separating funnel. The ester phase is discarded.
The aqueous phase is initially extracted by shaking twice at pH 5.8-6.0, by addition and dissolution of 4.0 g of potassium dihydrogen phosphate, with 700 ml of ethyl acetate each time. The ester phases are discarded. The aqueous phase is then extracted by shAking three times at pH 5.0, ad~usted by addition of about 5 ml of glacial acetic acid, with 700 ml of ethyl acetate each time. The aqueous phase is discarded. The ester phases contain the tripeptide ~ 7 ~ 1335493 which, on evaparation to dryness in vacuo, remains in the form of loosely packed crystals. The product is dried in a vacuum oven at 40C.
Weight: 51.2 g Yield: 42.0 % based on H-Tyr-OM~Y~Cl Purity: 98.2 % (determined with HPLC LiChrosorb Si 60/peptide buffer) Comparison ExEmple Z-Trp-Ser-Tyr-OH was prepared by the process specified in EP-A 156,280.
Yield: 30 %
Purity: 78.8 % (determined with HPLC LiChrosorb Si 60/peptide buffer)
Claims (11)
1. A process for the preparation of tripeptides of the general formula I
U - A - B - C - OH I
in which U denotes hydrogen or a urethane protective group A denotes a natural .alpha.-amino acid or derivatives thereof B denotes a natural .alpha.-amino acid or derivatives thereof and C denotes an aromatic .alpha.-amino acid, which comprises reacting a compound of the formula II
U' - B - OH II
in which U' is a urethane protective group which can be eliminated by hydrogenolysis, and B has the above-mentioned meaning, with a compound of the formula III
H - C - OR III
in which R represents alkyl having 1 to 4 carbon atoms, and C has the abovementioned meaning, in the presence of propylphosphonic anhydride, eliminating the protective group U' by hydrogenolysis, reacting the resulting compound of the formula IV
H - B - C - OR IV
with a compound of the formula V
U - A - OH V
in the presence of propylphosphonic anhydride, and finally eliminating R enzymatically.
U - A - B - C - OH I
in which U denotes hydrogen or a urethane protective group A denotes a natural .alpha.-amino acid or derivatives thereof B denotes a natural .alpha.-amino acid or derivatives thereof and C denotes an aromatic .alpha.-amino acid, which comprises reacting a compound of the formula II
U' - B - OH II
in which U' is a urethane protective group which can be eliminated by hydrogenolysis, and B has the above-mentioned meaning, with a compound of the formula III
H - C - OR III
in which R represents alkyl having 1 to 4 carbon atoms, and C has the abovementioned meaning, in the presence of propylphosphonic anhydride, eliminating the protective group U' by hydrogenolysis, reacting the resulting compound of the formula IV
H - B - C - OR IV
with a compound of the formula V
U - A - OH V
in the presence of propylphosphonic anhydride, and finally eliminating R enzymatically.
2. The process as claimed in claim 1, wherein U denotes benzyloxycarbonyl or tert.-butyloxycrbonyl.
3. The process as claimed in claim 1, wherein A and/or B denote Gly, Ala, Ser, Thr, Val, Leu, Ile, Glu, Gln, p-Glu, Tyr, Phe, Trp or His.
4. The process as claimed in claim 2, wherein A and/or B denote Gly, Ala, Ser, Thr, Val, Leu, Ile, Glu, Gln, p-Glu, Tyr, Phe, Trp or His.
5. The process as claimed in any one of claims 1 to 4, wherein A and/or B denote Ser, Thr, Trp or Phe.
6. The process as claimed in any one of claims 1 to 4, wherein C denotes Tyr or Phe.
7. The process as claimed in any one of claims 1 to 4, wherein U' denotes benzyloxycarbonyl.
8. The process as claimed in any one of claims 1 to 4, wherein R denotes methyl.
9. The process as claimed in any one of claims 1 to 4, wherein the reactions are carried out in the presence of propylphosphonic anhydride in an ethyl acetate/water mixture.
10. The process as claimed in any one of claims 1 to 4, wherein the enzymatic esterolysis is carried out with trypsin and/or .alpha.-chymotrypsin.
