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Background: Farfarae Flos (FF) is a frequently used traditional herbal medicine with outstanding antitussive actions. The adulteration of FF decoction pieces is common. Objective: This study aimed to study the effect of adulteration on the safety and quality of FF decoction pieces. Methods: The proportion of impurities was conducted by cone quartering method. A simple and accurate ultra-highperformance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method was established to simultaneous determinate three pyrrolizidine alkaloids (PAs) as endogenous toxic compounds in FF. The traditional medicinal parts (flower bud), impurities (pedicel and rhizome) and unselected samples were determined respectively. The values of estimated daily intake (EDI) and margin of exposure (MOE) were used for risk assessment. Results: Twenty batches of samples were collected from different habitats, and the proportion of impurities ranged from 17.51% to 41.27%. Pedicel and rhizome were the main impurities, accounting for more than 87.40% of the total impurities. The content of PAs in impurities was significantly higher. The EDI value range was 5.34 to 16.59 lg/kg bw/day, which was much higher than the standard safety value of 7.00X10-3μg/kg bw/day. The MOE values ranges for life long time and shorter exposure were 14.29 to 44.37 and 371.53 to 1153.63, respectively, indicating that at least 80% of the samples had safety risks. Correlation analysis showed that the proportion of adulterated impurities had significant correlation with the values of EDI and MOE. Conclusions: Adulteration of non medicinal parts may significantly increase the risk of medications of FF decoction pieces. Highlights: This study provides an efficient methodology reference for the control of PAs and a basis for adulteration to affect the safety and quality of FF decoction pieces. [ABSTRACT FROM AUTHOR]

Background: Studies on pyrrolizidine alkaloid (PA) contamination in honey produced in China are scarce. Previously reported HPLC-MS/MSmethods for the determination of PAs in honey often suffer from insufficient separation and uncertainties in PA isomers. Objective: To develop and validate an Ultra-HPLC (UHPLC)-MS/MS method for baseline separation of PA isomers towards precise determination of 32 PAs in Chinese wild honey. Methods: PAs were extracted from honey samples and separated on an ACQUITY BEH C18 (2.1mm 100mm, 1.7 lm) column with (A) 0.1% formic acid aqueous solution containing 5mM ammonium acetate and (B) methanol as mobile phase. The column temperature was maintained at 30 °C, and flow rate was 0.3 mL/min. Detection was performed by tandem mass spectrometry. The total run time was reduced to 18 min. Results: Thirty-one of 32 PAs were baseline separated efficiently within 18 min. The LOD and LOQ were 0.06-0.25 lg/kg and 0.22-0.82 lg/kg, respectively, except for that of clivorine, for which LOD and LOQ were 2.03 and 6.78 lg/kg, respectively. The average recoveries ranged between 66.3 and 95.1% and the average RSDs were 3.2 to 8%. The established method was used to analyze PAs in 22 types of Chinese wild honey, and the predominant PAs found in these honey samples were intermedine and lycopsamine. Conclusion: A high-throughput method for the determination of isomeric PAs in honey was developed and validated. Five of the 22 types of Chinese wild honey were contaminated with PAs concentrations of 2.2-207.0 lg/kg. Highlights: A new method capable of monitoring more PAs and providing better separation than previously reported protocols for the determination of multiclass PAs in honey is established. [ABSTRACT FROM AUTHOR]

Understanding the plants pollinators use through the year is vital to support pollinator populations and mitigate for declines in floral resources due to habitat loss. DNA metabarcoding allows the temporal picture of nectar and pollen foraging to be examined in detail. Here, we use DNA metabarcoding to examine the forage use of honeybees (Apis mellifera L.) within a florally diverse landscape within the UK, documenting the key forage plants used and seasonal progression over two years. The total number of plant taxa detected in the honey was 120, but only 16 of these were found with a high relative read abundance of DNA, across the main foraging months (April–September). Only a small proportion of the available flowering genera in the landscape were used by the honeybees. The greatest relative read abundance came from native or near-native plants, including Rubus spp. Trifolium repens , the Maleae tribe including Crataegus, Malus , and Cotoneaster , and Hedera helix. Tree species were important forage in the spring months, followed by increased use of herbs and shrubs later in the foraging season. Garden habitat increased the taxon richness of native, near-native and horticultural plants found in the honey. Although horticultural plants were rarely found abundantly within the honey samples, they may be important for increasing nutritional diversity of the pollen forage. [ABSTRACT FROM AUTHOR]

