Guanosine VOLUME

A solution of the monoacetyl compound (0.454 g, 1.4 mmol) and NaNOj (0.8 g, 11.6 mmol) in hot HjO (2.5 mL) was rapidly cooled to rt. Glacial HOAc (0.8 mL) was added and the mixture shaken until a clear Solution was obtained. After 1 h, an equal volume of H O was added the mixture was then kept overnight at rt. After evaporation, the residue was refluxed with a solution of N.i (0.1 g, 2.5 mmol) in McOH (15 mL) HjO was then added until a clear solution was obtained and the mixture was refluxed for 45 min. After neutralization with HOAc, the solution was evaporated to dryness and a solution of the residue in HjO (25 mL) was seeded with neutral guanosinc upon which crystallization occurred rapidly yield 0.28 g (71 %). Recrystallization (H,0, 7 mL) gave 0.223 g of product which charred without melting at 235 "C alone or in admixture with natural guanosine [a] —ll(c = 1.4%, 0.1 M NaOH). A sample of anhyd natural guanosine had [x] — 70 (c = 1.5%, 0.1 M NaOH). 

To a solution of 2 -deoxy-0 -(pentanuorophenyl)-Ar -(trifiuoroacetyl)guanosine (11 0.53 g, 1 mmol) in MeOH (20 mL) was added NaOMe (0.54 g, 10 mmol). The mixture was heated at 55 °C for 36 h and concentrated to dryness. The residue was dissolved in HjO and neutralized by addition of HOAc. The mixture was concentrated to a small volume and purified by reverse-phase HPLC (gradient of 2-lOVu MeCN in HjO over 30 min at a flow rate of 16 mLmin ). Concentration of appropriate fractions gave the product yield 0.24g (85%). 

Fig. 3-93. Separation of nucleosides using anion exchange chromatography. — Separator columns 2 IonPac AS4A eluent see Fig. 3-92 flow rate 1.5 mL/min detection suppressed conductivity injection volume 50 pL solute concentrations 10 ppm cytidine (1), 5 ppm adenosine (2), 10 ppm thymidine (3) and uridine (4), 13 ppm 2 -deoxyguanosine (5), 15 ppm guanosine (6), and inos-ine (7).

M FIGURE 3-5 Various graphic representations of the structure of Ras, a monomeric guanine nucleotide-binding protein. The inactive, guanosine diphosphate (GDP)-bound form is shown in all four panels, with GDP always depicted in blue spacefill, (a) The C backbone trace demonstrates how the polypeptide is packed into the smallest possible volume. 

Prepare serial dilutions in the range of 1 pAf to 10 mAf of cGMP, GTP, GDP, OMP, guanosine, cAMP, ATP, ADP, AMP, and adenosine in sucrose solution Mix equal volumes of gelatin solution (see Note 11) with the solutions prepared in step 1 and keep warm (approx 50°C). 

---