Mass spectrometric detection of chlorophyll a and the tetrapyrrole secondary metabolite tolyporphin A in the filamentous cyanobacterium HT-58-2. Approaches to high-throughput screening of intact cyanobacteria

Tolyporphins are unusual tetrapyrrole macrocycles produced by the filamentous cyanobacterium–microbial community HT-58-2, the only known source to date. Numerous cyanobacterial samples have been collected worldwide but most have not been screened for secondary metabolites. Identification of tolyporphins typically has entailed lipophilic extraction followed by chromatographic fractionation and spectroscopic and/or mass spectrometric analysis. For quantitation, lengthy lipophilic extraction, sample processing and HPLC separation are needed. Examination by MALDI-TOF-MS (with the matrix 1,5-diaminonaphthalene) of lipophilic crude extracts of small-scale HT-58-2 samples (2 mL) without chromatographic fractionation enabled semi-quantitation of tolyporphin A over a 41-day growth period. Screening for tolyporphin A in intact or slightly sheared and vortexed HT-58-2 samples (no lipophilic extraction), and confirmation of identity by tandem MS, were carried out by IR-MALDESI-FTMS. Tolyporphin A was identified by the molecular ion and four characteristic fragments. The molecular ion of chlorophyll [Formula: see text] also was observed. The sheared and vortexed sample contained substantial numbers of intact cells as demonstrated by regrowth of the filamentous cyanobacterium–microbial culture. The semi-quantitative and rapid qualitative methods developed herein should facilitate examination of other tolyporphin-producing organisms among the vast worldwide strains of cyanobacteria as well as investigation of the biosynthesis of tolyporphins.

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Nanopipettes: probes for local sample analysis

Chemical Science ◽

10.1039/c5sc00668f ◽

2015 ◽

Vol 6 (6) ◽

pp. 3334-3341 ◽

Cited By ~ 34

Author(s):

Anumita Saha-Shah ◽

Anna E. Weber ◽

Jonathan A. Karty ◽

Steven J. Ray ◽

Gary M. Hieftje ◽

...

Keyword(s):

Single Cells ◽

Mass Spectrometric ◽

Mass Spectrometric Analysis ◽

Small Scale ◽

Spectrometric Analysis ◽

Sample Analysis ◽

Local Mass ◽

Local Sample

Nanopipettes are demonstrated as probes for local mass spectrometric analysis with potential for small-scale extraction of analytes from single cells, tissue and organisms.

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Computer correction for molecular ion interferences observed in spark source mass spectrometric analysis of non-conducting materials

Spectrochimica Acta Part B Atomic Spectroscopy ◽

10.1016/0584-8547(84)80232-3 ◽

1984 ◽

Vol 39 (9-11) ◽

pp. 1497-1501 ◽

Cited By ~ 1

Author(s):

J.R. Bacon ◽

A.M. Ure

Keyword(s):

Mass Spectrometric ◽

Mass Spectrometric Analysis ◽

Spectrometric Analysis ◽

Molecular Ion ◽

Source Mass ◽

Conducting Materials

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Mass Spectrometric Analysis of N2-Formation Induced by the Oxidation of Hydrazine and Hydroxylamine in Flash Illuminated Thylakoid Preparations of the Filamentous Cyanobacterium Oscillatoria chalybea

Zeitschrift für Naturforschung C ◽

10.1515/znc-1991-7-820 ◽

1991 ◽

Vol 46 (7-8) ◽

pp. 629-634 ◽

Cited By ~ 2

Author(s):

P. He ◽

K. P. Bader ◽

G . H. Schmid

Keyword(s):