Polyamines are important metabolites in plant development and abiotic and biotic stress responses. Copper-containing amine oxidases (CuAOs) are involved in the regulation of polyamine levels in the cell. CuAOs oxidize primary amines to their respective aldehydes and hydrogen peroxide. In plants, aldehydes are intermediates in various biosynthetic pathways of alkaloids. CuAOs are thought to oxidize polyamines at only one of the primary amino groups, a process frequently resulting in monocyclic structures. These oxidases have been postulated to be involved in pyrrolizidine alkaloid (PA) biosynthesis. Here, we describe the identification and characterization of homospermidine oxidase (HSO), a CuAO of Heliotropium indicum (Indian heliotrope), involved in PA biosynthesis. Virus-induced gene silencing of HSO in H. indicum leads to significantly reduced PA levels. By in vitro enzyme assays after transient in planta expression, we show that this enzyme prefers Hspd over other amines. Nuclear magnetic resonance spectroscopy and mass spectrometry analyses of the reaction products demonstrate that HSO oxidizes both primary amino groups of homospermidine (Hspd) to form a bicyclic structure, 1-formylpyrrolizidine. Using tracer feeding, we have further revealed that 1-formylpyrrolizidine is an intermediate in the biosynthesis of PAs. Our study therefore establishes that HSO, a canonical CuAO, catalyzes the second step of PA biosynthesis and provides evidence for an undescribed and unusual mechanism involving two discrete steps of oxidation that might also be involved in the biosynthesis of complex structures in other alkaloidal pathways. [ABSTRACT FROM AUTHOR]

Sunn hemp (Crotalaria juncea) has been widely used as cover crop in the USA and due to superior biomass accumulation and nutritive value, there is interest from producers to use sunn hemp as forage for livestock. However, sunn hemp has pyrrolizidine alkaloids, which can affect animal performance. The objective of this study was to evaluate the effects of feeding sunn hemp hay to early-weaned beef calves in a dry lot in Florida. The experiment was conducted at the UF/IFAS Range Cattle Research and Education Center, Ona, FL from April to May 2021 (42 d). Treatments were calves receiving sunn hemp or stargrass (Cynodon nlemfuensis) hay distributed in a randomized complete block design with 12 replicates. Calves were supplemented with concentrate at 1.3% BW daily (18% CP and 78% TDN). Twenty-four Angus crossbred calves were weaned with 90 ± 14 kg BW and grazed annual ryegrass (Lolium multiflorum) with 1% BW concentrate supplementation for approximately 60 d. Subsequently, calves were randomly distributed and assigned to experimental units (pen) on 14 April 2021 with initial BW of 184 ± 10 kg. 'Ubon' sunn hemp hay was harvested approximately 49 d after seeding when 75% of the plants were initially flowering. Ground hay was fed daily targeting 10% refusals, and the initial CP and in vitro digestible organic matter concentrations of sunn hemp and stargrass hay were 9 and 48%, and 9 and 47%, respectively. The average concentration of pyrrolizidine alkaloids from sunn hemp hay was 58 μg/g DM. Early-weaned calves receiving stargrass had a tendency to have greater ADG than sunn hemp (0.5 vs. 0.4 kg/d, P = 0.09), and had greater forage DM intake (1.2 vs. 1.0% BW, P < 0.01) and total DM intake (2.5 vs. 2.3% BW, P < 0.01). Conversely, diets with sunn hemp had greater in vivo DM digestibility than stargrass (68 vs. 62%, P = 0.04). There was no difference between treatments on parasite egg count (mean = 45 eggs/3 g feces, P = 0.25) and coccidia egg count (log10 eggs/g feces, P = 0.35). Earlyweaned calves receiving sunn hemp consumed 0.25 g pyrrolizidine alkaloids/d; however, there was no detectable amount (< 0.05 μg/g DM) of pyrrolizidine alkaloids in the feces. The presence of pyrrolizidine alkaloids in sunn hemp may be the potential cause of decreased early-weaned calves forage intake and animal performance. [ABSTRACT FROM AUTHOR]

