Revision of Coelastrella (Scenedesmaceae, Chlorophyta) and first register of this green coccoid microalga for continental Norway.

A terrestrial green microalga was isolated at Ås, in Akershus County, Norway. The strain corresponded to a coccoid chlorophyte. Morphological characteristics by light and electron microscopy, in conjunction with DNA amplification and sequencing of the 18 s rDNA gene and ITS sequences, were used to identify the microalgae. The characteristics agree with those of the genus Coelastrella defined by Chodat, and formed a sister group with the recently described C. thermophila var. globulina. Coelastrella is a relatively small numbered genus that has not been observed in continental Norway before; there are no previous cultures available in collections of Norwegian strains. Gas chromatography analyses of the FAME-derivatives showed a high percentage of polyunsaturated fatty acids (44-45%) especially linolenic acid (C18:3n3; 30-34%). After the stationary phase, the cultures were able to accumulate several carotenoids as neoxanthin, pheophytin a, astaxanthin, canthaxanthin, lutein, and violaxanthin. Due to the scarcity of visual characters suitable for diagnostic purposes and the lack of DNA sequence information, there is a high possibility that species of this genus have been neglected in local environmental studies, even though it showed interesting properties for algal biotechnology.

Simultaneous Double-Pass Absorption and Fluorescence Excitation-Emission Matrix Spectroscopy for Measurements of Reaction Kinetics.

A novel Hadamard-transform excitation-emission matrix (EEM) spectrometer generates two-dimensional (2D) fluorescence matrices at a data acquisition rate of over 6 EEMs per minute and with a spectral resolution of 5.3 nm. Using Fresnel reflections from the sample cell, we could record optical transmission spectra synchronously with the 2D EEMs. The spectrometer was integrated into a custom-designed stopped-flow injection device to collect visible absorption and fluorescence EEM spectra of reacting solutions. Two different kinetic studies on two rapidly evolving chemical reactions with multiple overlapping spectral components were conducted by collecting over 8400 absorption spectra and EEMs. The third-order rate constant for the demetalation of chlorophyll-a to pheophytin-a was experimentally determined to be 450 ± 20 M-2·s-1 as derived from a parallel factor (PARAFAC) analysis where absorption and fluorescence data were combined. A PARAFAC analysis of data collected from the insertion of a copper atom into pheophytin-a resulted in several absorbing components and only a single fluorescing component. A reaction model with an association complex and a sitting-a-top (SAT) complex as intermediates explained the absorption data, resulting in a sequence of second-order reactions with rate constants of 4.0 ± 0.4, 2.7 ± 0.3, and 0.28 ± 0.02 M-1·s-1, respectively. The rate constant of the fluorescence decrease was determined to be 1.7 ± 0.2 M-1·s-1, which is consistent with the fluorescence component being attributed to both the pheophytin-a and the association complex.

Photosynthetic pigments in acid mine drainage: Seasonal patterns and associations with stressful abiotic characteristics.

Acid mine drainage represents an extreme type of water pollution. The environments develop especial hydrochemical and ecological characteristics, such as high concentrations of acidity and low biodiversity, with dominance of acidophilic organisms. Ecological criteria are assuming increasing relevance for assessing water quality, including in acid mine drainage-affected systems. Photosynthetic pigments, like chlorophyll, could be considered in this context as they are generally used to estimate phytoplankton biomass. The present work was focused on acid mine drainage and their relationships with chlorophyll a. It was developed in the historic mine of São Domingos (Iberian Pyrite Belt, SW Europe). The methodological approach comprised two sampling campaigns (October and February) to represent seasonal behaviour. The diversity of hydrological conditions was also considered through sampling sites established in the pit lake, acidic lagoons and affected stream. Hydrochemistry and chlorophyll a were analysed. The obtained results revealed very low pH values, with a minimum of 2.1. In general, hydrochemistry indicates higher concentrations of pollutants in summer. One of the sampling point, located in an acidic lagoon, stood out by highest concentrations, presenting maximum of sulfate (6564 mg/L), As (6.26 mg/L), and metals like Al (675 mg/L), Cd (1,30 mg/L), Zn (199 mg/L). These highest concentrations coincided with the maximum concentration of chlorophyll a (113 µg/L). The results suggested that the more contaminated the environment, more chlorophyll a was produced. Factor analyses emphasised the relationships between acid drainage properties and the photosynthetic activity, indicating a strong pH dependency of chlorophyll a production.

