[2] a ) C. MurLean, E . L. Mackor, Discuss. Faraday Soc. 34, 165 (1962); b) E. L. Mackor, C . MacLean, Pure Appl. Chem. 8, 393 (1964); c) D. M . Brouwer, C . MacLean, E . L . Mackor, Discuss. Faraday Soc. 39, 121 (1965): d) 7: Birchall, R. J . Gillespie, Can. J. Chem. 42, 502 (1964); e) G. A. Olah, R . H . Schlosberg, D. P. Kelly, G . D. Matrescu, J. Am. Chem. SOC.92, 2546 (1970). [3] J. L. Derlin, I l l , J . F . WOK R . W Taf, W J . Hehre, J. Am. Chem. Soc. 98, 1990 (1976). Additivity of alkyl substituent effects was assumed in that work. [4] The NMR spectra were obtained in the Fourier transform mode, at 25.2 MHz. using a Varian XL-100 instrument with a variable temperature probe. [ 5 ] Acid A is well over lo' times stronger than acid B. 161 The solution in acid B partially froze at lower temperature. [7] The peaks are still broad in acid A at + 10°C.
Facile Synthesis of Arabinoadenosine
3'-Phosphate[**I
By Rudolf Mengel and Harald Wiedner"] Only few examples of oxirane ring opening by phosphate ions are known in nucleoside chemistry"]. We demonstrated that adenosine can be transformed into its lyxo-epoxide ( I )r21 and that this product can be opened with mineral acid or alkali halide in the presence of boron trifluoride-etherate to give predominantly 3'-substituted halon~cleosides'~~. We have now examined the cleavage of the oxirane ring of ( 1 ) with phosphoric acid in hexamethylphosphoric triamide. Fourteen hours' heating to 100°C affords arabinoadenosine 3'-phosphate ( 2 ) which can be precipitated as the barium salt. On electrophoresis[4a1and chromatography[4b1the salt behaves in a manner resembling adenosine 5'-monophosphate and is cleaved by alkaline phosphatase''"] to arabinoadenosine together with traces of xyloadenosine[4'l. After reprecipitation of the salt and treatment with phosphatase, xyloadenosine can no longer be detected. Treatment with 5'-nucleotida~e[~~] (18 h) has no effect on ( 2 ) , while 3'-nu~leotidase[''~cleaves the compound to the extent of 90% within 18 h. The 90 MHz 'H-NMR spectrum of (2) was measured against sodium 2,2-dimethyl-2-silapentanesulfonate as internal standard in D,O to which had been added one drop of trifluoroacetic acid (to shift the HOD signal away from the signals due to the sugar protons). It showed signals at
O=+O
Arabinoadenosine 3'-phosphate (2) could serve as an "arabino" building block in oligonucleotide synthesis and as starting material for the synthesis of arabino-cAMP[61.Compound (2) also warrants interest in connection with the fact that arabinoadenosine has an inhibitory action on DNA polymerase, ribonucleotide reductase, adenyl cyclase, various viruses, and various animal tumors.
Procedure A solution of adenosine lyxoepoxide ( I ) (100mg, 0.4 mmol) and phosphoric acid (crystalline, 98 %; 200mg, 1.7mmol) in hexamethylphosphoric triamide (2 ml) is stirred for 14h at 100°C. After neutralization with conc. ammonia most of the solvent is removed in uacuo at 100°C. After addition of 10 % barium acetate solution (6 ml), triethylamine (0.4ml), and ethanol (12ml) a precipitate is formed. The mixture is allowed to stand for 12h at 4°C and then centrifuged; the residue is washed with ethanol ( 3 x IOml), and then with water (7 x 10ml). The combined aqueous extracts are evaporated to IOml, treated with ethanol (10ml), and left to stand for 10h at 4°C. The precipitate formed is filtered off and applied to a Sephadex DEAE A 25 column (2.3cm x 35 cm). Elution is performed with a gradient of triethylammonium hydrogen carbonate (0.05 to 0.2 M ; 2 liter); 25-ml fractions are collected. Fractions 33 to 47 contain 0.153 mmol of (2) (yield 38%). They are concentrated and freeze dried. Received: February 17, 1977 [Z 678 IE] German version: Angew. Chem. 89, 328 (1977) CAS Registry numbers: ( l ) , 40110-98-3: (Z), 54621-41-9
W E . Harue!, J . J . Michalsky, A . R . Todd, J. Chem. Soc. 1951, 2271: K . Tadashi, A. Mituse, J . Motonobu, N . Tokrrro, Japan. Patent Kokai 74, 18879 (CI. 16E 461) 1974: Chem. Abstr. 80, 121290e (1974). M. J . Robins, Y Fonron, R. Mengel, J. Org. Chem. 39, 1564 (1974). R . Mengel, H . Wiedner, Chem. Ber. 109, 1395 (1976). a) Triethylammonium hydrogen carbonate, pH =7.5, IOOOV: b) o n PEI cellulose with I N NHaOAc/ethanol ( I : I ) ; c) xyloadenosine and arabinoadenosine can be separated by electrophoresis in borate buffer (pH = 9) at 1000 V. a) Boehringer (Mannheim); b) [E.C. 3.1.3.51, Sigma Comp.; c) [E.C. 3.1.3.61, Sigma Comp. 7: A. Khwaja, R . Harris, R . K . Robins, Tetrahedron Lett. 1972, 4681, and references cited therein.-Cf. the analogous synthesis of 9-p-D-arabinofuranosylcytosine 3',5'-cyclophosphate: R. A . Long, G . L . Szekeres, T: A . Kliwaja, R . W. Sidwell, L. N . Simon, R . K. Robins, J. Med. Chem. 1 5 , 1215 (1972).
