Stereochemical aspects of the biosynthesis of the epimeric cyanogenic glucosides dhurrin and taxiphyllin.

Dhurrin, I ((S)-p-hydroxymandelonitrile-beta-D-glucopyranoside), and taxiphyllin, II (the (R) epimer), occur in the genera Sorghum and Taxus, respectively. Both derive biosynthetically from L-tyrosine via the hydroxylation of p-hydroxyphenylacetonitrile, III. (3R)- and (3S)-L-[3-3H1]tyrosine, prepared by enzymic hydroxylation of the corresponding phenylalanines, were fed separately to shoots from sorghum seedlings (Sorghum bicolor (Linn) Moench) and cuttings from Japanese Yew (Taxus cuspidata Sieb. and Zucc.) and the appropriate cyanogenic glycoside was isolated (I or II). The fraction of the 3H conserved in I and II was calculated from both parallel feeding and 3H:14C double labeling experiments. The results for II were the reverse of I. Both hydroxylations of III, which give rise to the enantiomeric products (S)- and (R)-p-hydroxymandelonitrile, occurred with retention of configuration. The retention mode is characteristic of hydroxylations at aliphatic carbons catalyzed by mixed function oxidases.     

In vitro biosynthesis of the cyanogenic glucoside taxiphyllin in Triglochin maritima

When l-tyrosine and NADPH were incubated with a microsomal fraction from Triglochin maritima seedlings, 4-hydroxymandelonitrile (measured as 4-hydroxybenzaldehyde and HCN) and 4-hydroxyphenylacetonitrile were synthesized. Taxiphyllin, a cyanogenic glucoside occurring in this plant, was formed when UDPglucose and a soluble protein fraction from T. maritima, were also included in the assay. The above glucosyl transferase activity also produced taxiphyllin when incubated with R,S-4-hydroxymandelonitrile and UDPglucose. No trace of dhurrin, the enantiomer of taxiphyllin, was detected. The highest microsomal activity was found in 5- to 6-day-old etiolated seedlings. It was absolutely necessary to remove all seed coats from the seedlings in order to obtain an active microsomal fraction.     

Frequency of cyanogenesis in tropical rainforests of far north Queensland, Australia

BACKGROUND AND AIMS: Plant cyanogenesis is the release of toxic cyanide from endogenous cyanide-containing compounds, typically cyanogenic glycosides. Despite a large body of phytochemical, taxonomic and ecological work on cyanogenic species, little is known of their frequency in natural plant communities. This study aimed to investigate the frequency of cyanogenesis in Australian tropical rainforests. Secondary aims were to quantify the cyanogenic glycoside content of tissues, to investigate intra-plant and intra-population variation in cyanogenic glycoside concentration and to appraise the potential chemotaxonomic significance of any findings in relation to the distribution of cyanogenesis in related taxa. METHODS: All species in six 200 m(2) plots at each of five sites across lowland, upland and highland tropical rainforest were screened for cyanogenesis using Feigl-Anger indicator papers. The concentrations of cyanogenic glycosides were accurately determined for all cyanogenic individuals. KEY RESULTS: Over 400 species from 87 plant families were screened. Overall, 18 species (4.5 %) were cyanogenic, accounting for 7.3 % of total stem basal area. Cyanogenesis has not previously been reported for 17 of the 18 species, 13 of which are endemic to Australia. Several species belong to plant families or orders in which cyanogenesis has been little reported, if at all (e.g. Elaeocarpaceae, Myrsinaceae, Araliaceae and Lamiaceae). A number of species contained concentrations of cyanogenic glycosides among the highest ever reported for mature leaves-up to 5.2 mg CN g(-1) d. wt, for example, in leaves of Elaeocarpus sericopetalus. There was significant variation in cyanogenic glycoside concentration within individuals; young leaves and reproductive tissues typically had higher cyanogen content. In addition, there was substantial variation in cyanogenic glycoside content within populations of single species. CONCLUSIONS: This study expands the limited knowledge of the frequency of cyanogenesis in natural plant communities, includes novel reports of cyanogenesis among a range of taxa and characterizes patterns in intra-plant and intra-population variation of cyanogensis.     

