CH587865A5 - Reagent for radio-immunoassay of testosterone - by reacting testerone-oxime with tyrosine ester - Google Patents
Reagent for radio-immunoassay of testosterone - by reacting testerone-oxime with tyrosine esterInfo
- Publication number
- CH587865A5 CH587865A5 CH1659573A CH1659573A CH587865A5 CH 587865 A5 CH587865 A5 CH 587865A5 CH 1659573 A CH1659573 A CH 1659573A CH 1659573 A CH1659573 A CH 1659573A CH 587865 A5 CH587865 A5 CH 587865A5
- Authority
- CH
- Switzerland
- Prior art keywords
- testosterone
- methyl ester
- carboxymethyl
- hormone
- tme
- Prior art date
Links
- MUMGGOZAMZWBJJ-DYKIIFRCSA-N Testostosterone Chemical compound O=C1CC[C@]2(C)[C@H]3CC[C@](C)([C@H](CC4)O)[C@@H]4[C@@H]3CCC2=C1 MUMGGOZAMZWBJJ-DYKIIFRCSA-N 0.000 title claims description 14
- 229960003604 testosterone Drugs 0.000 title claims description 7
- 239000003153 chemical reaction reagent Substances 0.000 title claims description 5
- 238000003127 radioimmunoassay Methods 0.000 title 1
- OUYCCCASQSFEME-UHFFFAOYSA-N tyrosine Natural products OC(=O)C(N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-UHFFFAOYSA-N 0.000 title 1
- -1 tyrosine ester Chemical class 0.000 title 1
- 239000002904 solvent Substances 0.000 claims abstract description 12
- QOSSAOTZNIDXMA-UHFFFAOYSA-N Dicylcohexylcarbodiimide Chemical compound C1CCCCC1N=C=NC1CCCCC1 QOSSAOTZNIDXMA-UHFFFAOYSA-N 0.000 claims abstract description 11
- 150000004702 methyl esters Chemical class 0.000 claims abstract description 11
- VDYLVWGBLQNNAW-UVTKPVMCSA-N testosterone (E)-3-(O-carboxymethyl)oxime Chemical compound OC(=O)CO/N=C/1CC[C@]2(C)[C@H]3CC[C@](C)([C@H](CC4)O)[C@@H]4[C@@H]3CCC2=C\1 VDYLVWGBLQNNAW-UVTKPVMCSA-N 0.000 claims abstract description 11
- MWZPENIJLUWBSY-VIFPVBQESA-N methyl L-tyrosinate Chemical compound COC(=O)[C@@H](N)CC1=CC=C(O)C=C1 MWZPENIJLUWBSY-VIFPVBQESA-N 0.000 claims abstract description 7
- 229940088597 hormone Drugs 0.000 claims description 22
- 239000005556 hormone Substances 0.000 claims description 22
- 238000000034 method Methods 0.000 claims description 22
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 18
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 16
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 claims description 12
- 238000012360 testing method Methods 0.000 claims description 10
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 claims description 8
- 230000015572 biosynthetic process Effects 0.000 claims description 8
- ZMANZCXQSJIPKH-UHFFFAOYSA-N Triethylamine Chemical compound CCN(CC)CC ZMANZCXQSJIPKH-UHFFFAOYSA-N 0.000 claims description 6
- 238000003786 synthesis reaction Methods 0.000 claims description 6
- 150000008064 anhydrides Chemical class 0.000 claims description 4
- 238000011088 calibration curve Methods 0.000 claims description 4
- 238000006243 chemical reaction Methods 0.000 claims description 4
- 230000001419 dependent effect Effects 0.000 claims description 4
- 230000000694 effects Effects 0.000 claims description 4
- 238000002360 preparation method Methods 0.000 claims description 4
- 239000000047 product Substances 0.000 claims description 4
- 239000007787 solid Substances 0.000 claims description 4
- 150000003431 steroids Chemical class 0.000 claims description 4
- 238000010998 test method Methods 0.000 claims description 4
- YOETUEMZNOLGDB-UHFFFAOYSA-N 2-methylpropyl carbonochloridate Chemical compound CC(C)COC(Cl)=O YOETUEMZNOLGDB-UHFFFAOYSA-N 0.000 claims description 2
- WDJUZGPOPHTGOT-OAXVISGBSA-N Digitoxin Natural products O([C@H]1[C@@H](C)O[C@@H](O[C@@H]2C[C@@H]3[C@@](C)([C@@H]4[C@H]([C@]5(O)[C@@](C)([C@H](C6=CC(=O)OC6)CC5)CC4)CC3)CC2)C[C@H]1O)[C@H]1O[C@@H](C)[C@H](O[C@H]2O[C@@H](C)[C@@H](O)[C@@H](O)C2)[C@@H](O)C1 WDJUZGPOPHTGOT-OAXVISGBSA-N 0.000 claims description 2