Side Reactions in Peptide Synthesis: tert-Butylation of Tryptophan By Erich Wiinsch, Ernst Jaeger, Lajos Kisfaludy, and Miklos LOW[*]
6=8.6 (s, 1, Hs), 8.46 (s, 1, HZ), 6.5 (d, Jt,-2,=5.2HZ, 1, HI,),4.86 (m, 1, H3,),4.74 (q, Jz,-3.=4Hz, 1 , H2,),4.29 (quart, 1, H4.), 3.95ppm ("d, 2H, Hs,,, H5.b).The chemical shift of the H atom attached to C-3' (shifted downfield by 0.5 ppm relative to adenosine) and the coupling constant between 31P and 3'-H, show the phosphate group to be located in position 3'.
Indoie and its derivatives are known to be very reactive. The amino acid tryptophan contains an indole ring as a third function, thus incorporating a relatively reactive moiety into the peptide chain. Side reactions due to indole have been described and postulated in peptide chemistry, e. g . also under conditions of acidolytic removal of protective groups from
["I
p] Prof. Dr. E. Wunsch, Dr. E. Jaeger
Priv.-Doz. Dr. R. Mengel, H. Wiedner Fachbereich Chemie der Universitat Postfach 7733, D-7750 Konstanz (Germany) [**] This work was supported by the Deutsche Forschungsgerneinschaft and represents part of the Doctoral Dissertation by H . Wiedner (Universitat Konstanz, 1976). Anyew Chem. J a r . Ed. Engl. 16 11977) No. 5
Max-Planck-Institut fur Biochemie, Abteilung fur Peptidchemie Schillerstrasse 42, 0-8000 Miinchen 2 (Germany) Dr. L. Kisfaludy, Dr. M. Low Forschungslaboratorium der Chemischen Werke Gedeon Richter AG H-1475 Budapest X, Pf. 27 (Hungary)
317
Facile Synthesis of Arabinoadenosine
3'-Phosphate[**I
By Rudolf Mengel and Harald Wiedner"] Only few examples of oxirane ring opening by phosphate ions are known in nucleoside chemistry"]. We demonstrated that adenosine can be transformed into its lyxo-epoxide ( I )r21 and that this product can be opened with mineral acid or alkali halide in the presence of boron trifluoride-etherate to give predominantly 3'-substituted halon~cleosides'~~. We have now examined the cleavage of the oxirane ring of ( 1 ) with phosphoric acid in hexamethylphosphoric triamide. Fourteen hours' heating to 100°C affords arabinoadenosine 3'-phosphate ( 2 ) which can be precipitated as the barium salt. On electrophoresis[4a1and chromatography[4b1the salt behaves in a manner resembling adenosine 5'-monophosphate and is cleaved by alkaline phosphatase''"] to arabinoadenosine together with traces of xyloadenosine[4'l. After reprecipitation of the salt and treatment with phosphatase, xyloadenosine can no longer be detected. Treatment with 5'-nucleotida~e[~~] (18 h) has no effect on ( 2 ) , while 3'-nu~leotidase[''~cleaves the compound to the extent of 90% within 18 h. The 90 MHz 'H-NMR spectrum of (2) was measured against sodium 2,2-dimethyl-2-silapentanesulfonate as internal standard in D,O to which had been added one drop of trifluoroacetic acid (to shift the HOD signal away from the signals due to the sugar protons). It showed signals at
O=+O
Arabinoadenosine 3'-phosphate (2) could serve as an "arabino" building block in oligonucleotide synthesis and as starting material for the synthesis of arabino-cAMP[61.Compound (2) also warrants interest in connection with the fact that arabinoadenosine has an inhibitory action on DNA polymerase, ribonucleotide reductase, adenyl cyclase, various viruses, and various animal tumors.