A galloylated cyanogenic glycoside from the Australian endemic rainforest tree Elaeocarpus sericopetalus (Elaeocarpaceae)

A cyanogenic glycoside - 6'-O-galloylsambunigrin - has been isolated from the foliage of the Australian tropical rainforest tree species Elaeocarpus sericopetalus F. Muell. (Elaeocarpaceae). This is the first formal characterisation of a cyanogenic constituent in the Elaeocarpaceae family, and only the second in the order Malvales. 6'-O-galloylsambunigrin was identified as the principal glycoside, accounting for 91% of total cyanogen in a leaf methanol extract. Preliminary analyses indicated that the remaining cyanogen content may comprise small quantities of sambunigrin, as well as di- and tri-gallates of sambunigrin. E. sericopetalus was found to have foliar concentrations of cyanogenic glycosides among the highest reported for tree leaves, up to 5.2 mg CN g(-1) dry wt.     

Cyclopentenylglycine,a precursor of deidaclin in Turnera ulmifolia

When [2-¹⁴C]cyclopentenylglycine was synthesized and fed to seedlings of Turnera ulmifolia, the label was incorporated into the nitrile group of the cyanogenic glycoside deidaclin. The amino acid cyclopentenylglycine was also found to occur naturally in Turnera ulmifolia. These findings indicate that cyclopentenyl cyanogenic glycosides are synthesized from the corresponding amino acids by the same pathway utilized in the biosynthesis of other cyanogenic glycosides.     

Cyanogenic Eucalyptus nobilis is polymorphic for both prunasin and specific beta-glucosidases

Cyanogenesis (i.e. the evolution of HCN from damaged plant tissue) requires the presence of two biochemical pathways, one controlling synthesis of the cyanogenic glycoside and the other controlling the production of a specific degradative beta-glucosidase. The sole cyanogenic glycoside in Eucalyptus nobilis was identified as prunasin (D-mandelonitrile beta-D-glucoside) using HPLC and GC-MS. Seedlings from three populations of E. nobilis were grown under controlled conditions and 38% were found to be acyanogenic, a proportion far greater than reported for any other cyanogenic eucalypt. A detailed study of the acyanogenic progeny from a single open-pollinated parent found that 23% lacked a cyanogenic beta-glucosidase, 32% lacked prunasin and 9% lacked both. Of the remaining seedlings initially identified as acyanogenics, 27% contained either trace amounts of beta-glucosidase or prunasin, while 9% contained trace amounts of both. Results support the hypothesis that the two components necessary for cyanogenesis are inherited independently. Trace amounts are likely to result from the presence of non-specific beta-glucosidases or the glycosylation of the cyanohydrin intermediate by non-specific UDP glycosyl transferases.     

Gynocardin from Baileyoxylon lanceolatum and a revision of cyanogenic glycosides in Achariaceae

The cyclopentenone cyanhydrin glycoside gynocardin was the only cyanogen isolated from foliage of monotypic Australian rainforest tree, Baileyoxylon lanceolatum (Achariaceae). The presence of cyanogenic compounds in plants can have considerable taxonomic utility. A review of previous reports of cyanogenesis in the recently revised Achariaceae revealed distinct taxonomic patterns as well as inconsistencies in the reporting of cyanogenic compounds. This variation appears to be due to tissue level localisation of cyanogenic compounds as well as discrepancies in results obtained from different detection methods. Recommendations are made for future investigations.     

Positive effects of cyanogenic glycosides in food plants on larval development of the common blue butterfly

