Combination of Pretreatments with Acetic Acid and Sodium Methoxide for Efficient Digoxin Preparation from Digitalis Glycosides in Digitalis lanata Leaves

We previously developed an HPLC method for determination of lanatoside C, digoxin and α-acetyldigoxin in digitalis glycosides isolated from Digitalis lanata leaves. Here, we present an improved HPLC-UV method to determine those compounds and deslanoside. We used the improved method to examine the effects of various pretreatments on the amounts of the four compounds isolated from the leaves, with the aim of maximizing the yield of digoxin. Leaves were extracted with 50% methanol, followed by clean-up on a Sep-Pak C18 cartridge prior to HPLC analysis. The amounts of lanatoside C, digoxin and α-acetyldigoxin per 100 mg of the leaves without pretreatment were 115.6, 7.45 and 23.8 μg, respectively (deslanoside was not detected). Pretreatment with acetic acid, which activated deglucosylation mediated by digilanidase present in the leaves, increased the amounts of digoxin and α-acetyldigoxin, while lanatoside C and deslanoside were not detected. Pretreatment with sodium methoxide, which hydrolyzed lanatoside C to deslanoside, increased the yields of deslanoside and digoxin, while lanatoside C and α-acetyldigoxin were not detected. The combination of both pretreatments afforded only digoxin in a yield of 115.1 μg/100 mg leaves. Use of the combined pretreatments appears to be effective for maximizing the yield of digoxin from the leaves.