UPLC-MSE profiling of Phytoplankton metabolites: application to the identification of pigments and structural analysis of metabolites in Porphyridium purpureum.

A fast and high-resolution UPLC-MSE analysis was used to identify phytoplankton pigments in an ethanol extract of Porphyridium purpureum (Pp) devoid of phycobiliproteins. In a first step, 22 standard pigments were analyzed by UPLC-MSE to build a database including retention time and accurate masses of parent and fragment ions. Using this database, seven pigments or derivatives previously reported in Pp were unequivocally identified: ß,ß-carotene, chlorophyll a, zeaxanthin, chlorophyllide a, pheophorbide a, pheophytin a, and cryptoxanthin. Minor amounts of Divinyl chlorophyll a, a chemotaxonomic pigment marker for prochlorophytes, were also unequivocally identified using the database. Additional analysis of ionization and fragmentation patterns indicated the presence of ions that could correspond to hydroxylated derivatives of chlorophyll a and pheophytin a, produced during the ethanolic extraction, as well as previously described galactosyldiacylglycerols, the thylakoid coenzyme plastoquinone, and gracilamide B, a molecule previously reported in the red seaweed Gracillaria asiatica. These data point to UPLC-MSE as an efficient technique to identify phytoplankton pigments for which standards are available, and demonstrate its major interest as a complementary method for the structural elucidation of ionizable marine molecules.

Plastoquinones from Sargassum yezoense; chemical structures and effects on the activation of peroxisome proliferator-activated receptor gamma.

Four new plastoquinones, together with two known compounds, sargahydroquinoic acid and sargaquinoic acid, were isolated from the brown alga, Sargassum yezoense collected from the eastern coast of Korea. The structures of these compounds were elucidated based on spectroscopic analyses including NMR and MS. Their structures designated as meroterphenol A (1), B (2), C (3) and D (4) were characterized by a 6-methyl-1,4-benzohydroquinone moiety with an oxygenated diterpenoic acid chain. Meroterphenols A-D showed potent activation effects on peroxisome proliferator-activated receptor gamma (PPARγ).

Effects of plastoquinones from the brown alga Sargassum micracanthum and a new chromene derivative converted from the plastoquinones on acute gastric lesions in rats.

Previously, we reported the anti oxidative and anti viral effects of plastoquinones (compounds 1, 2) extracted from the seaweed Sargassum micracanthum (Kuetzing) Endlicher and a new chromene compound (compound 3), which was converted from the plastoquinones. Recently, we have also demonstrated the antiulcer effects of these compounds and assessed the effects using a rat model of acute gastric lesion and fundus strips isolated from rats. In hydrochloric acid/ethanol rat ulcer tests: 1) oral administrations of compounds 1, 2, and 3 1--10, 3--30 and 10--30 mg/kg, respectively, and omeprazole 3--30 mg/kg showed dose-dependent antiulcer effects: 2) the antiulcer effects after intraduodenal administration of the respective compounds at the dose of 30 mg/kg were found to be significant: and 3) a decrease in the hexosamine level of the gastric mucosa was slightly improved by oral administration of compounds 1, 2, and 3 30 mg/kg. In indomethacin-induced gastric ulcer tests, the antiulcer effects of compounds 1, 2, and 3 10 mg/kg (p.o.) were not significant. Compounds 1, 2, and 3 showed slight contracting effects on the fundus isolated from rats and these effects were inhibited by pretreatment with AH6809, an inhibitor of prostaglandin DP, EP(1), and EP(2) receptors. These results suggest that the protection of the mucosa via endogenous prostaglandins might be related to the antiulcer effects of compounds 1, 2, and 3.

Tumor cell proliferation and cyclooxygenase inhibitory constituents in horseradish (Armoracia rusticana) and Wasabi (Wasabia japonica).