Background: Pyrrolizidine alkaloids (PAs) are naturally occurring plant toxins associated with potential hepatic and carcinogenic diseases in humans and animals. The concern over PAs has increased as the consumption of herbal medicines has increased. Objective: This study aimed to develop and validate a sensitive analytical method to determine 28 PAs in five herbal medicines using liquid chromatography (LC)-electrospray ionization (ESI)-tandem mass spectrometry (MS/MS). Additionally, this study identified and quantified the amount of PAs in 10 samples of each herbal medicine. Methods: The pretreatment in the proposed LC-MS/MS analysis comprised solvent extraction using 0.05M H2SO4 in 50% methanol and clean-up step using an mixed-mode cationic exchange (MCX)-solid-phase extraction (SPE) cartridge. The PA contents in herbal medicines were measured by using the developed method. Results: The proposed method had recoveries ranging from 72.5-123.7% for the Atractylodis Rhizoma Alba, 70.6-151.7% for Alba Chrysanthmi Flos, 80.6-130.9% for Leonuri Herba, 70.3-122.9% for Gastrodiae Rhizoma, and 67.1-106.9% for Glycyrrhizae Radix. Even though a few samples showed recoveries in unsatisfactory values, the proposed method indicated entirely sufficient recoveries and precision in most samples. In monitoring results, only Leonuri Herba contained two PAs, which indicated Retrorsine (4/10) of 84.7-120.9 μg/kg and Senkirkine (10/10) of 60.9-170.7 μg/kg. Conclusion: The results obtained from this study demonstrate that the proposed method is fit for purpose to determine 28 PAs in herbal medicines. Therefore it could serve as a regulatory method capable of being used for controlling the risks of PAs in certain medicinal plants and dietary supplements. Highlights: An LC-MS/MS method for the determination of 28 pyrrolizidine alkaloids in herbal medicines was developed and validated through this study. The proposed method is considered as an useful method for monitoring pyroolizidine alkaloids in herbal medicines. [ABSTRACT FROM AUTHOR]

Porous organic polymers have gained great research interest in the field of adsorption. A benzoxazine porous organic polymer (BoxPOP) constructed from p -phenylenediamine, 1,3,5-trihydroxybenzene and paraformaldehyde was fabricated and explored as an adsorbent for solid-phase extraction (SPE) of four chlorophenols from water and honey samples. Under the optimized SPE conditions, the response linearity for the analysis of the SPE extract of the chlorophenols by high performance liquid chromatography-diode array detector was observed in the range of 0.2–40.0 ng mL−1 for water samples and 5.0–400.0 ng g−1 for honey samples. The method detection limits of the analytes were 0.06–0.08 ng mL−1 for water samples and 1.5–2.0 ng g−1 for honey samples. The recoveries of the analytes from fortified water and honey samples ranged from 84.8 to 119.0% with the relative standard deviations below 8.4%. The results indicate that the prepared BoxPOP is an effective adsorbent for the chlorophenols. The established method provides an alternative approach for the determination of chlorophenols in real samples. [ABSTRACT FROM AUTHOR]