Cyanogenesis is a widespread chemical defence mechanism in plants against herbivory. However, some specialised herbivores overcome this protection by different behavioural or metabolic mechanisms. In the present study, we investigated the effect of presence or absence of cyanogenic glycosides in birdsfoot trefoil (Lotus corniculatus, Fabaceae) on oviposition behaviour, larval preference, larval development, adult weight and nectar preference of the common blue butterfly (Polyommatus icarus, Lycaenidae). For oviposition behaviour there was a female-specific reaction to cyanogenic glycoside content; i.e. some females preferred to oviposit on cyanogenic over acyanogenic plants, while other females behaved in the opposite way. Freshly hatched larvae did not discriminate between the two plant morphs. Since the two plant morphs differed not only in their content of cyanogenic glycoside, but also in N and water content, we expected these differences to affect larval growth. Contrary to our expectations, larvae feeding on cyanogenic plants showed a faster development and stronger weight gain than larvae feeding on acyanogenic plants. Furthermore, female genotype affected development time, larval and pupal weight of the common blue butterfly. However, most effects detected in the larval phase disappeared for adult weight, indicating compensatory feeding of larvae. Adult butterflies reared on the two cyanogenic glycoside plant morphs did not differ in their nectar preference. But a gender-specific effect was found, where females preferred amino acid-rich nectar while males did not discriminate between the two nectar mimics. The presented results indicate that larvae of the common blue butterfly can metabolise the surplus of N in cyanogenic plants for growth. Additionally, the female-specific behaviour to oviposit preferably on cyanogenic or acyanogenic plant morphs and the female-genotype-specific responses in life history traits indicate the genetic flexibility of this butterfly species and its potential for local adaptation.     

Cyanohydrin glycosides of Passiflora: distribution pattern, a saturated cyclopentane derivative from P. guatemalensis, and formation of pseudocyanogenic alpha-hydroxyamides as isolation artefacts

Nineteen species of Passiflora (Passifloraceae) were examined for the presence of cyanogenic glycosides. Passibiflorin, a bisglycoside containing the 6-deoxy-beta-D-gulopyranosyl residue, was isolated from P. apetala, P. biflora, P. cuneata, P. indecora, P. murucuja and P. perfoliata. In some cases this glycoside co-occurs with simple beta-D-glucopyranosides: tetraphyllin A, deidaclin, tetraphyllin B, volkenin, epivolkenin and taraktophyllin. P. citrina contains passicapsin, a rare glycoside with the 2,6-dideoxy-beta-D-xylo-hexopyranosyl moiety, while P. herbertiana contains tetraphyllin A, deidaclin, epivolkenin and taraktophyllin, P. discophora tetraphyllin B and volkenin, and P. x violacea tetraphyllin B sulfate. The remaining species were noncyanogenic. The glycosides were identified by 1H and 13C NMR spectroscopy following isolation by reversed-phase preparative HPLC. From P. guatemalensis, a new glucoside named passiguatemalin was isolated and identified as a 1-(beta-D-glucopyranosyloxy)-2,3-dihydroxycyclopentane-1-carbonitrile. An isomeric glycoside was prepared by catalytic hydrogenation of gynocardin. alpha-Hydroxyamides corresponding to the cyanogenic glycosides were isolated from several Passiflora species. These alpha-hydroxyamides, presumably formed during processing of the plant material, behave as cyanogenic compounds when treated with commercial Helix pomatia crude enzyme preparation. Thus, the enzyme preparation appears to contain an amide dehydratase, which converts alpha-hydroxyamides to cyanohydrins that liberate cyanide; this finding is of interest in connection with analysis of plant tissues and extracts using Helix pomatia enzymes.     

Sambunigrin and cyanogenic variability in populations of Sambucus canadensis L. (Caprifoliaceae)

The presence or absence of cyanogenic glycosides was determined for individuals from nine populations of Sambucus canadensis L. (elderberry) of east-central Illinois. In most of the populations tested, all or most of the individuals did not produce these compounds, in one, all were cyanogenic, whereas in another population this trait was highly variable. Addition of the enzyme emulsin to negative tests did not result in any further release of cyanide. The glycoside responsible is (S)-sambunigrin.     

Short Report : The cyanogenic glycoside barterin from Barteria fistulosa is epitetraphyllin B

Reisolation of the cyanogenic glycoside of Barteria fistulosa demonstrates that the structure of barterin, previously reported from this plant, is identical with that of epitetraphyllin B.     