Cyclooxygenase and human tumor cell growth inhibitory extracts of horseradish (Armoracia rusticana) and wasabi (Wasabia japonica) rhizomes upon purification yielded active compounds 1-3 from horseradish and 4 and 5 from wasabi rhizomes. Spectroscopic analyses confirmed the identities of these active compounds as plastoquinone-9 (1), 6-O-acyl-beta-d-glucosyl-beta-sitosterol (2), 1,2-dilinolenoyl-3-galactosylglycerol (3), linolenoyloleoyl-3-beta-galactosylglycerol (4), and 1,2-dipalmitoyl-3-beta-galactosylglycerol (5). 3-Acyl-sitosterols, sinigrin, gluconasturtiin, and phosphatidylcholines isolated from horseradish and alpha-tocopherol and ubiquinone-10 from wasabi rhizomes isolated were inactive in our assays. At a concentration of 60 microg/mL, compounds 1 and 2 selectively inhibited COX-1 enzyme by 28 and 32%, respectively. Compounds 3, 4, and 5 gave 75, 42, and 47% inhibition of COX-1 enzyme, respectively, at a concentration of 250 microg/mL. In a dose response study, compound 3 inhibited the proliferation of colon cancer cells (HCT-116) by 21.9, 42.9, 51.2, and 68.4% and lung cancer cells (NCI-H460) by 30, 39, 44, and 71% at concentrations of 7.5, 15, 30, and 60 microg/mL, respectively. At a concentration of 60 microg/mL, compound 4 inhibited the growth of colon, lung, and stomach cancer cells by 28, 17, and 44%, respectively. This is the first report of the COX-1 enzyme and cancer cell growth inhibitory monogalactosyl diacylglycerides from wasabi and horseradish rhizomes.

Antioxidant and antiviral activities of plastoquinones from the brown alga Sargassum micracanthum, and a new chromene derivative converted from the plastoquinones.

Two plastoquinones were isolated from the methanolic extract of the brown alga Sargassum micracanthum, and these were identified as a known 2-geranylgeranyl-6-methylbenzoquinone and its hydroquinone, respectively, based on spectroscopic analysis. The absolute configuration of the secondary hydroxyl group was determined by the modified Mosher's method using the new chromene derivative converted from plastoquinones. One of the plastoquinones and the chromene exhibited significant antioxidant activities, such as an inhibitory effect on lipid peroxidation and a radical scavenging effect on 1,1-diphenyl-2-picrylhydrazyl (DPPH). The benzoquinone-type compound and the chromene derivative were found to have potent antiviral activity against human cytomegalovirus (HCMV).

Evaluation of the anti-inflammatory and antioxidant activities of the plastoquinone derivatives isolated from Roldana barba-johannis.

A phytochemical study of Roldana barba-johannis afforded two triterpenic esters, an eremophilanolide and three compounds structurally related to the antioxidants vitamin E and plastoquinone. Evaluation of the anti-inflammatory and antioxidant activities of sargahydroquinoic acid, sargachromenol, their mixture, and their methyl esters showed that most of them are antioxidant and anti-inflammatory agents.

Recruitment of a foreign quinone into the A(1) site of photosystem I. II. Structural and functional characterization of phylloquinone biosynthetic pathway mutants by electron paramagnetic resonance and electron-nuclear double resonance spectroscopy.

Electron paramagnetic resonance (EPR) and electron-nuclear double resonance studies of the photosystem (PS) I quinone acceptor, A(1), in phylloquinone biosynthetic pathway mutants are described. Room temperature continuous wave EPR measurements at X-band of whole cells of menA and menB interruption mutants show a transient reduction and oxidation of an organic radical with a g-value and anisotropy characteristic of a quinone. In PS I complexes, the continuous wave EPR spectrum of the photoaccumulated Q(-) radical, measured at Q-band, and the electron spin-polarized transient EPR spectra of the radical pair P700(+) Q(-), measured at X-, Q-, and W-bands, show three prominent features: (i) Q(-) has a larger g-anisotropy than native phylloquinone, (ii) Q(-) does not display the prominent methyl hyperfine couplings attributed to the 2-methyl group of phylloquinone, and (iii) the orientation of Q(-) in the A(1) site as derived from the spin polarization is that of native phylloquinone in the wild type. Electron spin echo modulation experiments on P700(+) Q(-) show that the dipolar coupling in the radical pair is the same as in native PS I, i.e. the distance between P700(+) and Q(-) (25.3 +/- 0.3 A) is the same as between P700(+) and A(1)(-) in the wild type. Pulsed electron-nuclear double resonance studies show two sets of resolved spectral features with nearly axially symmetric hyperfine couplings. They are tentatively assigned to the two methyl groups of the recruited plastoquinone-9, and their difference indicates a strong inequivalence among the two groups when in the A(1) site. These results show that Q (i) functions in accepting an electron from A(0)(-) and in passing the electron forward to the iron-sulfur clusters, (ii) occupies the A(1) site with an orientation similar to that of phylloquinone in the wild type, and (iii) has spectroscopic properties consistent with its identity as plastoquinone-9.