Berberine (BBR) is a protoberberine alkaloid extracted from plants such as Coptis japonica (Ranunculaceae). In a previous report, we demonstrated the existence of a 11-hydroxylation pathway employed by BBR-utilizing bacteria for metabolism of BBR. In the present study, we report the identification of the genes brhA, brhB, and brhC as encoding a multicomponent BBR 11-hydroxylase in Burkholderia sp. strain CJ1. BrhA is belonging to the Rieske non-heme iron oxygenase (RO) family, a class of enzymes known to catalyze the first step in bacterial aromatic-ring hydroxylation. We further demonstrate that BrhA activity requires BrhB (ferredoxin reductase) and BrhC (ferredoxin) as electron transport chain components. A BLAST search revealed that BrhA exhibits 38% and 33% sequence identity to dicamba O-demethylase (DdmC; AY786443) and chloroacetanilide herbicides N-dealkylase (CndA; KJ461679), respectively. To our knowledge, this work represents the first report of a bacterial oxygenase catalyzing the metabolism of a polycyclic aromatic-ring alkaloid. Abbreviations: BBR: berberine; D-BBR: demethyleneberberine; H-BBR: 11-hydroxyberberine; HD-BBR: 11-hydroxydemethyleneberberine; HDBA: 2-hydroxy-3,4-dimethoxybenzeneacetic acid; PAL: palmatine; H-PAL: 11-hydroxypalmatine; BRU: berberrubine; Fd: ferredoxin; FdR: ferredoxin reductase; ETC: electron transport chain Components and functions of the BBR 11-hydroxylase from Burkholderia sp. Strain CJ1. [ABSTRACT FROM AUTHOR]

β-Carboline alkaloids exhibit a broad spectrum of pharmacological and biological activities and are widely distributed in nature. Genetic information on the biosynthetic mechanism of β-carboline alkaloids has not been accumulated in bacteria, because there are only a few reports on the microbial β-carboline compounds. We previously isolated kitasetaline, a mercapturic acid derivative of a β-carboline compound, from the genetically modified Kitasatospora setae strain and found a plausible biosynthetic gene cluster for kitasetaline. Here, we identified and characterized three kitasetaline (ksl) biosynthetic genes for the formation of the β-carboline core structure and a gene encoding mycothiol-S-conjugate amidase for the modification of the N-acetylcysteine moiety by using heterologous expression. The proposed model of kitasetaline biosynthesis shows unique enzymatic systems for β-carboline alkaloids. In addition, feeding fluorotryptophan to the heterologous Streptomyces hosts expressing the ksl genes led to the generation of unnatural β-carboline alkaloids exerting novel/potentiated bioactivities. [ABSTRACT FROM AUTHOR]

Objective: Androgen excess in childhood is a common presentation and may signify sinister underlying pathology. Data describing its patterns and severity are scarce, limiting the information available for clinical decision processes. Here, we examined the differential diagnostic value of serum DHEAS, an drostenedione (A4) and testosterone in childhood androgen excess. Design: Retrospective review of all children undergoing serum androgen measurement at a single center over 5 years. Methods: Serum A4 and testosterone were measured by tandem mass spectrometry and DHEAS by immunoassay. Patients with at least one increased androgen underwent phenotyping by clinical notes review. Results: In 487 children with simultaneous DHEAS, A4 and testosterone m easurements, we identified 199 with androgen excess (140 pre- and 59 post-pubertal). Premature adre narche (PA) was the most common pre-pubertal diagnosis (61%), characterized by DHEAS excess in 85%, while A4 and testosterone were only increased in 26 and 9% respectively. PCOS was diagnosed in 40% of post-pubertal subjec ts, presenting equally frequent with isolated excess of DHEAS (29%) or testosterone (25%) or increases in both A4 and t estosterone (25%). CAH patients (6%) predominantly had A4 excess (86%); testosterone and DHEAS were increased in 5 0 and 33% respectively. Concentrations increased above the two-fold upper limit of normal were mostly observed i n PA for serum DHEAS (>20-fold in the single case of adrenocortical carcinoma) and in CAH for serum androstenedione. Conclusions: Patterns and severity of childhood androgen excess provide pointers to the underlying diagnosis and can be used to guide further investigations. [ABSTRACT FROM AUTHOR]

