Pergamon 0031-9422(94)00825--6 Phytochemistry, Vol. 38. No. 6, pp. 1547 1550. 1995 Elsevier Science Ltd Printed in Great Britain 0031 9422/95 $9.50 + 0.00 TWO STEROIDAL ALKALOIDS FROM VERATRUM VIRIDE KHALID A. EL SAYED,* JAMES D. MCCHESNEY,t AHMED F. HALIM, AHMED M. ZAGHLOUL and MARKUS VOEHL Pharmacognosy Department, Faculty of Pharmacy, Mansoura University, Mansoura 35511, Egypt; fThe Research Insti Pharmaceutical Sciences and Department of Pharmacognosy, School of Pharmacy, University of Mississippi, MS 38 +Department of Chemistry, Vanderbilt University, Nashville, TN 37235, U.S.A. (Received in revised form 14 September 1994) Key Word lndex--Veratrum viride; Liliaceae; roots; rhizomes; steroidal alkaloids; rubivirine; veramivirine. Abstraet--A phytochemical study of the roots and rhizomes of Veratrum viride led to the isolation of the two ne steroidal alkaloids, rubivirine, identified as 12fl-hydroxyisorubijervine, and veramivirine, identified as 12~-hy- droxyveramiline. Structural elucidation of the two compounds was aided by 2D-NMR spectral analyses. INTRODUCTION The roots and rhizomes of Veratrum viride have been extensively studied for their hypotensive [1] and cytotoxic [2] activities and in the therapy of myasthenia gravis [3]. About 25 steroidal alkaloids have been iso- lated from this species so far [4]. The isolation of the known alkaloids, veramarine, zygadenine, angeloyl- zygadenine and 15-O-2-methylbutyroylgermine, as well as l~,3]~-dihydroxy 5g-jervanin-12-en-11-one, a new jer- vanine alkaloid, has been reported [5]. Further phytochemical work on this species has led to the isola- tion of two new steroidal alkaloids, rubivirine (1) and veramivirine (2). 19 11 9-1 OH3 It8 # Compound R1 R2 1 Rubivirine [5-OH CH2OH 3 Rubijervine a-OH CH3 4 l Isorubijervine H CH2OH 5 Epirubijervine ffOH CH3 6 Solanidine In CH3 RESULTS AND DISCUSSION The alkaloid-containing fractions were separated by a combination of flash chromatography on silica gel, preparative TLC and fractional recrystallization to give rubivirine (1) and veramivirine (2). Both compounds gave positive Dragendorff, iodoplatinate and Mayer tests, in- dicating their alkaloidal nature. They also gave positive Liebermann-Burchard and Salkowski tests, indicating their steroidal nature. Rubivirine (1) was recrystallized from methanol to give needles. Thermospray-LC-MS analysis displayed a [M + HI + at m/z 430 suggesting the molecular formula C27H43NO3. The methyl singlet which absorbed at 60.92 in the 1HNMR (Table 1) is correlated with the methyl carbon at 6 19.2 and was assigned to the 19- methyl group. On the other hand, the two methyl doub- lets at 6 0.87 (J = 6.2 Hz) and 1.06 (J = 7.4 Hz) are corre- *Author to whom correspondence should be addressed. lated with the two methyl carbons at 6 18.9 and 19.4, and ascribed to the 21- and 27-methyl groups. These assign- ments revealed the 22,26-epiminocholestane skeleton of this alkaloid [6]. The ~t-axial proton H-12 resonating at 6 3.30 as a d d (J = 7.7 and 4.3 Hz) is correlated with a methine carbon at 669.0 and assigned to C-12 on the basis of HETCOR. The proton signals, which reson- ated at 63.97 (1H, d, J = 1 2 . 2 H z ) and 3.51 (1H, d, J = 12.0Hz), and correlatedwith the oxygenated methylene carbon at 62.4, were assigned to a C-18 hy- droxymethyl group. Confirmation of the position of the hydroxymethyl group was achieved by comparison of the 13CNMR spectra (Table 2) of 1 with that reported for solanidine (6), which is 12,18-dideoxyrubivirine [7]. The hydroxyl group substitution at C-18 caused significant shifts at C-13 ( + 4.9), C-17 ( - 5 . 9 ) as well as C-18 ( + 45.4). It also caused a significant shift at C-12 ( - 2.9) as compared with rubijervine (3), which is 12~-hy- droxysolanidine [5]. The equatorial proton H-26 ab- sorbed at 62.91 as a d d (J = 9.1 and 2 Hz) and is gem- inally coupled to the axial H-26 proton which resonated 1547