- 239000000427 antigen Substances 0.000 claims description 2
- 102000036639 antigens Human genes 0.000 claims description 2
- 108091007433 antigens Proteins 0.000 claims description 2
- WDJUZGPOPHTGOT-XUDUSOBPSA-N digitoxin Chemical compound C1[C@H](O)[C@H](O)[C@@H](C)O[C@H]1O[C@@H]1[C@@H](C)O[C@@H](O[C@@H]2[C@H](O[C@@H](O[C@@H]3C[C@@H]4[C@]([C@@H]5[C@H]([C@]6(CC[C@@H]([C@@]6(C)CC5)C=5COC(=O)C=5)O)CC4)(C)CC3)C[C@@H]2O)C)C[C@@H]1O WDJUZGPOPHTGOT-XUDUSOBPSA-N 0.000 claims description 2
- 229960000648 digitoxin Drugs 0.000 claims description 2
- 238000000921 elemental analysis Methods 0.000 claims description 2
- 238000001704 evaporation Methods 0.000 claims description 2
- 230000008020 evaporation Effects 0.000 claims description 2
- 239000000706 filtrate Substances 0.000 claims description 2
- 238000001914 filtration Methods 0.000 claims description 2
- 230000001900 immune effect Effects 0.000 claims description 2
- 230000028993 immune response Effects 0.000 claims description 2
- 238000002329 infrared spectrum Methods 0.000 claims description 2
- 238000002372 labelling Methods 0.000 claims description 2
- 238000004519 manufacturing process Methods 0.000 claims description 2
- 238000005259 measurement Methods 0.000 claims description 2
- 230000008018 melting Effects 0.000 claims description 2
- 238000002844 melting Methods 0.000 claims description 2
- VXYFARNRGZWHTJ-FVGYRXGTSA-N methyl (2s)-2-amino-3-(4-hydroxyphenyl)propanoate;hydrochloride Chemical compound Cl.COC(=O)[C@@H](N)CC1=CC=C(O)C=C1 VXYFARNRGZWHTJ-FVGYRXGTSA-N 0.000 claims description 2
- 239000012454 non-polar solvent Substances 0.000 claims description 2
- 230000000144 pharmacologic effect Effects 0.000 claims description 2
- 230000002285 radioactive effect Effects 0.000 claims description 2
- 239000011541 reaction mixture Substances 0.000 claims description 2
- 238000001953 recrystallisation Methods 0.000 claims description 2
- 210000002966 serum Anatomy 0.000 claims description 2
- 239000003270 steroid hormone Substances 0.000 claims description 2
- 238000003756 stirring Methods 0.000 claims description 2
- 239000000725 suspension Substances 0.000 claims description 2
- 238000001308 synthesis method Methods 0.000 claims description 2
- 238000002211 ultraviolet spectrum Methods 0.000 claims description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 2
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07J—STEROIDS
- C07J41/00—Normal steroids containing one or more nitrogen atoms not belonging to a hetero ring
- C07J41/0005—Normal steroids containing one or more nitrogen atoms not belonging to a hetero ring the nitrogen atom being directly linked to the cyclopenta(a)hydro phenanthrene skeleton
- C07J41/0016—Oximes
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/74—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving hormones or other non-cytokine intercellular protein regulatory factors such as growth factors, including receptors to hormones and growth factors
- G01N33/743—Steroid hormones
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Molecular Biology (AREA)
- Engineering & Computer Science (AREA)
- Hematology (AREA)
- General Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Urology & Nephrology (AREA)
- Biomedical Technology (AREA)
- Immunology (AREA)
- Cell Biology (AREA)
- Biotechnology (AREA)
- Microbiology (AREA)
- Endocrinology (AREA)
- Food Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Steroid Compounds (AREA)
Abstract
Testosterone-3-(O-carboxymethyl)-Oximetyrosine methyl ester is prepared by reacting testosterone-3-(O-carboxymethyl)-oxime and tyrosine methyl ester in the presence of N,N-dicyclohexylcarbodiimide in an apolar solvent at 0-25 degrees C. Then the solvent is evaporated and the crude residue is crystallised.
