Henk 't Hart

Alkaloids of some Asian Sedum species

Abstract The leafy parts of 16 Asian species belonging to the three sections of Sedum were investigated for the presence of alkaloids. Only in seven species of Sedum sect. Sedum were alkaloids found. Sedum bulbiferum, S. japonicum, S. lepidopodium, S. morrisonensis, S. oryzifolium, S. polytrichoides and S. sarmentosum contain 14 pyrrolidine and piperidine alkaloids. Sedum oryzifolium differs significantly from the other species. It contains the typical S. acre alkaloids, sedamine, allosedamine and sedinone, as well as a series of monosubstituted piperidines and pyrrolidines. In addition, two new pyrrolidine alkaloids were detected in S. oryzifolium. i.e. both diastereoisomers of 1-phenyl-2-(2-N-methylpyrrolidyl)-ethanol for which the names pyrrolsedamine and pyrrolallosedamine are proposed (to express their structural relationship with sedamine and allosedamine). In five species of Sedum sect. Aizoon and four species of S. sect. Telephium, no alkaloids could be detected. The results support the hypothesis that in the family Crassulaceae, alkaloids are restricted to species of the ‘Acre’ lineage.

Sedum surculosum andS. jaccardianum (Crassulaceae) share a unique 70 bp deletion in the chloroplast DNA trnL (UAA)-trnF (GAA) intergenic spacer

TrnL (UAA)-trnF (GAA) chloroplast DNA spacer sequences of three species ofMonanthes, Sedum surculosum (=Monanthes atlanticum) andS. jaccardianum were compared.S. surculosum, the systematic position of which has been disputed ever since its discovery, shares a phylogenetically highly significant 70 bp deletion withS. jaccardianum. In addition to this large deletion the two Moroccan species ofS. ser.Monanthoidea differ in three more indels as well as in four nucleotide substitutions from the species ofMonanthes. These data render strong support for the monophyly ofS. ser.Monanthoidea andMonanthes. Spacer length in seven species and one subspecies ofMonanthes is relatively uniform.

ALKALOIDS OF SOME EUROPEAN AND MACARONESIAN SEDOIDEAE AND SEMPERVIVOIDEAE (CRASSULACEAE)

Some 22 pyrrolidine and piperdine alkaloids were detected in the leafy parts of Sedum acre, S. aetnense, S. anglicum, S. brissemoreti, S. farinosum, S. fusiforme, S. lancerottense, S. melanantherum, and S. nudum. In addition to the alkaloids known from S. acre, 1-(2-pyrrolidyl)-propan-2-one and 2-monosubstituted piperidine alkaloids bearing butan-2-one, butan-2-ol, pentan-2-one and pentan-2-ol sidechains were identified. Phenylethylamine was isolated from the vegetative parts of S. album. In S. lydium, S. meyeri-johannis, and 16 species of S. series Rupestria, Aeonium, Greenovia, Jovibarba and Sempervivum no alkaloids could be detected. The results indicate a correlation between the presence of alkaloids and the major evolutionary trends in the European and Macaronesian Crassulaceae.

The systematic and evolutionary significance of exudate flavonoids in Aeonium

Leaf exudates of 32 species of Aeonium were examined for the presence of flavonoids. Thirty two flavonoids were detected in exudates of half of the species. The flavonoids were identified as methyl ethers of kaempferol, 6-hydroxykaempferol, quercetin, myricetin and scutellarein. The distribution of the exudate flavonoids is mostly in agreement with infrageneric sectional classifications based on morphological and molecular characters. The combination of kaempferol 3,7-dimethyl ether and quercetin 3,7-dimethyl ether is characteristic of section Aeonium. Myricetin methyl ethers are restricted to section Goochia and section Petrothamnium. Section Leuconium, section Canariense and section Patinaria are particularly poor in exudate flavonoids. 6-Hydroxyflavonoids occur in leaf waxes of six species belonging to sections Goochia, Canariense, Chrysocome and Pittonium, probably as a result of inheritance of this feature from a common ancestor. The presence of exudate flavonoids shows weak correlations with habitat and the presence of glandular hairs.

Evolutionary Aspects of Crassulacean Acid Metabolism in the Crassulaceae

The CAM pathway is taxonomically widespread and is believed to be used by about 16 000 species of vascular plants (see p. 9), a figure that includes more than 6% of all angiosperms. In addition, CAM has been reported in certain advanced groups of lower vascular plants (Isoetes; see Chap. 18) and in the specialized gymnosperm Welxvitschia mirabilis (Schulze et al. 1976; Winter 1985).

