Phytochemistry, Vol. 30, No. 2, pp.495496, 1991 Printed in Great Britain. zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA p-HYDROXYNONANOPHENONE IN CELL SUSPENSION OF A zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA HUMULUS LUPULUS CULTIVAR 0031~9422/91 $3.00+0.00 0 1991 Pergamon Pnss plc CULTURES AMITABH CHANDRA, CARLOS R. LANGEZAAL and JOHANNES J. C. SCHEFFER* Division of Pharmacognosy, Center for Bio-Pharmaceutical Sciences, Gorlaeus Laboratories, P.O. Box 9502, 2300 RA Leiden, The Netherlands (Received 21 May 1990) zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQP Key Word Idex-Ifumulus lupulus; Cannabaceae; hop; cell suspension culture; secondary metabolite; aryl alkyl ketone; p-hydroxynonanophenone. zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA Abstract-A new aryl alkyl ketone was isolated from a cell suspension culture of zyxwvutsrqponmlkjihgfedcbaZYXWVUTSR Humulus lupulus cv Wye Northdown. Its structure is established as 1-phenyl+l’-hydroxy)-n-nonanone, using spectroscopic methods. INTRODUCTION Cell cultures of Humulus lupulus have been reported to produce some steroids [l], but a biotechnological ap- proach to induce such cultures to produce the bitter principles and the essential oil components present in hops have not yet materialized [2, 31. Therefore, we started a phytochemical investigation of cell suspension cultures of H. lupulus cv Wye Northdown, to look for the possible precursors of the hop flavour components, which are of great importance to the brewing industry [4]. We report on the isolation of l-phenyl+‘-hydroxy)-n- nonanone (1) from the chloroform extract of freeze-dried cultures, and on its structure determination. RESULTS AND DISCU!SSION Hop callus was induced from female leaf explants of H. lupwlus cv Wye Northdown, using Gamborg’s B5 medium [S]. The growth of the cell suspensions reached a stationary phase in ca three weeks. Subsequently the suspension cultures were freeze-dried and homogenized with chloroform to yield an extract, which after extensive chromatographic separations, afforded 1 as a new com- pound. 495 hydroxyphenone (R-CH,-CO-C,H,OI-I) system in 1[7, 83. Non-availability of further signals for oxygenated and unsaturated protons and carbons in the NMR spectra, the presence of an important mass fragment at m/z 177 [M-C4HJ+ and the McLafferty signal at m/z 135 [M -C7Hi5]+, suggested R to be an n-heptyl chain. Fur- thermore, the ‘HNMR signals at 63.24 (2H, brt, .I = 7.9 Hz, H-2), the overlapping multiplets at 6 1.25 (12H; H-3 to H-8), and the 13CNMR signals from 623.37 to 33.50 (C-3 to C-8) and at 615.0 (C-9 Me), together established the R-CH,CO- moiety in 1 to be n-non- anone with further substitution at C-l. Based on these data, the structure of 1 was established to be I-phenyl-(4’- hydroxy)-n-nonanone. The structure of 1 follows from its mass spectrum showing an [Ml+ peak at m/z 234, accounting for Cr5Hzz02. The presence of strong bands at 3360, 1675, 1610, 1510 and 1460 cm-’ in its IR spectrum, together with a peak at m/z 216 [M-HaO]+, indicated the presence of a phenolic and ketonic groups. The com- pound also gave a positive ferric chloride test. The ‘H NMR signals at 66.94 and 7.80 (each 2H, d, J = 8.5 Hz, H-3’ and H-S, H-2’ and H-6’), and the downfield singlet at 69.98, clearly indicated the phenyl ring to be p-hydroxy substituted. The 13C NMR signals at 6 191.0, 38.79, 129.0 and 132.0 (C-l, C-2, C-l’ and C-2’, respectively), combined with the characteristic mass peaks for phenolic and aromatic ketones [6] at m/z 216 [M-H20]+, 121 [C,H,OH-CO]+ and 93 CW-WHI+ clearly established the presence of a p- Secondary metabolites possessing an n-alkanone moiety as in the case of 1, have been detected in cell cultures of Ruta graueolens, which contained for example methyl nonyl ketone [9-123. Compound 1 cannot be related biosynthetically to either the bitter components or the essential oil components of hops, but the possibility of formation from shikimic acid via p-hydroxyaceto- phenone or pungenin aglycone, during the lignin bio- synthesis [13] appears to be more feasible. *Author to whom correspondence should be addressed. Leaf explants from female H. lupulus L. cv. Wye Northdown were surface sterilized. Callus was initiated and maintained on agar (2%) containing Gamborg’s B5 medium [5-J supplemented with 2% sucrose, 2 mg 1-r 2,4-D and 0.2 mg 1-l kinetin. Cell suspension cultures were induced from this callus using the same, liquid medium. They were maintained by subculturing every 14 days in 250 ml conical flasks containing 5Oml medium and agitated on a rotatory shaker (ca 120 strokes mitt-‘) at 25” with permanent light. “ad-l 1 EXPRRIMENTAL