Mass Spectrometry ◽

Photosystem Ii ◽

Oxygen Uptake ◽

Mass Spectrometric ◽

Spectrometric Analysis ◽

Filamentous Cyanobacterium ◽

High Concentration ◽

Single Turnover ◽

Oscillatoria Chalybea ◽

Light Flashes

In tobacco chloroplasts hydrazine-dependent dinitrogen formation measured by mass spectrometry as the consequence of short saturating light flashes is always linked to a substantial oxygen uptake (G. Renger, K. P. Bader, and G. H. Schmid, Biochim. Biophys. Acta 1015, 288, 1990). However, in thylakoids of the filamentous cyanobacterium Oscillatoria chalybea this dinitrogen formation is not linked to an apparent O2-uptake, even at the high concentration of 1 mм hydrazine. Whereas in tobacco chloroplasts Tris-treatment does not affect hydrazine dependent dinitrogen formation up to a concentration of 3 mм hydrazine, Tris-treatment of thylakoids of O. chalybea affects strongly both oxygen evolution and dinitrogen evolution under a single turnover flash as well as under ten flashes. In contrast to tobacco chloroplasts, the presence of hydrazine up to concentrations of 3 mм does not substantially affect photosynthetic O2-evolution. The observed dinitrogen evolution is affected by DCMU regardless whether induced by a single turnover flash or by ten flashes, whereas in tobacco dinitrogen evolution and the O2-uptake linked to it (which is not observed in the cyanobacterium) were clearly not affected by DCMU in the single turnover flash. In Oscillatoria the earlier described Photosystem II-mediated H2O2 formation and decomposition is influenced by hydrazine. In the presence of 300 μм hydrazine the usually present O2-uptake leading to H2O2 formation appears diminished.

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Mass spectrometric analysis of a photosystem-II-mediated oxygen uptake phenomenon in the filamentous cyanobacterium, Oscillatoria chalybea

Biochimica et Biophysica Acta (BBA) - Bioenergetics ◽

10.1016/0005-2728(88)90234-4 ◽

1988 ◽

Vol 936 (2) ◽

pp. 179-186 ◽

Cited By ~ 17

Author(s):

K.P. Bader ◽

G.H. Schmid

Keyword(s):

Photosystem Ii ◽

Oxygen Uptake ◽

Mass Spectrometric ◽

Mass Spectrometric Analysis ◽

Spectrometric Analysis ◽

Filamentous Cyanobacterium ◽

Oscillatoria Chalybea

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Rapid Compound Integrity Assessment for High-Throughput Screening Hit Triaging

SLAS DISCOVERY Advancing Life Sciences ◽

10.1177/2472555220919097 ◽

2020 ◽

pp. 247255522091909

Author(s):

Wilson Z. Shou ◽

Samuel W. Gerritz ◽

David Harden ◽

R. Michael Lawrence ◽

Peter Chase ◽

...

Keyword(s):

Liquid Chromatography ◽

High Throughput ◽

High Throughput Screening ◽

High Speed ◽

Mass Spectrometric ◽

Spectrometric Analysis ◽

Liquid Sample ◽

Integrity Assessment ◽

Novel Approach

Hits from high-throughput screening (HTS) assays are typically evaluated using cheminformatics and/or empirical approaches before a decision for follow-up (activity confirmation and/or sample resynthesis) is made. However, the compound integrity (i.e., identity and purity) of these hits often remains largely unknown at this stage, since many compounds in the screening collection could undergo various changes such as degradation, polymerization, and precipitation during storage over time. When compound integrity is actually assessed for HTS hits postassay to address this issue, the process often increases the overall cycle time by weeks due to the reacquisition of the samples and the lengthy liquid chromatography–ultraviolet/mass spectrometric analysis time. Here we present a novel approach where compound integrity data are collected concurrently with the concentration–response curve (CRC) stage of HTS, with both assays occurring either in parallel on two distributions from the same liquid sample or serially using the original source liquid sample. The rapid generation of compound integrity data has been enabled by a high-speed ultra-high-pressure liquid chromatography–ultraviolet/mass spectrometric platform capable of analyzing ~2000 samples per instrument per week. From this parallel approach, both compound integrity and CRC potency results for screening hits become available to medicinal chemists at the same time, which has greatly enhanced the decision-making process for hit follow-up and progression. In addition, the compound integrity results from recent hits provide a real-time and representative “snapshot” of the sample integrity of the entire compound collection, and the data can be used for in-depth analyses of the screening collection.

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