Developmental toxicity of the cyanogenic glycoside linamarin in the golden hamster

Cassava, a staple food in many tropical countries, has been suspected as a cause of human congenital defects. Ingestion of the material during pregnancy has been reported to induce limb defects, microcephaly, open eye, and growth retardation in rats. Linamarin is a natural cyanogenic glycoside that occurs in high concentrations in cassava. In the present study, pregnant hamsters received an oral dose of 70,100, 120 or 140 mg/kg linamarin or an equivalent volume of isotonic saline during the early primitive streak stage of gestation. A dose of 120 or 140 mg/kg of the glycoside was associated with an increased incidence of vertebral and rib anomalies as well as the production of encephaloceles in the offspring. These larger doses of linamarin also resulted in obvious maternal toxicity. Linamarin treatment had no effect on fetal body weight, ossification of fetal skeletons, embryonic mortality, or litter size. Although ingestion of the cyanogenic glycoside was associated with a significant teratogenic response, the effects occurred only at doses that elicited signs of maternal intoxication.     

Cyanogenic allosides and glucosides from Passiflora edulis and Carica papaya

Leaf and stem material of Passiflora edulis (Passifloraceae) contains the new cyanogenic glycosides (2R)-beta-D-allopyranosyloxy-2-phenylacetonitrile (1a) and (2S)-beta-D-allopyranosyloxy-2-phenylacetonitrile (1b), along with smaller amounts of (2R)-prunasin (2a), sambunigrin (2b), and the alloside of benzyl alcohol (4); the major cyanogens of the fruits are (2R)-prunasin (2a) and (2S)-sambunigrin (2b). The major cyanogenic glycoside of Carica papaya (Caricaceae) is 2a; only small amounts of 2b also are present. We were not able to confirm the presence of a cyclopentenoid cyanogenic glycoside, tetraphyllin B, in Carica papaya leaf and stem materials. In detailed 1H NMR studies of 1a/b and 2a/b, differences in higher order effects in glucosides and allosides proved to be valuable for assignment of structures in this series. The diagnostic chemical shifts of cyanogenic methine and anomeric protons in 1a/b are sensitive to anisotropic environmental effects. The assignment of C-2 stereochemistry of 1a/b was made in analogy to previous assignments in the glucoside series and was supported by GLC analysis of the TMS ethers.     

Constitutive polymorphic cyanogenesis in the Australian rainforest tree, Ryparosa kurrangii (Achariaceae)

Cyanogenesis, the liberation of volatile hydrogen cyanide from endogenous cyanide-containing compounds, is a proven plant defence mechanism and the particular cyanogens involved have taxonomic utility. The cyclopentenoncyanhydrin glycoside gynocardin was the only cyanogen isolated from foliar tissue of the rare Australian rainforest tree, Ryparosa kurrangii (Achariaceae). Mechanical damage simulating foliar herbivory did not induce a significant increase in the expression of cyanogenesis over a 24h period, indicating cyanogenic herbivore defence in R. kurrangii is constitutive. The cyanogenic potential of mature leaves was quantitatively polymorphic between trees in a natural population, ranging from 0.54 to 4.77 mg CN g(-1) dry wt leaf tissue.     

Cyanogenic glycosides: a case study for evolution and application of cytochromes P450

Cyanogenic glycosides are ancient biomolecules found in more than 2,650 higher plant species as well as in a few arthropod species. Cyanogenic glycosides are amino acid-derived β-glycosides of α-hydroxynitriles. In analogy to cyanogenic plants, cyanogenic arthropods may use cyanogenic glycosides as defence compounds. Many of these arthropod species have been shown to de novo synthesize cyanogenic glycosides by biochemical pathways that involve identical intermediates to those known from plants, while the ability to sequester cyanogenic glycosides appears to be restricted to Lepidopteran species. In plants, two atypical multifunctional cytochromes P450 and a soluble family 1 glycosyltransferase form a metabolon to facilitate channelling of the otherwise toxic and reactive intermediates to the end product in the pathway, the cyanogenic glycoside. The glucosinolate pathway present in Brassicales and the pathway for cyanoalk(en)yl glucoside synthesis such as rhodiocyanosides A and D in Lotus japonicus exemplify how cytochromes P450 in the course of evolution may be recruited for novel pathways. The use of metabolic engineering using cytochromes P450 involved in biosynthesis of cyanogenic glycosides allows for the generation of acyanogenic cassava plants or cyanogenic Arabidopsis thaliana plants as well as L. japonicus and A. thaliana plants with altered cyanogenic, cyanoalkenyl or glucosinolate profiles.     