Description
Die vorliegende Erfindung betrifft ein Verfahren zur Herstellung von Testosteron-3-(O-carboxymethyl)oximtyro- sinmethylester.
Immunologische Hormonversuchsmethoden beruhen auf der Fähigkeit eines gegebenen Hormons, bei einer gegebenen immunologischen Reaktion als Antigen zu wirken.
Gestützt auf dieses Prinzip sind in letzter Zeit Radioimmunversuchsmethoden entwickelt worden, bei denen eine unbekannte Menge des zu bestimmenden unmarkierten (kalten) Hormons in Gegenwart einer bekannten Menge desselben markierten Hormons mit einer geeigneten Menge eines spezifischen Antiserums geimpft wird. So kann eine unbekannte Hormonmenge leicht aus der Radioaktivitätsbestimmung des Antigen-Antikörperkomplexes durch Vergleich mit einer vorgängig aufgenommenen Eichkurve ermittelt werden. Eine solche Eichkurve wird unter Verwendung von spezifischem Antiserum, kaltem Hormon und markiertem Hormon in bekannten Mengen aufgenommen, wobei kaltes und markiertes Hormon proportional zu ihren Mengen an der Bildung des genannten Antigen-Antikörperkomplexes beteiligt sind.
Offensichtlich ist die Radioaktivitätszählung des genannten Antigen-Antikörperkomplexes umgekehrt proportional zur eingesetzten Menge an kaltem Hormon, wenn die Menge an Antiserum und markiertem Hormon gleich bleibt.
Eine Radioimmunversuchsmethode zur Messung von Steroid-Hormonen, welche auf der Verwendung von 125J markierten Steroid-Tyrosinmethylester-Konjugaten beruht, ist von Oliver et al., in J. Clin. Investig. 47, 1035 (1968) vorgeschlagen und in bezug auf ihre Anwendung auf die Messung pharmakologischer Digitoxinkonzentrationen im menschlichen Blutserum spezifisch beschrieben worden.
Bei der Anwendung auf den Radioimmunversuch von Testosteron bedingt diese Methode die Synthese des konjugaten Testosteron-3-(O-carboxymethyl)oximtyrosinmethylesters (im folgenden T-3-TME genannt), dessen Markierung mit 125J und Verwendung als radioaktives Hapten bei der radioimmunologischen Reaktion.
Midgley et al. haben in Recent Progress in Hormone Research, 27, 236 (1971) festgestellt, dass die relative molare Aktivität von T-3-TME beim Testosteron-3-Radioimmunversuch dem 0,7-fachen des Wertes von Testosteron entspricht.
Dies bedeutet, dass z'var ein konjugates Steroid wie beispielsweise T-3-TME nicht exakt im selben Verhältnis wie das entsprechende native Steroid an den Antikörper gebunden sein kann, dass jedoch die Differenz klein ist, sofern überhaupt eine solche vorliegt.
Aus den obigen Ausführungen geht die Brauchbarkeit des T3-TME-Konjugates beim Radioimmunversuch von Testosteron klar hervor.