EPICUTICULAR WAXES OF SEDUM SERIES RUPESTRIA

Epicuticular waxes from 55 plants of the seven species of Sedum series Rupestria and four artificial hybrids have been examined by GC and GC-MS. The taxa were S. amplexicaule, S. forsterianum, S. montanum ssp. montanum, S. montanum ssp. orientale, S. ochroleucum, S. pruinatum, S. rupestre ssp. erectum, S. rupestre ssp. rupestre, S. sediforme, and four hybrids between S. forsterianum and S. rupestre ssp. erectum. Alkanes, alcohols, aldehydes, triterpenoids and wax esters were the main wax components. Comparison of wax composition and wax ultrastructure of glaucous and glossy plants led to the conclusion that triterpenoids are primarily responsible for glaucousness. Some 14 triterpenes were detected, the major being germanicyl formate, fern-8-en-3-yl formate, taraxerone and taraxeryl acetate. Fern-8-en-3-yl formate was isolated from S. forsterianum. The distribution of triterpenes generally supported the phylogeny of the species based on a combination of biosystematic and molecular data, but failed to provide additional evidence for the presumed allopolyploid origin of S. rupestre ssp. rupestre from S. forsterianum and S. rupestre ssp. erectum.

The evolution of crassulacean acid metabolism inAeonium inferred from carbon isotope composition and enzyme activities

SummaryVariation in crassulacean acid metabolism (CAM) was investigated in thirtyAeonium species under controlled, well-watered conditions. Carbon isotope ratios and the specific activities of the CAM enzymes phosphoenolpyruvate carboxylase (PEPC) and malic enzyme ME_ are highly correlated and indicate a large variation in CAM. The δ13C values range from -13.6 to -27.1‰; PEPC and ME activities vary seven- and five-fold, respectively. Since these observations were made under controlled conditions, the differences are genetically determined and represent adaptations to different habitats. The CAM activity of the species is correlated with the characteristics of their natural habitats. CAM activity is also highly correlated with the growth- and life-forms of the species and accordingly agrees well with the infrageneric classifications which are partly based on these characters. Our results support the hypothesis that the Sempervivoideae have evolved from an EurasiaticSedum, which used C3 photosynthesis or weak CAM, and that evolution withinAeonium was from undifferentiatedSedum-like dwarf shrubs with C3 photosynthesis to woody candelabrum-shaped shrubs exhibiting strong CAM. The monocarpic, rosulate species with intermediate CAM may have evolved via an independent secondary route.

Variation in Crassulacean acid metabolism within the genus Sedum : carbon isotope composition and water status dependent phosphoenolpyruvate carboxylase activity

Summary Variation in Crassulacean acid metabolism (CAM) was examined in a large group of Sedum species, using two criteria: 13 C/ 12 C ratio and water status dependent specific activity of the CAM key enzyme phosphoenolpyruvate carboxylase (PEP-C). A large amount of variation in CAM was found within the genus Sedum . In three European species C3-like photosynthesis was found, as indicated by water status independent, low specific PEP-C activity and δ 13 C values of around -30‰. Three Mexican species showed water status independent high specific PEP-C activity and δ 13 C values of -15 to -22‰, indicating constitutive CAM. In the majority of European species the specific PEP-C activity was water status dependent; δ 13 C values ranged from -25 to -31‰. Specific PEP-C activity and carbon isotope ratios are significantly correlated, species with high specific activities showing less negative δ 13 C values. The Michaelis constant for PEP of the PEP-C enzymes of these Sedum species was determined. The observed values differed from 120-800 µmo1. The observed variation in CAM was compared with taxonomic relations of the examined species, leaf thickness, Km(PEP) and life cycle length.

Phosphoenolpyruvate carboxylase in Sedum rupestre (Crassulaceae) : drought-enhanced expression and purification

Summary Upon exposure to water stress, the specific activity of the key Crassulacean acid metabolism enzyme, phosphoenolpyruvate (PEP) carboxylase, increased several-fold in Sedum rupestre L. Photoperiodic long days were without effect. PEP carboxylase was purified from drought-stressed S. rupestre. The enzyme had an apparent molecular weight of 400,000 and was composed of four 100 kilodalton subunits. It had a Michaelis constant of 200 micromolar for PEP and was inhibited by malate with a Ki of 700 micromolar. During drought stress the apparent affinity for PEP did not change. Monospecific polyclonal antibodies were raised against the purified protein. Immunoblot analysis after different periods of drought stress showed that the increase in PEP carboxylase activity was the result of de novo synthesis of PEP carboxylase protein.

Seasonal Variation of Phosphoenolpyruvate Carboxylase Specific Activity in Fifteen Species Exhibiting Facultative or Obligate Crassulacean Acid Metabolism

Summary For 16 months the specific activity of the CAM key-enzyme PEP carboxylase (PEP-C) was determined monthly in twelve Sedum species previously shown to exhibit facultative or obligate CAM, as well as in Aeonium castello-paivae, Sempervivum nevadense and Kalanchoe daigremontiana . In all species examined the PEP-C activity showed a seasonal fluctuation with maxima during the summer from 8- to 30-fold higher than winter values. For most species the seasonal fluctuation correlated significantly with temperature and, to a lesser extent, with irradiation; no correlation was present with precipitation in garden-grown plants. Plants grown in greenhouse and garden showed corresponding seasonal fluctuations. Specific PEP-C activity, however, was restored to the summer level upon transfer to a growth chamber (21/15 °C, LD) in October. To determine the controlling climatic factor for this growth room-induced increase in PEP-C activity, the influence of light intensity, photoperiod and temperature on PEP-C activity was investigated in Sedum rupestre. Daytime temperature appeared to be the most influential factor, whereas light intensity had a small but significant effect.