Frequency and distribution of cyanogenic glycosides in Eucalyptus L'Hérit

In this study approximately 420 of the described species of Eucalyptus were examined for cyanogenesis. Our work has identified an additional 18 cyanogenic species, 12 from living tissues and a further six from herbarium samples. This brings the total of known cyanogenic species to 23, representing approximately 4% of the genus. The taxonomic distribution of the species within the genus is restricted to the subgenus Symphyomyrtus, with only two exceptions. Within Symphyomyrtus, the species are in three closely related sections. The cyanogenic glycoside was found to be predominantly prunasin (1) in the 11 species where this was examined. We conclude that cyanogenesis is plesiomorphic in Symphyomyrtus (i.e. a common basal trait) but has probably arisen independently in the other two subgenera, consistent with recent phylogenetic treatments of the genus. The results of this study have important implications for the selection of trees for plantations to support wildlife, and to preserve genetic diversity.     

Growth cost and ontogenetic expression patterns of defence in cyanogenic Eucalyptus spp.

Plant defences can incur allocation costs and such costs incurred early in ontogeny may result in opportunity costs with effects evident later in life. A unified understanding of the growth cost of defence requires the identification of plants with varying ontogenetic trajectories of preferably resource demanding defences and an appropriate measurement of the growth cost of these defences. To develop such tools, we first compared nitrogen-based chemical defence (cyanogenic glycosides) in juvenile and adult foliage of three species of Eucalyptus (Myrtaceae). We found marked differences between the species, with two having much lower concentrations of foliar cyanogenic glycosides in seedlings compared to adults. We next used seedlings of two species to measure the resource (nitrogen) and growth cost of deploying cyanogenic glycosides. We found evidence that for every 1.0 nitrogen invested in cyanogenic glycosides, 1.49 additional nitrogens were effectively added to the leaves. We also found that deployment of cyanogenic glycosides was associated with a reduction in net assimilation rate (NAR) at constant leaf nitrogen. We did not, however, detect an overall growth cost associated with cyanogenic glycoside deployment because the rise in leaf nitrogen associated with this deployment apparently counteracted the reduction in NAR.     

Cyanogenesis in Acacia pachyphloia

The cyanogenic glycoside, proacacipetalin, is reported from Acacia pachyphloia (Acacia subgenus Acacia). This represents the first record of a glycoside with an aliphatic aglycone from a species of Acacia indigenous to Australia. This finding reinforces the taxonomic distinctions between subgenus Acacia and subgenus Phyllodineae.     

Taxiphyllin:苦竹笋中具有酪氨酸酶抑制活性的氰苷

从苦竹笋Pleioblastus amarus（Keng）Keng f．中首次分得一个氰苷化合物,通过光谱分析及其物理化学性质鉴定为taxiphyllin（1）。该化合物在体外能显著抑制酪氨酸酶活性,是一个强有效的酪氨酸酶抑制剂。     

Relationship between cyanogenesis and latex stability on tapping panel dryness in rubber trunk girth

The presence of high cyanogenic glycoside concentrations may predispose plant to the tapping panel dryness (TPD). This study aimed to verify the involvement of cyanogenesis in the reduction of latex stability and in the establishment of TPD. The following parameters were evaluated in rubber tree trunk bark: concentration of cyanogenic glycosides with determination of cyanogenic potential (HCNp) and latex stability with lutoid bursting index (LBI). The study of the relationship between cyanogenesis and TPD was performed by semiquantitative comparison of hydrogen cyanide (HCN) gas released from the trunk bark under the following conditions: without (0%) and with (100%) TPD. The positive correlations between HCNp values and LBI indicate that cyanogenic glycosides present in the bark reduce latex stability, resulting in low yield due to the short duration of flow during tapping. The largest amount of HCN released by trunk bark tissues when the plant exhibits TPD symptoms strengthens the evidence of the involvement of this compound in the establishment of this condition.