Effect of Oven-Drying on the Recovery of Valuable Compounds from Ulva rigida, Gracilaria sp. and Fucus vesiculosus.

The effect of oven-drying at 25, 40 and 60 °C was evaluated on three macroalgae of relevance in Europe, namely Ulva rigida, Gracilaria sp. and Fucus vesiculosus, with respect to quality aspects, including their potential to be exploited as a source of valuable compounds. Notably, as compared to freeze-drying, oven-drying at 25 °C promoted the extraction of chlorophylls and carotenoids from U. rigida, as well as those of phycoerythrin and chlorophyll a from Gracilaria sp., while 40 °C favored the recovery of fucoxanthin and pheophytin a from F. vesiculosus. On the other hand, the use of oven-drying had a negative impact on the extraction of phenolic compounds from this alga, also diminishing the antioxidant activity of the resulting extracts. Instead, the impact of oven-drying of raw material on the recovery of specific polysaccharides differed among the macroalgae. While the amounts of ulvans and fucoidans obtained from macroalgae dried at higher temperatures tended to be superior, the recovery of agar was not affected with the drying temperatures applied to Gracilaria sp. The overall results showed that oven-drying might serve as a good alternative to stabilize Ulva rigida, Gracilaria sp. and Fucus vesiculosus, especially if extraction of pigments and polysaccharides is aimed, thought the appropriate temperature applied must be adapted for each macroalgae.

Morphophysiological variation and metal concentration in the thallus of Parmotrema tinctorum (Despr. ex Nyl.) Hale between urban and forest areas in the subtropical region of Brazil.

Anthropic activities such as the emission of pollutants resulting from industrial and agropastoral activities promote several changes in urban and forest areas. Lichens are organisms that are used in air quality evaluations due to their sensitivity to these changes. The aim of this study is to analyze the presence of morphophysiological damages and the metal concentration in samples of the lichen Parmotrema tinctorum, in urban and forest areas, checking for possible parameter variations between these areas, in the different matrices and seasons in the Southern region of Brazil. Six areas were selected (urban and forest), assigned to the rural/urban and urban/industrial matrices belonging to the watershed of the Rio dos Sinos, Brazil. The following parameters were analyzed: Index of photobiont vitality (IPV), photosynthetic pigments (chlorophyll, carotene, and pheophytin) and metals (Cu, Cr, Zn, Pb, and Ni) in the thallus of the lichen. The data were analyzed by an ANOVA one way, Pearson correlation test, and principal component analysis (PCA). Variations in the morphophysiological parameters were recorded in all the areas revealing significant differences. The lowest IPV values and highest concentration of metals were recorded in the urban environment, while low chlorophyll levels were found in the forest areas. The PCA showed a distinction between the areas and the season. The level of urbanization, vehicle traffic, and the weather conditions might have influenced the results. The use of P. tinctorum, the index of photobiont vitality and chlorophyll and pheophytin content, has proved to be an efficient tool to diagnose the air quality in the areas analyzed, allowing its use as a model in air monitoring studies, both in urban and forest areas, as well as in distinct matrices in the subtropical region.

Rapid Determination of Chlorophyll and Pheophytin in Green Tea Using Fourier Transform Infrared Spectroscopy.

The chlorophyll, pheophytin, and their proportions are critical factors to evaluate the sensory quality of green tea. This research aims to establish an effective method to determine the quantification of chlorophyll and pheophytin in green tea, based on Fourier transform infrared (FT⁻IR) spectroscopy. First, five brands of tea were collected for spectral acquisition, and the chlorophyll and pheophytin were measured using the reference method. Then, a relation between these two pigments and FT⁻IR spectroscopy were developed based on chemometrics. Additionally, the characteristic IR wavenumbers of these pigments were extracted and proved to be effective for a quantitative determination. Successively, non-linear models were also built based on these characteristic wavenumbers, obtaining coefficients of determination of 0.87, 0.80, 0.85 and 0.89; and relative predictive deviations of 2.77, 2.62, 2.26 and 3.07 for the four pigments, respectively. These results demonstrate the feasibility of FT⁻IR spectroscopy for the determination of chlorophyll and pheophytin.