Pyrrolizidine alkaloids (PAs) are characteristicmetabolites of some plant families and forma powerful defensemechanism against herbivores. More than 600 different PAs are known. PAs are ester alkaloids composed of a necine base and a necic acid, which can be used to divide PAs in different structural subcategories. Themain target organs for PAmetabolismand toxicity are liver and lungs. Additionally, PAs are potentially genotoxic, carcinogenic and exhibit developmental toxicity. Only for very few PAs, in vitro and in vivo investigations have characterized their toxic potential. However, these investigations suggest that structural differences have an influence on the toxicity of single PAs. To investigate this structural relationship for a large number of PAs, a quantitative structural-activity relationship (QSAR) analysis for hepatotoxicity of over 600 different PAs was performed, using Random Forest- and artificial Neural Networks-algorithms. Thesemodels were trained with a recently established dataset specific for acute hepatotoxicity in humans. Using this dataset, a set of molecular predictors was identified to predict the hepatotoxic potential of each compound in validated QSAR models. Based on thesemodels, the hepatotoxic potential of the 602 PAs was predicted and the following hepatotoxic rank order in 3main categories defined (1) for necine base: otonecine>retronecine>platynecine; (2) for necine basemodification: dehydropyrrolizidine≫tertiary PA=N-oxide; and (3) for necic acid: macrocyclic diester≥open-ring diester>monoester. A further analysis with combined structural features revealed that necic acid has a higher influence on the acute hepatotoxicity than the necine base. [ABSTRACT FROM AUTHOR]

Close associations of certain lepidopteran taxa with pyrrolizidine alkaloids (PAs), a typical class of plant secondary metabolites, have been well documented from the perspective of evolutionary ecology. Male danaine butterflies are thought to utilize PAs as precursors for the production of dihydropyrrolizines [e.g. danaidone (DO) and hydroxydanaidal (HD)] in their two distinct androconial organs, viz. alar scent organs (sex brands) and abdominal hairpencils. However, little is known about the quantitative profiles of these compounds in danaines, the mechanism for their formation in the androconial organs, or their biological functions, particularly in mating behaviour. The present study addressed these unanswered questions posed for males of the danaine butterfly, Parantica sita. Chemical analyses of androconial extracts revealed considerable seasonal/regional and individual variations of the amounts of DO (the major dihydropyrrolizine produced) and 7 R-HD (the 7 R-enantiomer of HD detected in this study) found in the two organs. These variations seemed to depend primarily on the age of the male and the phenological traits of PA-containing plants available. Males were found to acquire an adequate capability to produce DO ~1 week after eclosion. DO was shown to be produced exclusively in the sex brand and subsequently physically transferred to the hairpencil through a contact behaviour between the two organs, here termed 'perfuming behaviour'. The results of behavioural experiments with PA-fed and PA-unfed males that were allowed to compete for mates, combined with the positive electroantennographic (EAG) responses of the female, to both DO and 7 R-HD, led to the conclusion that either or both of these compounds can act as the sex pheromone. Oral administration of PAs to males indicated that DO can be biosynthesized from various PA precursors, while 7 R-HD, unlike in arctiid moths, is derived only from PAs with the 7 R-configuration. The putative biosynthetic pathways of DO and 7 R-HD, and the evolutionary provenance of the binate androconial system in the Danainae are also discussed. [ABSTRACT FROM AUTHOR]

Four Greek endemic Boraginaceae plants, Onosma erecta Sibth. & Sm., Onosma kaheirei Teppner, Onosma leptantha Heldr., and Cynoglossum columnae L. (aerial parts), were screened for their content of pyrrolizidine alkaloids (PAs). TLC with the Mattocks-Molyneux visualization reagent was used as a preliminary qualitative test for PA or PA N-oxide detection. The extracts of the species found to contain PAs and their N-oxides were further analyzed by GC/MS, so as to identify their structures by means of the mass spectra and retention index values of known PAs already published in the literature. Twenty-three PAs were identified. For additional peaks, recognized as possible PAs by their MS pattern, no exact structures were tentatively suggested, as a result of lack of matching literature data. Furthermore, a quantitative PA profile of the species was obtained. [ABSTRACT FROM AUTHOR]