Die Synthese von T-3-TME ist nicht speziell beschrieben worden, Die einzige vorgeschlagene Methode ist die Isobutylchlorformiat (gemischtes Anhydrid) Synthesemethode, welche von Oliver et al. am Beispiel der Synthese vom 3 -O-succinyldigitoxigenintyrosinmethylester-Konjugat beschrieben worden ist. Wenn jedoch die gemischte Anhydrid Methode zur Synthese von T-3-TME gebraucht wird, kann keine feststellbare Ausbeute an gewünschtem Produkt erzielt werden.
Das erfindungsgemässe Verfahren zur Herstellung von Testosteron-3-(O-carboxymethyl)oximtyrosinmethylester ist dadurch gekennzeichnet, dass Testosteron-3-(O-carboxymethyl)oxim in Gegenwart von N,N-Dicyclohexylcarbodiimid mit Tyrosinmethylester in einem apolaren Lösungsmittel bei einer Temperatur von 0-25 C umgesetzt wird, das Lösungsmittel abgedampft und der rohe Rückstand kristallisiert wird.
Bevorzugte Verfahrensbedingungen sind die folgenden: Äquimolare Mengen der Reagenzien bei leichtem molarem Überschuss an N,N-Dicyclohexylcarbodiimid; apolare Lösungsmittel wie z.B. Methylenchlorid, N,N-Dimethylformamid und Chloroform; Reaktionstemperaturen von 0 bis 25ob, vorzugsweise etwa 10ob.
Das folgende Beispiel soll das erfindungsgemässe Verfahren noch besser veranschaulichen:
Beispiel
Unter Rühren wurden zu einer Suspension von 2,31 g Tyrosinmethylesterhydrochlorid in 80 ml Methylenchlorid 1,4 ml Triäthylamin und, 30 Minuten später, 3,61 g Testosteron-3 (O-carboxymethyl)oxim zugefügt.
Die Reaktionsmischung wurde auf -10"C gekühlt, mit
2,47 g N,N-dicyclohexylcarbodiimid versetzt, wieder auf + 10oC gebracht und 24 Stunden bei dieser Temperatur gehalten.
Nach dem Abfiltrieren und Eindampfen im Vakuum bis zur Trockenheit wurde der halbfeste Rückstand in 50 ml 50%igem Äthanol gelöst und die Lösung filtriert. Das Filtrat wurde im Vakuum eingedampft, wodurch ein öliger Rückstand erhalten wurde. Dieser wurde durch Zugabe von etwa 20 ml dest.
Wasser zu einer Lösung des Rückstandes in 10 ml 95 %-igem Äthanol in Form eines kristallinen Festkörpers ausgefällt.
Die Ausbeute an kristallinem Testosteron-3-(O-carboxymethyl)oxim-TME betrug 2,15 g (40%).
Nach Umkristallisierung aus wässrigem Äthanol betrug der
Schmelzpunkt 94-95"C.
Elementaranalyse für C31H4206N2:
C H N Berechnet 69,12% 7,86% 5,20% Gefunden 68,95 % 8,36% 5,06%
Die UV- und IR-Spektren stimmten völlig mit der erwarteten Struktur von T-3-TME überein.
Das so erhaltene Produkt wurde nach Greenwood et al., Biochem. J. 89, 114 (1963) mit 125J markiert, wodurch ein, für den Radioimmunversuch von Testosteron-3 geeignetes Reagenz von hoher spezifischer Wirksamkeit erhalten wurde.
PATENTANSPRUCH
Verfahren zur Herstellung von Testosteron-3-(O-carboxymethyl)oximtyrosinmethylester, dadurch gekennzeichnet, dass Testosteron-3-(O-carboxymethyl)oxim in Gegenwart von N,N-Dicyclohexylcarbodiimid mit Tyrosinmethylester in einem apolaren Lösungsmittel bei einer Temperatur von 0-25oC umgesetzt wird, das Lösungsmittel abgedampft und der rohe Rückstand kristallisiert wird.