11. The process as claimed in any one of claims 1 to 4, wherein U and U' denote benzyloxycarbonyl, A denotes Trp, B
denotes Ser, C denotes Tyr and R denotes methyl.
denotes Ser, C denotes Tyr and R denotes methyl.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| DE3839379A DE3839379A1 (en) | 1988-11-22 | 1988-11-22 | METHOD FOR PRODUCING TRIPEPTIDES |
| DEP3839379.4 | 1988-11-22 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CA1335493C true CA1335493C (en) | 1995-05-09 |
Family
ID=6367619
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA000614545A Expired - Lifetime CA1335493C (en) | 1988-11-22 | 1989-09-29 | Process for the preparation of tripeptides |
Country Status (10)
| Country | Link |
|---|---|
| EP (1) | EP0370399B1 (en) |
| JP (1) | JP2843618B2 (en) |
| AT (1) | ATE124053T1 (en) |
| AU (1) | AU626608B2 (en) |
| CA (1) | CA1335493C (en) |
| DE (2) | DE3839379A1 (en) |
| DK (1) | DK173690B1 (en) |
| ES (1) | ES2075028T3 (en) |
| IE (1) | IE67293B1 (en) |
| IL (1) | IL92368A0 (en) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| DE102004029811A1 (en) * | 2004-06-19 | 2006-01-12 | Clariant Gmbh | Process for the preparation of alkenes by elimination of water from alcohols with alkylphosphonic anhydrides |
| DE102004029812A1 (en) * | 2004-06-19 | 2006-05-24 | Clariant Gmbh | Process for the preparation of nitriles from aldehyde oximes by reaction with alkylphosphonic anhydrides |
Family Cites Families (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| SE7801373L (en) * | 1978-02-07 | 1979-08-08 | Kabi Ab | EASY SPLABLE SUBSTRATE FOR QUANTIFIATION OF PROTEASES |
| US4293648A (en) * | 1979-12-12 | 1981-10-06 | G. D. Searle & Co. | Process for esterification of α-L-aspartyl-L-phenylalanine |
| DE3411224A1 (en) * | 1984-03-27 | 1985-10-10 | Hoechst Ag, 6230 Frankfurt | METHOD FOR THE RACEMATAR PRODUCTION OF INTERMEDIATE PEPTIDE PRODUCTS OF THE GONADORELIN AND GONADORELINANALOGA SYNTHESIS AND NEW INTERMEDIATE PRODUCTS IN THIS METHOD |
| DE3438189A1 (en) * | 1984-10-18 | 1986-04-24 | Hoechst Ag, 6230 Frankfurt | METHOD FOR PRODUCING AROMATICALLY SUBSTITUTED L-AMINO ACIDS |
| US4888385A (en) * | 1987-04-30 | 1989-12-19 | Millipore Corporation | BOP reagent for solid phase peptide synthesis |
| US4946942A (en) * | 1988-03-11 | 1990-08-07 | Bioresearch, Inc. | Urethane-protected amino acid-N-carboxyanhydrides |
-
1988
- 1988-11-22 DE DE3839379A patent/DE3839379A1/en not_active Withdrawn
-
1989
- 1989-09-29 CA CA000614545A patent/CA1335493C/en not_active Expired - Lifetime
- 1989-11-17 AT AT89121277T patent/ATE124053T1/en not_active IP Right Cessation
- 1989-11-17 EP EP89121277A patent/EP0370399B1/en not_active Expired - Lifetime
- 1989-11-17 ES ES89121277T patent/ES2075028T3/en not_active Expired - Lifetime
- 1989-11-17 DE DE58909308T patent/DE58909308D1/en not_active Expired - Lifetime
- 1989-11-20 IL IL92368A patent/IL92368A0/en not_active IP Right Cessation
- 1989-11-21 DK DK198905844A patent/DK173690B1/en not_active IP Right Cessation
- 1989-11-21 IE IE371789A patent/IE67293B1/en not_active IP Right Cessation
- 1989-11-21 JP JP1300960A patent/JP2843618B2/en not_active Expired - Lifetime
- 1989-11-21 AU AU45328/89A patent/AU626608B2/en not_active Expired
Also Published As
| Publication number | Publication date |
|---|---|
| DK173690B1 (en) | 2001-06-25 |