Procedure A solution of adenosine lyxoepoxide ( I ) (100mg, 0.4 mmol) and phosphoric acid (crystalline, 98 %; 200mg, 1.7mmol) in hexamethylphosphoric triamide (2 ml) is stirred for 14h at 100°C. After neutralization with conc. ammonia most of the solvent is removed in uacuo at 100°C. After addition of 10 % barium acetate solution (6 ml), triethylamine (0.4ml), and ethanol (12ml) a precipitate is formed. The mixture is allowed to stand for 12h at 4°C and then centrifuged; the residue is washed with ethanol ( 3 x IOml), and then with water (7 x 10ml). The combined aqueous extracts are evaporated to IOml, treated with ethanol (10ml), and left to stand for 10h at 4°C. The precipitate formed is filtered off and applied to a Sephadex DEAE A 25 column (2.3cm x 35 cm). Elution is performed with a gradient of triethylammonium hydrogen carbonate (0.05 to 0.2 M ; 2 liter); 25-ml fractions are collected. Fractions 33 to 47 contain 0.153 mmol of (2) (yield 38%). They are concentrated and freeze dried. Received: February 17, 1977 [Z 678 IE] German version: Angew. Chem. 89, 328 (1977) CAS Registry numbers: ( l ) , 40110-98-3: (Z), 54621-41-9
W E . Harue!, J . J . Michalsky, A . R . Todd, J. Chem. Soc. 1951, 2271: K . Tadashi, A. Mituse, J . Motonobu, N . Tokrrro, Japan. Patent Kokai 74, 18879 (CI. 16E 461) 1974: Chem. Abstr. 80, 121290e (1974). M. J . Robins, Y Fonron, R. Mengel, J. Org. Chem. 39, 1564 (1974). R . Mengel, H . Wiedner, Chem. Ber. 109, 1395 (1976). a) Triethylammonium hydrogen carbonate, pH =7.5, IOOOV: b) o n PEI cellulose with I N NHaOAc/ethanol ( I : I ) ; c) xyloadenosine and arabinoadenosine can be separated by electrophoresis in borate buffer (pH = 9) at 1000 V. a) Boehringer (Mannheim); b) [E.C. 3.1.3.51, Sigma Comp.; c) [E.C. 3.1.3.61, Sigma Comp. 7: A. Khwaja, R . Harris, R . K . Robins, Tetrahedron Lett. 1972, 4681, and references cited therein.-Cf. the analogous synthesis of 9-p-D-arabinofuranosylcytosine 3',5'-cyclophosphate: R. A . Long, G . L . Szekeres, T: A . Kliwaja, R . W. Sidwell, L. N . Simon, R . K. Robins, J. Med. Chem. 1 5 , 1215 (1972).
Side Reactions in Peptide Synthesis: tert-Butylation of Tryptophan By Erich Wiinsch, Ernst Jaeger, Lajos Kisfaludy, and Miklos LOW[*]
6=8.6 (s, 1, Hs), 8.46 (s, 1, HZ), 6.5 (d, Jt,-2,=5.2HZ, 1, HI,),4.86 (m, 1, H3,),4.74 (q, Jz,-3.=4Hz, 1 , H2,),4.29 (quart, 1, H4.), 3.95ppm ("d, 2H, Hs,,, H5.b).The chemical shift of the H atom attached to C-3' (shifted downfield by 0.5 ppm relative to adenosine) and the coupling constant between 31P and 3'-H, show the phosphate group to be located in position 3'.
Indoie and its derivatives are known to be very reactive. The amino acid tryptophan contains an indole ring as a third function, thus incorporating a relatively reactive moiety into the peptide chain. Side reactions due to indole have been described and postulated in peptide chemistry, e. g . also under conditions of acidolytic removal of protective groups from
["I
p] Prof. Dr. E. Wunsch, Dr. E. Jaeger
Priv.-Doz. Dr. R. Mengel, H. Wiedner Fachbereich Chemie der Universitat Postfach 7733, D-7750 Konstanz (Germany) [**] This work was supported by the Deutsche Forschungsgerneinschaft and represents part of the Doctoral Dissertation by H . Wiedner (Universitat Konstanz, 1976). Anyew Chem. J a r . Ed. Engl. 16 11977) No. 5
Max-Planck-Institut fur Biochemie, Abteilung fur Peptidchemie Schillerstrasse 42, 0-8000 Miinchen 2 (Germany) Dr. L. Kisfaludy, Dr. M. Low Forschungslaboratorium der Chemischen Werke Gedeon Richter AG H-1475 Budapest X, Pf. 27 (Hungary)
317