The lurlenes, a new class of plastoquinone-related mating pheromones from Chlamydomonas allensworthii (Chlorophyceae).

Chlamydomonas allensworthii is a recently described species, which differs phenotypically and in its reproductive behaviour from other members of this genus of flagellated green protists. Its female cells temporarily or constantly excrete a luring signal into the medium to attract the male cells for mating. The sperm is susceptible to the pheromone either all the time or only after a maturation process. Maturation in this species is accompanied with a colour shift from green to khaki (strain from Lemon cove CA) or to olive (strain from Catarina TX), apparently due to degradation of chloroplast material. In the bioassay with polyacrylamide beads (plain or DEAE-substituted) to which the lures are bound, acting as mock females, Catarina-strain males are only attracted to Catarina strain lures, whereas Lemon-Cove-strain males are attracted to Lemon-Cove-strain lures and to a lesser extent, to Catarina-strain lures. Being amphiphilic acid, Lemon-Cove-strain lure is more tightly bound to DEAE beads. Catarina-strain lure is an uncharged amphiphile. The two signal compounds have been isolated and identified by chemical analysis and mass and NMR spectroscopies as O(5)-beta-D-xylosylated degradation products of the chloroplast electron transporter hydroplastoquinone, its polyprenoid side chain being oxidatively cleaved at the fourth double bond. Lurlenic acid (lurlene L) is the resulting acid and lurlenol (lurlene C) the corresponding alcohol. The pheromonal activities of both compounds have a threshold at 1 pM, and are destroyed by deglycosylation and subsequent oxidation to the quinonoid aglycon. This class of amphiphilic lures, having the same basic glycosylated hydroquinone structure, is named lurlenes.

L protein, encoded by psbL, restores normal functioning of the primary quinone acceptor, QA, in isolated D1/D2/CP47/Cytb-559/I photosystem II reaction center core complex.

Plastoquinone-9 (PQ-9)-depleted PSII reaction center core complex, consisting of CP47/D1/D2/Cytb-559/I, was isolated from spinach PSII particles. PQ-9, lipids and several proteins were extracted from the original PSII particles and separated by several steps of chromatography to be reconstituted into the isolated complex. PQ-9 reconstituted in the complex with the help of thylakoid lipids (digalactosyldiglyceride) did not function as QA by itself. However, PQ-9 simultaneously reconstituted with L protein and the thylakoid lipids successfully functioned as QA in the complex. Other proteins of PSII origin, such as CP43, H, K, nuclear encoded 4.1 and 5.0 kDa proteins, are unable to restore the QA activity in the complex.

Intermediates of tocopherol biosynthesis in the unicellular alga Scenedesmus obliquus. The presence of three isomeric methylphytylbenzoquinones.

Three isomers of methylphytylbenzoquinone have been isolated from lipids of the unicellular alga Scenedesmus obliquus, the most abundant being 2-methyl-6-phytylbenzoquinone (65% of the total). The 2-methyl-3-phytyl and 2-methyl-5-phytyl isomers amounted to 8 and 27% respectively. Previously problems have been encountered in the separation of the 3-phytyl and the 6-phytyl isomers, but in the present study it was found that they separated readily as quinols. Phytyl plastoquinone was also found and the relevance of these compounds to the biosynthesis of alpha-tocopherol is discussed. As well as phylloquinone, a hydroxyphylloquinone was detected, and studies indicated that it is the 5' carbon atom to which the hydroxy group is attached. Such a compound has been found by workers using other unicellular algae.