Homospermidine synthase (HSS), the first pathway-specific enzyme of pyrrolizidine alkaloid biosynthesis, is known to have its origin in the duplication of a gene encoding deoxyhypusine synthase. To study the processes that followed this gene duplication event and gave rise to HSS , we identified sequences encoding HSS and deoxyhypusine synthase from various species of the Convolvulaceae. We show that HSS evolved only once in this lineage. This duplication event was followed by several losses of a functional gene copy attributable to gene loss or pseudogenization. Statistical analyses of sequence data suggest that, in those lineages in which the gene copy was successfully recruited as HSS , the gene duplication event was followed by phases of various selection pressures, including purifying selection, relaxed functional constraints, and possibly positive Darwinian selection. Site-specific mutagenesis experiments have confirmed that the substitution of sites predicted to be under positive Darwinian selection is sufficient to convert a deoxyhypusine synthase into a HSS. In addition, analyses of transcript levels have shown that HSS and deoxyhypusine synthase have also diverged with respect to their regulation. The impact of protein–protein interaction on the evolution of HSS is discussed with respect to current models of enzyme evolution. [ABSTRACT FROM AUTHOR]

Homospermidine synthase (HSS) is the first specific enzyme in pyrrolizidine alkaloid (PA) biosynthesis, a pathway involved in the plant's chemical defense. HSS has been shown to be recruited repeatedly by duplication of a gene involved in primary metabolism. Within the lineage of the Boraginales, only one gene duplication event gave rise to HSS. Here, we demonstrate that the tissue-specific expression of HSS in three boraginaceous species, Heliotropium indicum, Symphytum officinale, and Cynoglossum officinale, is unique with respect to plant organ, tissue, and cell type. Within H. indicum, HSS is expressed exclusively in nonspecialized cells of the lower epidermis of young leaves and shoots. In S. officinale, HSS expression has been detected in the cells of the root endodermis and in leaves directly underneath developing inflorescences. In young roots of C. officinale, HSS is detected only in cells of the endodermis, but in a later developmental stage, additionally in the pericycle. The individual expression patterns are compared with those within the Senecioneae lineage (Asteraceae), where HSS expression is reproducibly found in specific cells of the endodermis and the adjacent cortex parenchyma of the roots. The individual expression patterns within the Boraginales species are discussed as being a requirement for the successful recruitment of HSS after gene duplication. The diversity of HSS expression within this lineage adds a further facet to the already diverse patterns of expression that have been observed for HSS in other PA-producing plant lineages, making this PA-specific enzyme one of the most diverse expressed proteins described in the literature. [ABSTRACT FROM AUTHOR]

Pyrrolizidine alkaloids (PAs) are the most common plant constituents that poison livestock, wildlife and humans. Riddelliine is a prototype genotoxic PA and has been nominated to be classified as a reasonably anticipated human carcinogen by the US National Toxicology Program (NTP) in the 12th Report on Carcinogens. Riddelliine’s nomination is due to the high incidence of liver tumours that were observed in both mice and rats in the NTP tumourigenicity bioassay study. In this current study, we explored whether riddelliine treatment could alter microRNA (miRNA) expression in rat liver and whether the possible deregulation of miRNA was related to mutagenicity and carcinogenicity of riddelliine. Groups of six rats were administered riddelliine at a mutagenic dose of 1 mg/kg body weight or with control vehicle 5 days a week for 12 weeks. A group of six rats treated with aristolochic acid, a renal carcinogen, was used as a tissue-specific negative control. The animals were sacrificed 1 day after the last treatment and the livers were isolated for miRNA expression analysis using miRNA microarrays. miRNA expression was significantly altered by riddelliine treatment. Principal component analysis and hierarchical clustering analysis showed that the miRNA expression profiles were clearly classified into two groups, riddelliine treatment versus other samples. Forty-seven miRNAs were significantly dysregulated by riddelliine treatment, among which 38 were up-regulated and 9 were down-regulated. Functional analysis of these differentially expressed miRNAs by riddelliine revealed that these miRNAs were involved in liver carcinogenicity and toxicity, such as liver proliferation, liver necrosis/cell death, hepatocellular carcinoma, liver hepatomegaly, liver inflammation and liver fibrosis. These results suggest that miRNAs actively respond to a mutagenic dose of riddelliine and the pattern of miRNA expression has the potential to be used as a biomarker of genotoxicity and carcinogenicity for riddelliine and possibly other PAs. [ABSTRACT FROM AUTHOR]