| JPH02219587A (en) | 1990-09-03 |
| JP2843618B2 (en) | 1999-01-06 |
| DK584489A (en) | 1990-05-23 |
| AU626608B2 (en) | 1992-08-06 |
| ATE124053T1 (en) | 1995-07-15 |
| EP0370399A3 (en) | 1991-09-18 |
| EP0370399B1 (en) | 1995-06-21 |
| IE893717L (en) | 1990-05-22 |
| DE58909308D1 (en) | 1995-07-27 |
| DK584489D0 (en) | 1989-11-21 |
| DE3839379A1 (en) | 1990-05-23 |
| AU4532889A (en) | 1990-05-31 |
| EP0370399A2 (en) | 1990-05-30 |
| IL92368A0 (en) | 1990-07-26 |
| ES2075028T3 (en) | 1995-10-01 |
| IE67293B1 (en) | 1996-03-20 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| US5015728A (en) | Process for the preparation of insulin derivatives, the B chain of which is lengthened c-terminally | |
| US6251625B1 (en) | Process for preparing peptides and N-carbamoyl-protected peptides | |
| US5382679A (en) | Process for the preparation of glutathione S-acyl derivatives, compounds obtained from said process and an intermediate useful for the preparation thereof | |
| KR0163224B1 (en) | Process for manufacture of 1-deamino-8-d-arginine vasopressin | |
| JPH0141317B2 (en) | ||
| CA1174973A (en) | Process for preparing polypeptide derivatives | |
| EP0149594A2 (en) | Enzymatic coupling of n-formyl amino acids and/or peptide residues | |
| CA1335493C (en) | Process for the preparation of tripeptides | |
| CA1195273A (en) | Process for converting preproinsulin analogs into insulin | |
| US5191065A (en) | Process for the preparation of tripeptides | |
| US4426325A (en) | Process for the preparation of compounds containing carboxylic acid amide groups, in particular or peptides | |
| SE452318B (en) | AMINO ACIDS FOR USE AS INTERMEDIATES IN THE PRODUCTION OF BESTATIN | |
| Hemmi et al. | STUDIES ON A NEW IMMUNOACTIVE PEPTIDE, FK-156 IV. SYNTHESIS OF FK-156 AND ITS GEOMETRIC ISOMER | |
| US4107158A (en) | Process for making an octapeptide useful for the treatment of diabetes | |
| Stepanov et al. | Subtilisin and α-chymotrypsin catalyzed synthesis of peptides containing arginine and lysine p-nitroanilides as c-terminal moieties | |
| CA2223911C (en) | The preparation of active peptides | |
| EP0149593B1 (en) | Novel thiopeptolide substrates for vertebrate collagenase | |
| US3749704A (en) | N-(omega-amino lower alkyl)-amides of 1,17-modified acth peptides | |
| Garg et al. | The synthesis of protected glycopeptides containing the amino acid sequences 34–37 and 34–38 of bovine ribonuclease B | |
| HU221619B1 (en) | Process for synthetise peptides and intermediates thereof | |
| Ohno et al. | Partial enzymic deprotection in the synthesis of a protected octapeptide bearing a free terminal carboxyl group | |
| US3247178A (en) | Synthesis of peptides containing alpha, omega-diamino acids protected by phthalyl and t-butyloxycarbonyl groups | |
| US4212796A (en) | Process for the preparation of cysteine-containing peptides | |
| Metzger et al. | Synthesis of a B-lymphocyte activating α-methylserine containing lipopentapeptide | |
| G. Bayryamov et al. | The Two Pathways for Effective Orthogonal Protection of L-Ornithine, for Amino Acylation of 5'-O-Pivaloyl Nucleosides, Describe the General and Important Role for the Successful Imitation, During the Synthesis of the Model Substrates for the Ribosomal Mimic Reaction |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| MKEX | Expiry |