Scheme 2. Synthesis of dehydrated madecassic acid conjugated with silybin. Reagents and conditions: a) Pyridine, SOCl2, pyridine, 2 h, rt, 47%. B) i) oxalyl chloride, DCM, rt, 14 h, ii) amino acid (glycine; or β-alanine), TEA, DCM, rt, 18 h; iii) Silybin, dicyclohexylcarbodiimide (DCC), DMAP, THF, 0 o C, rt, 18 h; c) Ac2O, pyridine, rt, 12 h. Conjugation of madecassic acid to silybin via an amino acid as a spacer (glycine, β-alanine, or 11-aminoundecanoic acid, 11-AUDA), was carried out in a two-step reaction sequence. Treatment of triacetyl madecassic acid with oxalyl chloride to furnish the intermediate acyl chloride acid was followed by reaction with one of the three amino acids, and then subsequent coupling with silybin 1 under Steglich conditions, using DCC/DMAP in dry tetrahydrofuran (THF) at room temperature. The corresponding conjugates 8-10 were obtained in overall yields of 20-26%. Analysis of NMR spectroscopy of these conjugates 8-10 confirmed the regioselective esterification to the 3-OH group of silybin moiety. A comparison of the 1 H NMR spectra of silybin ( ) with its conjugated ester exhibited distinctly that the chemical shift of proton H-3 in silybin moiety was downfield shifted from a range of 4.65-4.64 up to 5.82-5.84 ppm. Only a few silybin ester derivatives in positions C-3 have been reported so far. Antoszczak et al. reported the synthesis of conjugates of silybin with the antibiotics salinomucin and monensin. The authors obtained the conjugates through an ester linkage at the 23-OH group with the yields of 43% and 35%, respectively and no conjugates at the 3-OH group of silybin . However, our results are in good agreement with the results by Křen who reported a selective attachment on the 3-OH group of silybin moiety under Steglich esterification conditions to form the 3-O-galloylsilybin. Acetylation of compounds 6, 8-10 with acetic anhydride in pyridine at room temperature provided the corresponding acetylated products . Their structures were evidenced by the analysis of the NMR and MS spectroscopic data.