In this example, a coelenterazine having structure VIII (“dideoxycoelenterazine”) was synthesized from 2-amino-3-benzyl-5-phenylpyrazine and 2-acetoxy-3-phenylypropenal. The compound 2-amino-3-benzyl-5-phenylpyrazine was prepared according to previously described methods (Kishi, Y. et al, Tet. Lett. 2747 (1972); Cormier, M. et al, Biochemistry, 18(11), 2204 (1979); Hirano, T. et al Tetrahedron 53 (38) 12903-12916 (1997)). The compound 2-acetoxy-3-phenylypropenal was synthesized as follows. To a solution of phenylpyruvic acid (25 g, 152 mM) in dry pyridine (250 mL) was added acetic anhydride (170 mL). The solution was stirred overnight under an inert atmosphere and monitored by thin layer chromatography (TLC) using 10% methanol in CH2Cl2. The pyridine was evaporated under reduced pressure, and the resuling syrup was taken up in CH2Cl2 (700 mL) and washed 3 times with 200 mL portions of a 0.1N aqueous HCl solution. The organic phase was dried over Na2SO4 and filtered, and the filtrate was concentrated under reduced pressure to a viscous amber syrup. This syrup was purified by chromatography on N.P. silica with CH2Cl2 to give 24 g of 2-actoxy-3-phenylpropenoic acid. This enol acetate intermediate was dissolved in 150 mL THF and cooled to 0° C. Oxalyl chloride (51 mL, 580 mM) was then added dropwise with stirring, and the solution allowed to stir for an additional 20 minutes at 0° C. Dimethylformamide (DMF) (7.5 mL) was added dropwise, and this solution was then stirred for 4 hours at 0° C., concentrated under reduced pressure and co-evaporated twice with toluene to yield the acid chloride. This acid chloride intermediate (14.5 g, 87 mM) was dissolved in 200 mL of a 1:1 CH2Cl2/THF mixture. This solution was cooled to −70° C., and LiAl(OtBu)3H (152 mL, 152 mM) was added dropwise while maintaining the temperature of the reaction below −70° C. After addition of LiAl(OtBu)3H was complete, stirring was continued for 2 hours at −70° C. The cooling bath was removed, aqueous HCl (2N, 100 mL) was added and the reaction was allowed to warm to room temperature. The mixture was extracted into 500 mL ether, and the combined organic phases were dried over Na2SO4 and filtered. The filtrate was concentrated under reduced pressure to yield a viscous oil, which was purified by column chromatography using N.P. silica (250 g) and a step gradient of ethyl acetate in hexane. The desired compound was eluted in 10% ethyl acetate in heptane to yield 13.5 g of 2-acetoxy-3-phenylypropenal as an amber oil (80% yield, aldehyde singlet at 9 ppm in 1H NMR, ms.=189).
