Omanische Heilpflanze im Fokus der Phytochemie IPB-Wissenschaftler und Partner aus Dhofar haben jüngst die omanische Heilpflanze Terminalia dhofarica unter die phytochemische Lupe genommen. Die Pflanze ist reich an…
Geschmack ist vorhersagbar: Mit FlavorMiner. FlavorMiner heißt das Tool, das IPB-Chemiker und Partner aus Kolumbien jüngst entwickelt haben. Das Programm kann, basierend auf maschinellem Lernen (KI), anhand der…
Schmidt, J.; Boettcher, C.; Kuhnt, C.; Kutchan, T. M.; Zenk, M. H.;Poppy alkaloid profiling by electrospray tandem mass spectrometry and electrospray FT-ICR mass spectrometry after [ring-13C6]-tyramine feedingPhytochemistry68189-202(2007)DOI: 10.1016/j.phytochem.2006.10.003
Papaver alkaloids play a major role in medicine and pharmacy. In this study, [ring-13C6]-tyramine as a biogenetic precursor of these alkaloids was fed to Papaver somniferum seedlings. The alkaloid pattern was elucidated both by direct infusion high-resolution ESI-FT-ICR mass spectrometry and liquid chromatography/electrospray tandem mass spectrometry. Thus, based on this procedure, the structure of about 20 alkaloids displaying an incorporation of the labeled tyramine could be elucidated. These alkaloids belong to different classes, e.g. morphinan, benzylisoquinoline, protoberberine, benzo[c]phenanthridine, phthalide isoquinoline and protopine. The valuable information gained from the alkaloid profile demonstrates that the combination of these two spectrometric methods represents a powerful tool for evaluating biochemical pathways and facilitates the study of the flux of distant precursors into these natural products.
Publikation
Nualkaew, N.; De-Eknamkul, W.; Kutchan, T. M.; Zenk, M. H.;Membrane-bound geranylgeranyl diphosphate phosphatases: Purification and characterization from Croton stellatopilosus leavesPhytochemistry671613-1620(2006)DOI: 10.1016/j.phytochem.2005.12.014
Geranylgeranyl diphosphate phosphatase is an enzyme catalyzing the dephosphorylation of geranylgeranyl diphosphate (GGPP) to form geranylgeraniol (GGOH). The enzyme activity of GGPP phosphatase was detected in leaves of Croton stellatopilosus, a Thai medicinal plant containing plaunotol, a commercial anti-peptic acyclic diterpenoid. Enzymological studies of GGPP phosphatase in C. stellatopilosis leaves revealed that the enzyme is a membrane-bound protein that could be removed from 20,000g pellet by 0.1% Triton X-100 without significant loss of enzyme activity. The solubilized enzyme preparation was separated into two activity peaks, PI and PII, by BioGel A gel filtration chromatography. PI and PII were both partially purified and characterized. PI appeared to be a tetrameric enzyme with its native molecular mass of 232 kDa and subunit size of 58 kDa, whereas PII was a monomeric enzyme with a molecular mass of 30–34 kDa. Both phosphatases utilized GGPP as the preferred substrate over farnesyl and geranyl diphosphates. The apparent Km values for GGPP of PI and PII appeared to be 0.2 and 0.1 mM, respectively. Both activities were Mg2+ independent and exhibited slightly acidic pH optima, 6.0–6.5 for PI and 6.5–7.0 for PII. The catalytic activities of PII was strongly inhibited by 1.0 mM of Zn2+, Mn2+ and Co2+, whereas that of PI was not affected. Both enzyme preparations were very stable upon storage at −20 °C for 45 days without significant loss of phosphatase activity. The presence of GGPP phosphatase enzymes in C. stellatopilosus is consistent with its putative involvement in the biosynthetic pathway of plaunotol although whether PI or PII is the actual enzyme involved in the pathway remains to be clarified.
Publikation
Hirata, K.; Poeaknapo, C.; Schmidt, J.; Zenk, M. H.;1,2-Dehydroreticuline synthase, the branch point enzyme opening the morphinan biosynthetic pathwayPhytochemistry651039-1046(2004)DOI: 10.1016/j.phytochem.2004.02.015
A synthase which oxidizes (S)-reticuline to 1,2-dehydroreticuline has been found to occur in seedlings of opium poppy (Papaver somniferum L.). Due to its instability, this enzyme could only be partly purified (ca. 5-fold enrichment). Partial characterization at this stage of purification showed that it does not need a redox cofactor and accepts both (S)-reticuline and (S)-norreticuline as substrates. [1-2H, 13C]-(R,S)-reticuline was enzymatically converted into [1-13C]-dehydroreticuline, which has been identified by mass spectrometry. Release of the hydrogen atom in position C-1 of the isoquinoline alkaloid during the oxidative conversion, was exploited as a sensitive assay system for this enzyme. The enzyme has a pH optimum of 8.75, a temperature optimum of 37 °C and the apparent KM value for the substrate reticuline was shown to be 117 μM. Moreover it could be demonstrated by sucrose density gradient centrifugation that the enzyme is located in vesicles of varying size. In combination with the previously discovered strictly stereoselective and NADPH dependent 1,2-dehydroreticuline reductase the detection of this enzyme, the 1,2-dehydroreticuline synthase, provides the necessary inversion of configuration and completes the pathway from two molecules of L-tyrosine via (S)-norcoclaurine to (R)-reticuline in opium poppy involving a total number of 11 enzymes.A synthase which oxidizes (S)-reticuline to 1,2-dehydroreticuline has been found to occur in seedlings of opium poppy (Papaver somniferum L.)
Publikation
Poeaknapo, C.; Fisinger, U.; Zenk, M. H.; Schmidt, J.;Evaluation of the mass spectrometric fragmentation of codeine and morphine after 13C-isotope biosynthetic labelingPhytochemistry651413-1420(2004)DOI: 10.1016/j.phytochem.2004.05.005
All major fragment ions of codeine and morphine were elucidated using LC–electrospray MS/MS and high resolution FT-ICR-MS combined with an IRMPD system. Nanogram quantities of labeled codeine were isolated and purified from Papaver somniferum seedlings, which were grown for up to 9 days in the presence of [ring-13C6]-l-tyrosine, [ring-13C6]-tyramine and [1,2-13C2], [6-O-methyl 13C]-(R,S)-coclaurine. The labeling degree of codeine up to 57% into morphinans was observed.