THE SYNTHESIS OF 2'-DEOXY[6-15N]ADENOSINE HAS BEEN REPEATED FOR THIS FUNDING PERIOD BY MR. ASHBURN OF FURMAN UNIVERSITY. HE IS AN GRADUATE RESEARCH ASSISTANT WHO SPENT 6/93-7/94 WITH THE SIR RECEIVING TRAINING IN THIS SYNTHESIS. HE TOOK 5 GRAMS OF 6-CHLOROPURINE (PURCHASED FROM ALDRICH CHEMICAL CO.) WAS PLACED IN A STAINLESS STEEL BOMB AND THE CONTENTS WERE CHILLED AND THE VESSEL WAS EVACUATED. 2.0 EQUIVALENTS OF 99% 15NH3 WAS CYROPUMPED INTO THE VESSEL AND THE SYSTEM WAS THEN CLOSED. THE VESSEL WAS HEATED AT 150 OC OVERNIGHT. THE REACTION (SCHEME 1) WAS DETERMINED TO BE COMPLETE BY THIN LAYER CHROMATOGRAPHY (10 % MEOH/CH2CL2 V/V). THE [6-15N]ADENINE WAS ISOLATED IN GREATER THAN 85 % YIELD. THE EXCESS AMMONIA WAS ISOLATED AS THE AMMONIUM CHLORIDE. THE [6-15N]ADENINE WAS THEN CONVERTED TO [6-15N]-2'-DEOXYADENOSINE USING THE FOLLOWING METHOD. ADENINE HYDROCHLORIDE (2.5 G) WAS THEN DISSOLVED IN 360 ML OF 10 MM K2HPO4 AT ROOM TEMPERATURE. TO THIS WAS ADDED 25 G OF THYMIDINE, WITH STIRRING. THE PH WAS THE ADJUSTED TO 7.4 WITH 1 N KOH. TO THIS WAS ADDED 900 UNITS OF THYMIDINE PHOSPHORYLASE (SIGMA, T-7006) AND 62 MG OF PURINE NUCLEOSIDE PHOSPHORYLASE (SIGMA, N-3264, LOT 23H0543). THE REACTION WAS STIRRED AT ROOM TEMPERATURE FOR 3 DAYS. TLC ANALYSIS INDICATED AN INCOMPLETE REACTION. STIRRING WAS CONTINUED AND THE REACTION WAS BROUGHT TO 37 OC. AFTER 8 DAYS THE MIXTURE WAS FILTERED AND THE FILTRATE WAS APPLIED TO A BIO-RAD AG1X2 HYDROXIDE FORM RESIN, AND 2.35 G OF [6-15N]-2'-DEOXYADENOSINE WAS ISOLATED.
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