The article presents a study which aims to identify the cytotoxic and antitrypanosomal activities of Ethiopian plants that produce pyrrolizidine alkaloids. The study used extracts from Solanecio angulatus, which underwent alkaloid extraction and has showed good antitrypanosomal mechanisms. It used various methods such as cell culture, assaying, and hydrogen-ion concentration (pH) measurement. Results show that the dichloromethane was the plant's most active extract.

Pyrrolizidine alkaloids (PAs) are typical compounds of plant secondary metabolism and are believed to be part of the plant's chemical defense. Within the monocotyledonous plants, PAs have been described in only a few genera, mainly orchids, including Phalaenopsis. Because phylogenetic analyses suggest an independent origin of PA biosynthesis within the monocot lineage, we have analyzed the developmentally regulated expression of homospermidine synthase (HSS), the first pathway-specific enzyme of PA biosynthesis, at the cell level. HSS is expressed in the tips of aerial roots exclusively in mitotically active cells. Raphide crystal idioblasts present within the root apical meristem do not show HSS expression. In addition, young flower buds, but not mature flowers, express HSS and have been shown by tracer feeding experiments to be able to catalyze PAs. This second site of PA biosynthesis ensures high concentrations of PAs in the reproductive structures of the Phalaenopsis flower, even after the flower opens. Thus, in spite of its identical function in PA biosynthesis, HSS shows in Phalaenopsis a completely different spatial and developmental expression pattern in comparison to other PA-producing species. These results show that the proverbial diversity of plant secondary metabolism is not just a matter of structural diversity, but is also multifaceted in terms of pathway regulation and expression. [ABSTRACT FROM AUTHOR]

Examines a method for the extraction and identification of pyrrolizidine alkaloids (PA) and their N-oxides in botanical materials and commercial comfrey-containing products, using liquid chromatography electrospray ionization mass spectrometry. Optimization of the extraction procedure and the chromatographic conditions; Application of the method to the analysis of herbal remedies; Measurable quantities of PA in products that were labeled to contain comfrey.

Species of several unrelated families within the angiosperms are able to constitutively produce pyrrolizidine alkaloids as a defense against herbivores. In pyrrolizidine alkaloid (PA) biosynthesis, homospermidine synthase (HSS) catalyzes the first specific step. HSS was recruited during angiosperm evolution from deoxyhypusine synthase (DHS), an enzyme involved in the posttranslational activation of eukaryotic initiation factor 5A. Phylogenetic analysis of 23 cDNA sequences coding for HSS and DHS of various angiosperm species revealed at least four independent recruitments of HSS from DHS: one within the Boraginaceae, one within the monocots, and two within the Asteraceae family. Furthermore, sequence analyses indicated elevated substitution rates within HSS-coding sequences after each gene duplication, with an increased level of nonsynonymous mutations. However, the contradiction between the polyphyletic origin of the first enzyme in PA biosynthesis and the structural identity of the final biosynthetic PA products needs clarification. [ABSTRACT FROM AUTHOR]

Discusses a rapid cleanup method for the isolation and concentration of pyrrolizidine alkaloids (PA) in comfrey root. Solid-phase extraction method using an Ergosil cleanup column; Extraction process; Percent recoveries of PA.

Discusses the development of a simple and reliable high-performance liquid chromatographic assay for a simultaneous determination of isoline, a potent hepatotoxic pyrrolizidine alkaloid. Analysis of the metabolism of isoline by rat and mouse microsomal enzyme systems; Role of hepatic esterase in the detoxification againstisoline-induced hepatotoxicity; Means of incubating rat liver microsomes with isoline as substrate.