UNTERANSPRÜCHE
1. Verfahren nach Patentanspruch, dadurch gekennzeichnet, dass als apolares Lösungsmittel Methylenchlorid, N,N-Dimethylformamid oder Chloroform verwendet wird.
2. Verfahren nach Patentanspruch oder Unteranspruch 1, dadurch gekennzeichnet, dass äquimolare Mengen Testosteron-3-(O-carboxymethyl)oxim und Tyrosinmethylester in Gegenwart eines geringen molaren Überschusses an N,N-Dicyclohexylcarbodiimid miteinander umgesetzt werden.
3. Verfahren nach Patentanspruch oder Unteranspruch 1, dadurch gekennzeichnet, dass der rohe Rückstand aus wässrigem Äthanol umkristallisiert wird.
**WARNUNG** Ende DESC Feld konnte Anfang CLMS uberlappen**.
The present invention relates to a process for the production of testosterone-3- (O-carboxymethyl) oximtyrosine methyl ester.
Hormone immunological testing methods rely on the ability of a given hormone to act as an antigen in a given immunological response.
Based on this principle, radioimmune test methods have recently been developed in which an unknown amount of the unlabeled (cold) hormone to be determined is inoculated in the presence of a known amount of the same labeled hormone with a suitable amount of a specific antiserum. Thus, an unknown amount of hormone can easily be determined from the radioactivity determination of the antigen-antibody complex by comparison with a previously recorded calibration curve. Such a calibration curve is recorded using specific antiserum, cold hormone and labeled hormone in known amounts, with cold and labeled hormones participating in the formation of said antigen-antibody complex in proportion to their amounts.
Obviously, the radioactivity count of said antigen-antibody complex is inversely proportional to the amount of cold hormone used if the amount of antiserum and labeled hormone remains the same.
A radioimmune test method for measuring steroid hormones which is based on the use of 125 I-labeled steroid-tyrosine methyl ester conjugates is described by Oliver et al., In J. Clin. Investig. 47, 1035 (1968) and specifically described with respect to their application to the measurement of pharmacological digitoxin concentrations in human blood serum.
When applied to the radioimmune test of testosterone, this method requires the synthesis of the conjugate testosterone-3- (O-carboxymethyl) oximtyrosine methyl ester (hereinafter referred to as T-3-TME), its labeling with 125J and use as a radioactive hapten in the radioimmunological reaction.
Midgley et al. have found in Recent Progress in Hormone Research, 27, 236 (1971) that the relative molar activity of T-3-TME in the testosterone-3 radioimmune test is 0.7 times the value of testosterone.
This means that z'var a conjugate steroid such as T-3-TME cannot be bound to the antibody in exactly the same ratio as the corresponding native steroid, but that the difference is small, if there is any.
From the above, the usefulness of the T3-TME conjugate in the radioimmune test of testosterone is clear.
The synthesis of T-3-TME has not been specifically described. The only method proposed is the isobutyl chloroformate (mixed anhydride) synthesis method, which is described by Oliver et al. has been described using the example of the synthesis of 3 -O-succinyldigitoxigenintyrosine methyl ester conjugate. However, when the mixed anhydride method is used to synthesize T-3-TME, no detectable yield of the desired product can be achieved.
The inventive method for the preparation of testosterone-3- (O-carboxymethyl) oximtyrosine methyl ester is characterized in that testosterone-3- (O-carboxymethyl) oxime in the presence of N, N-dicyclohexylcarbodiimide with tyrosine methyl ester in an apolar solvent at a temperature of 0 -25 C is reacted, the solvent is evaporated and the crude residue is crystallized.
Preferred process conditions are as follows: equimolar amounts of the reagents with a slight molar excess of N, N-dicyclohexylcarbodiimide; apolar solvents such as Methylene chloride, N, N-dimethylformamide and chloroform; Reaction temperatures from 0 to 25ob, preferably about 10ob.