Effects of nitrogen fertilizer and harvesting stage on photosynthetic pigments and phytol contents of Italian ryegrass silage.

This study aimed to investigate the effects of nitrogen (N) fertilizer application and harvesting stage on the contents of chlorophyll, phytol and carotenoids (ß-carotene and lutein) in Italian ryegrass herbage before and after ensiling, and the extent of phytol preservation after ensiling. Three rates of N fertilizer (0, 60 and 120 kg N/ha) were applied by top-dressing as an additional fertilizer. The herbage harvested at booting stage (27 weeks of age) or heading stage (29 weeks of age) were wilted for 1 day, then ensiled for 60 days using a small-scale pouch system. In the pre-ensiled herbages, increasing N fertilizer application increased the contents of crude protein and photosynthetic pigments, and these contents were also higher at the booting stage compared with the heading stage. In the silage, increasing N fertilizer application also increased the contents of crude protein, the photosynthetic pigments and their derivatives (pheophytin and pheophorbide), while harvesting stage did not affect the contents of ß-carotene, chlorophylls or pheophorbide. Nitrogen fertilizer application and early harvesting of herbage increased lutein and phytol contents in Italian ryegrass silage. Lutein and phytol in Italian ryegrass herbage are indicated to be well preserved during ensiling.

Phytoconstituents from Sidastrum micranthum (A. St.-Hil.) Fryxell (Malvaceae) and antimicrobial activity of pheophytin a

abstract Sidastrum micranthum (A. St.-Hil.) Fryxell, a member of the Malvaceae family, is called malva preta in Brazil. As this species is commonly used to treat bronchitis, cough, and asthma, better knowledge of its chemical compounds is important. The phytochemical study of its hexane extract, using chromatographic techniques, led to isolation of six compounds: the triterpene isoarborinol, a mixture of sitosterol and stigmasterol, sitosterol-3-O-β-D-glucopyranoside, pheophytin a, and 132-hydroxy-(132-S)-pheophytin a. Structural identification of these compounds was carried out using spectroscopic methods such as IR and 1D and 2D NMR (HOMOCOSY, HMQC, HMBC, and NOESY). Compounds isolated from S. micranthum were screened for their in vitro antifungal and antibacterial activity against twenty fungal and bacterial standard strains. Pheophytin a exhibited antimicrobial action against all microorganisms tested.

resumo Sidastrum micranthum (A. St.-Hil.) Fryxell, pertencente à família Malvaceae, é conhecida no Brasil como "malva preta". A espécie é popularmente usada contra bronquite, tosse e asma, mostrando a relevância de conhecer melhor sua composição química. O estudo fitoquímico do extrato hexânico da espécie, utilizando técnicas cromatográficas, conduziu ao isolamento de seis compostos: o triterpeno isoarborinol, mistura de sitosterol e estigmasterol, sitosterol-3-O-β-D-glicopiranosídeo, feofitina a e de 132-hidroxi-(132-S)-feofitina a. A identificação estrutural destes compostos foi realizada com base em métodos espectroscópicos, tais como IV, RMN 1D e 2D (HOMOCOSY, HMQC, HMBC e NOESY). As substâncias isoladas de Sidastrum micranthum foram avaliadas quanto às suas atividades antimicrobianas in vitro, contra vinte cepas fúngicas e bacterianas. A feofitina a mostrou ação antimicrobiana contra todos os microrganismos testados.

Extensive fragmentation of pheophytin-a by infrared multiphoton dissociation tandem mass spectrometry.

