Analysis and Modification of Ergot Alkaloid Profiles in Fungi

Panaccione, Daniel G.; Ryan, Kathleen J.; Schardl, Christopher L.; Florea, Simona

doi:10.1016/b978-0-12-394290-6.00012-4

Cited by 48 publications

(55 citation statements)

References 58 publications

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“…Unknown A fluoresced more intensely at the 272-and 372-nm wavelength settings than at the 310-and 410-nm wavelength settings (Fig. 4), which is typical of ergot alkaloids lacking a double bond between positions 9 and 10 of the ergoline nucleus (12). In contrast, unknown B fluoresced more intensely at the 310-and 410-nm wavelengths than at the 272-and 372-nm wavelengths, indicating the presence of a 9,10 double bond (12).…”

Section: Fig 4 Qualitative Analysis Of Ergot Alkaloids From Transformmentioning

confidence: 91%

“…Conidia in each extract were counted to provide an estimate of fungal biomass. Extracts clarified by centrifugation were then analyzed by reverse-phase high-performance liquid chromatography (HPLC) with fluorescence detection (12). Briefly, samples were separated on a 150-mm by 4.6-mm C 18 column (Phenomenex Prodigy ODS3, 5-m particle size; Torrance, CA) with a multilinear binary gradient of 5% acetonitrile plus 50 mM ammonium acetate to 75% acetonitrile plus 50 mM ammonium acetate.…”

Section: Methodsmentioning

confidence: 99%

“…Samples were analyzed with two fluorescence detectors; excitation and emission wavelengths are indicated. Lysergic acid and other ergot alkaloids with a 9,10 double bond fluoresce more strongly at 310 and 410 nm, whereas other ergot alkaloids fluoresce maximally with settings of 272 and 372 nm (12). Ergot alkaloids with 9,10 double bonds form diastereoisomers at carbon 8.…”

Section: Fig 4 Qualitative Analysis Of Ergot Alkaloids From Transformmentioning

confidence: 99%

“…Abbreviations: LA, lysergic acid; ILA, isolysergic acid; IS, isosetoclavine; S, setoclavine; UB, unknown B; Ch, chanoclavine; Ag, agroclavine; UA, unknown A. The peak eluting at 53 min is present in nontransformed A. fumigatus (15), and its fluorescence ratio indicates that it is not an ergot alkaloid (12).…”

Section: Fig 4 Qualitative Analysis Of Ergot Alkaloids From Transformmentioning

confidence: 99%

“…1) (4,10,11). Ergot alkaloidproducing fungi in the Trichocomaceae, such as the opportunistic human pathogen Aspergillus fumigatus, produce clavine-based derivatives in which the D ring is saturated and carbon 17 remains reduced as a methyl group (12,13).…”

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confidence: 99%

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Heterologous Expression of Lysergic Acid and Novel Ergot Alkaloids in Aspergillus fumigatus

Robinson

Panaccione

2014

Appl Environ Microbiol

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Different lineages of fungi produce distinct classes of ergot alkaloids. Lysergic acid-derived ergot alkaloids produced by fungi in the Clavicipitaceae are particularly important in agriculture and medicine. The pathway to lysergic acid is partly elucidated, but the gene encoding the enzyme that oxidizes the intermediate agroclavine is unknown. We investigated two candidate agroclavine oxidase genes from the fungus Epichloë festucae var. lolii ؋ Epichloë typhina isolate Lp1 (henceforth referred to as Epichloë sp. Lp1), which produces lysergic acid-derived ergot alkaloids. Candidate genes easH and cloA were expressed in a mutant strain of the mold Aspergillus fumigatus, which typically produces a subclass of ergot alkaloids not derived from agroclavine or lysergic acid. Candidate genes were coexpressed with the Epichloë sp. Lp1 allele of easA, which encodes an enzyme that catalyzed the synthesis of agroclavine from an A. fumigatus intermediate; the agroclavine then served as the substrate for the candidate agroclavine oxidases. Strains expressing easA and cloA from Epichloë sp. Lp1 produced lysergic acid from agroclavine, a process requiring a cumulative six-electron oxidation and a double-bond isomerization. Strains that accumulated excess agroclavine (as a result of Epichloë sp. Lp1 easA expression in the absence of cloA) metabolized it into two novel ergot alkaloids for which provisional structures were proposed on the basis of mass spectra and precursor feeding studies. Our data indicate that CloA catalyzes multiple reactions to produce lysergic acid from agroclavine and that combining genes from different ergot alkaloid pathways provides an effective strategy to engineer important pathway molecules and novel ergot alkaloids.

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Section: Fig 4 Qualitative Analysis Of Ergot Alkaloids From Transformmentioning

confidence: 91%

Section: Methodsmentioning

confidence: 99%

Section: Fig 4 Qualitative Analysis Of Ergot Alkaloids From Transformmentioning

confidence: 99%

Section: Fig 4 Qualitative Analysis Of Ergot Alkaloids From Transformmentioning

confidence: 99%

mentioning

confidence: 99%

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Heterologous Expression of Lysergic Acid and Novel Ergot Alkaloids in Aspergillus fumigatus

Robinson

Panaccione

2014

Appl Environ Microbiol

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Fungal Endophytes of Grasses and Morning Glories, and Their Bioprotective Alkaloids

2014

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Analysis of paspalic acid, lysergic acid, and iso‐lysergic acid by capillary zone electrophoresis with UV‐ and quadrupole time‐of‐flight mass spectrometric detection

Himmelsbach

Ferdig²,

Rohrer³

2013

Electrophoresis

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CZE was investigated for separation of lysergic, iso-lysergic, and paspalic acid. BGEs were optimized regarding separation selectivity and analysis time as well as MS compatibility. BGEs using asparagine, Na-tetraborate, or ammonium acetate yielded satisfactory resolution when 40% of methanol was added and the pH adjusted to 8.3. Applying acidic BGEs also allowed fast separations but the poor stability under acidic conditions of the selected analytes prevented further use. With ultraviolet (UV) detection, LODs were 0.45 and 0.40 mg/L for paspalic acid and lysergic acid, respectively. Run-to-run precision of peak areas was 1.8% for lysergic acid and 1.9% for paspalic acid and day-to-day precision was 2.4 and 4.0%, respectively. When MS detection was used LODs improved to 0.09 mg/L for paspalic acid and 0.07 mg/L for lysergic acid. Repeatability results were excellent for a CZE-MS method without internal standard ranging from 3.4% for the highest standard concentration to 5.8% for the lowest concentration. Recovery and matrix effects were studied with samples taken from different stages of the manufacturing process and yielded an average recovery of 100.8% and a RSD of 5.7%.

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Analysis and Modification of Ergot Alkaloid Profiles in Fungi

Cited by 48 publications

References 58 publications

Heterologous Expression of Lysergic Acid and Novel Ergot Alkaloids in Aspergillus fumigatus

Heterologous Expression of Lysergic Acid and Novel Ergot Alkaloids in Aspergillus fumigatus

Fungal Endophytes of Grasses and Morning Glories, and Their Bioprotective Alkaloids

Analysis of paspalic acid, lysergic acid, and iso‐lysergic acid by capillary zone electrophoresis with UV‐ and quadrupole time‐of‐flight mass spectrometric detection

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