Journal of Natural Pmdwtr Vol. 5 3 , No. 2 , p p . 325-332, Mar-Apr 1990
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CHEMOTAXONOMY OF FERNS, 3. TRITERPENOIDS FROM POLYPOD1UM POL YPODlOlDES HIROWKIAGETA+and YOKOARAI S h a a Colkge o j P h a m e u t i r a l Sciences, Machiah, Tokyo 194,Japan AesTucr.-From the fresh rhizomes of Polypodiumpolypodioidesvarious triterpenoids belonging to the hopane, serratane, cycloartane, malabaricane, and polypodane groups were isolated. A new triterpenoid acetal, onon acetal 1141,was also isolated, and its structure was characterized based on its physical data. Polypodiumpolypodioides (the type species of the genus Marginaria) was found to be chemically very similar to Polypodium vulgare (the type species of Polypodium). Thus, the genus name Marginaria should be discontinued.
Polypodium polypodioides (L.) Watt. is distributed in North America and is usually found growing on large oak trees. It is called the “resurrection fern” because the fronds curl up when dry but revive after a rain. This fern was named Marginaria ceteracina by Bory in 1824 (1,2), and designated as the type species of the genus Marginaria, which included many Polypodiaceous species growing in Central and South America (3). As previously reported (4,5), some Japanese Polypodium species, such as Polypodium niponicum Mett. and P . f m o s a n u m Baker, contain very characteristic triterpenoid constituents; this information was used to separate these species from the genus Polypodium. Because these Japanese ferns were once described as Marginaria niponica (Mett .) Nakai and Marginaria f m o s a n a (Baker) Nakai (6), we decided to examine the chemical constituents of the type species ofMarginaria. This paper therefore deals with the chemical studies on the n-hexane extract of the fresh rhizomes of P . polypodioides, the type species. Besides various kinds of known triterpenoids, including those characteristic of Polypodium species, a new hopane acetal, named orton acetal [147, and its derivative were isolated and their structures characterized. RESULTS The fresh rhizomes of P . polypodioides, collected in Wilmington, North Carolina, were extracted twice with n-hexane to give an oily extract. Inasmuch as the tlc pattern of this extract indicated triterpenoid hydrocarbons, esters, alcohols, and sterols, the extract was chromatographed on Si gel and yielded seven fractions: triterpenoid hydrocarbons (fraction I), sterol esters (fraction II), cycloartanoid esters (fraction III), cycloartanoid acetates (fraction IV), an acetal, a ketone, and alcohols (fraction V), sterols (fraction VI), and hemiacetal (fraction VII). The constituents of each fraction were identified as follows.
TRITERPENOID HYDROCARBONS.-Fraction I was separated by AgNO, si gel cc to give three white crystal triterpenoid hydrocarbons, which were identified as hop17(2 1)-ene [l} ( 4 ) ,serrat- 14-ene [2} (7,8) and hop-22(29)-ene [37 (4).The occurrence of 1among ferns is not common, while 3 was found in almost all kinds of ferns. Compound 2 is a very characteristic component of Polypodium, such as Polypodium vulgare L. (the type species of this genus), Polypodium fauriei Christ, and Polypodium boreale Haufler (=Polypodium virginianum L.) of Japan (9). More polar oily triterpenoid hydrocarbons were also detected from the later eluted portion of fraction I and were identified as the bicyclic a-polypodatetraene [4}(10)and y-polypodatetraene 151 (lo), the tricyclic 13PH-malabaricatriene 167 (11) and the linear squalene [7(10) by comparison of the gc and ms data with those of authentic samples.
Wol. 53, No. 2
Journal of Natural Products
326
fl \
\
7
12 R=Ac
RO
dP+ 11 8 R R=Ac =H
13 R=Ac
STEROLESTERS.--The waxy ester, fraction 11, was hydrolyzed with 5% KOH in EtOH followed by workup to give three alcohols, which were identified as sitosterol (€lo%),stigmasterol (13%), and campesterol (7%) by gc-ms. The acidic fraction was also identified as palmitic acid (>go%) by the gc-ms of its methyl ester. Thus, this fraction was confirmed to be a mixture of the three sterol palmitates. CYCLOARTANOID ESTERS.-A mixture of oily esters was separated by chromatography from fraction 111and suggested to be fatty acid esters of cycloartanoid triterpenes. The 'H-nmr spectrum of the mixture exhibited the proton signals of the cyclopropane methylene of 3 1-norcycloartane and cycloartane 18 0.149 and 0.401 ( d , J = 4 . 1 Hz),
Mar-Apr 19901
Ageta and Arai:
eH
Triterpenoids from Polypodium
327
OH
OMe
14
..'
0.341 and 0.576 (d, 4.1 Hz), respectively], terminal methylenes (6 4.663, 4.667, 4.7 16), olefinic methyl (6 1.603)and low field doublet methyls {6 1.026, 1.031 ( J = 6.8 Hz)] with several methyl singlets and aliphatic proton signals. The proton signals of the fatty acid parts were also observed at 6 1.303, 1.3 12, 2.035, 2.293, 2.770, and 5.34 1. The alcohols obtained by hydrolysis of the ester mixture gave three spots on tlc and separated into three components, 3 I-norcyclolaudenol (81 (12,13), a mixture of cycloartenol 191 (12,13) and cyclolaudenol 1101 (5), and a sterol mixture. These were identified by comparison of the gc-ms data with those of authentic samples. Linoleic acid was also identified from the acidic part by comparison of the 'H-nmr and ms data of its methyl ester with an commercial sample. Thus, this fraction was established to be a mixture of the three cycloartanoid alcohol linoleates accompanied by sterol linoleates. CYCLOARTANOID AcETATES.-Although fraction Iv gave one spot on the tlc, it was found to be a mixture of cycloartanoid triterpene acetates. The 'H-nmr spectrum showed very similar proton signals to those of linoleic acid esters: 6 0.15 1and 0.400 (d, J = 4 . 1 Hz), 0.341 and 0.575 (d, 4.1), 4.665, 4.714, 1.605, 1.025 (d, 6.8), and 1.030 (d, 6.8). The gc of the mixture exhibited three main peaks at re1 Rt 3.94,4.05, and 4.52 with five additional small peaks. The three peaks were identified as 3 l-norcyclolaudenyl acetate E l l ] (5), cycloartenyl acetate {12] (12,13), and cyclolaudenyl acetate 1131(5) by comparison of the re1 Rt and the typical fragment ion peaks ofauthentic cycloartanoid acetates. ACETAL.