The following example is intended to illustrate the method according to the invention even better:
example
With stirring, 1.4 ml of triethylamine and, 30 minutes later, 3.61 g of testosterone-3 (O-carboxymethyl) oxime were added to a suspension of 2.31 g of tyrosine methyl ester hydrochloride in 80 ml of methylene chloride.
The reaction mixture was cooled to -10 "C, with
2.47 g of N, N-dicyclohexylcarbodiimide were added, brought back to + 10oC and kept at this temperature for 24 hours.
After filtering off and evaporation in vacuo to dryness, the semi-solid residue was dissolved in 50 ml of 50% ethanol and the solution was filtered. The filtrate was evaporated in vacuo to give an oily residue. This was by adding about 20 ml of dist.
Water precipitated into a solution of the residue in 10 ml of 95% ethanol in the form of a crystalline solid.
The yield of crystalline testosterone-3- (O-carboxymethyl) oxime TME was 2.15 g (40%).
After recrystallization from aqueous ethanol, the
Melting point 94-95 "C.
Elemental analysis for C31H4206N2:
C H N Calculated 69.12% 7.86% 5.20% Found 68.95% 8.36% 5.06%
The UV and IR spectra were completely consistent with the expected structure of T-3-TME.
The product thus obtained was according to Greenwood et al., Biochem. J. 89, 114 (1963) marked with 125J, whereby a reagent suitable for the radioimmune test of testosterone-3 and having a high specific activity was obtained.
PATENT CLAIM
Process for the preparation of testosterone-3- (O-carboxymethyl) oximtyrosine methyl ester, characterized in that testosterone-3- (O-carboxymethyl) oxime in the presence of N, N-dicyclohexylcarbodiimide with tyrosine methyl ester in an apolar solvent at a temperature of 0-25oC is reacted, the solvent is evaporated and the crude residue is crystallized.
SUBCLAIMS
1. The method according to claim, characterized in that methylene chloride, N, N-dimethylformamide or chloroform is used as the apolar solvent.
2. The method according to claim or dependent claim 1, characterized in that equimolar amounts of testosterone 3- (O-carboxymethyl) oxime and tyrosine methyl ester are reacted with one another in the presence of a small molar excess of N, N-dicyclohexylcarbodiimide.
3. The method according to claim or dependent claim 1, characterized in that the crude residue is recrystallized from aqueous ethanol.
** WARNING ** End of DESC field could overlap beginning of CLMS **.
Claims (1)
Priority Applications (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| NL7314599A NL7314599A (en) | 1973-10-24 | 1973-10-24 | PROCEDURE FOR PREPARING A REAGENT FOR RADIOIMMUNOLOGICAL DETERMINATION OF TESTOSTERONE. |
| CH1659573A CH587865A5 (en) | 1973-10-24 | 1973-11-26 | Reagent for radio-immunoassay of testosterone - by reacting testerone-oxime with tyrosine ester |
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| NL7314599A NL7314599A (en) | 1973-10-24 | 1973-10-24 | PROCEDURE FOR PREPARING A REAGENT FOR RADIOIMMUNOLOGICAL DETERMINATION OF TESTOSTERONE. |
| CH1659573A CH587865A5 (en) | 1973-10-24 | 1973-11-26 | Reagent for radio-immunoassay of testosterone - by reacting testerone-oxime with tyrosine ester |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CH587865A5 true CH587865A5 (en) | 1977-05-13 |
Family
ID=25718198
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CH1659573A CH587865A5 (en) | 1973-10-24 | 1973-11-26 | Reagent for radio-immunoassay of testosterone - by reacting testerone-oxime with tyrosine ester |
Country Status (2)
| Country | Link |
|---|---|
| CH (1) | CH587865A5 (en) |
| NL (1) | NL7314599A (en) |
-
1973
- 1973-10-24 NL NL7314599A patent/NL7314599A/en unknown
- 1973-11-26 CH CH1659573A patent/CH587865A5/en not_active IP Right Cessation
Also Published As
| Publication number | Publication date |
|---|---|
| NL7314599A (en) | 1975-04-28 |
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