RATIONALE: In a previous tandem mass spectrometry (MS/MS) study on chlorophyll-a, infrared multiphoton dissociation (IRMPD) was demonstrated as a more effective fragmentation method than collision-induced dissociation (CID) and electron-induced dissociation (EID), where odd-electron product ions were observed ubiquitously in CID and IRMPD. To further understand the role of the macrocycle and the central Mg atom in the MS/MS process, the fragmentation behaviour of pheophytin-a, the Mg-free chlorophyll-a, was investigated. METHODS: CID, IRMPD, and EID were applied to the singly protonated pheophytin-a using an ultra-high-resolution Fourier transform ion cyclotron resonance (FTICR) mass spectrometer. The result is compared with the MS/MS study of chlorophyll-a. RESULTS: For pheophytin-a, some different fragmentation patterns from chlorophyll-a were obtained by all three MS/MS methods, but IRMPD still appears the most efficient method of generating product ions. The detection of odd-electron fragments in the CID and IRMPD spectra of protonated pheophytin-a suggests that the macrocyclic structure effectively stabilizes radicals, and these radical ions seem to have a relatively higher abundance in the presence of the central Mg atom. CONCLUSIONS: The strong absorption in the infrared region of pheophytin-a and secondary free radical rearrangement are proposed to explain the extensive frgmentation in IRMPD spectra. In addition, a comparison of the IRMPD spectra of chlorophyll-a and pheophytin-a shows that the macrocycle in the absence of the Mg atom is much more fragile.

High-performance liquid chromatography with fluorescence detection for the rapid analysis of pheophytins and pyropheophytins in virgin olive oil.

Pheophytins and pyropheophytin are degradation products of chlorophyll pigments, and their ratios can be used as a sensitive indicator of stress during the manufacturing and storage of olive oil. They increase over time depending on the storage condition and if the oil is exposed to heat treatments during the refining process. The traditional analysis method includes solvent- and time-consuming steps of solid-phase extraction followed by analysis by high-performance liquid chromatography with ultraviolet detection. We developed an improved dilute/fluorescence method where multi-step sample preparation was replaced by a simple isopropanol dilution before the high-performance liquid chromatography injection. A quaternary solvent gradient method was used to include a fourth strong solvent wash on a quaternary gradient pump, which avoided the need to premix any solvents and greatly reduced the oil residues on the column from previous analysis. This new method not only reduces analysis cost and time but shows reliability, repeatability, and improved sensitivity, especially important for low-level samples.

Identification and quantification of Cu-chlorophyll adulteration of edible oils.

Cu-pyropheophytin a, the major Cu-pigment of Cu-chlorophyll, was determined in edible oil by high-resolution mass spectrometry with a high-performance liquid chromatography-quadrupole (HPLC-Q)-Orbitrap system and by HPLC coupled with a photodiode-array detector. Respective limit of detection and limit of quantification levels of 0.02 µg/g and 0.05 µg/g were obtained. Twenty-nine commercial oil products marked as olive oil, grapeseed oil and blended oil, all sourced directly from a food company that committed adulteration with Cu-chlorophyll, were investigated. In this company, four green dyes illegally used in oils were seized during factory investigation by the health authorities. The food additive Cu-pyropheophytin a was found in all confiscated samples in concentrations between 0.02 and 0.39 µg/g. Survey results of another 235 commercial oil samples manufactured from other companies, including olive pomace oil, extra virgin olive oil, olive oil, grapeseed oil and blended oil, indicated high positive incidences of 63%, 39%, 44%, 97% and 8%, respectively, with a concentration range between 0.02 and 0.54 µg/g. High Cu-chlorophyll concentrations are indications for fraudulent adulteration of oils.

The 17-propionate esterifying variants of bacteriochlorophyll-a and bacteriopheophytin-a in purple photosynthetic bacteria.

Most purple photosynthetic bacteria contain bacteriochlorophyll(BChl)-a (a magnesium complex) and bacteriopheophytin(BPhe)-a (its free base) as their photoactive pigments. These pigments are composed of two parts: a cyclic tetrapyrrole as the chromophore and a long hydrocarbon-chain as the propionate-type esterifying group at the 17-position. The hydrocarbon-chain is usually an isoprenoid-type C20 phytyl (Phy) group in both the pigments. In the ester group of BChl-a, several variants such as geranylgeranyl (GG), dihydrogeranylgeranyl (DHGG) and tetrahydrogeranylgeranyl (THGG) groups were found in the final stage of BChl-a biosynthesis. On the other hand, the esterifying variants in BPhe-a have not been studied as much due to the lower levels of this pigment relative to BChl-a. The esterifying group does not affect the electronic absorption properties of such pigments in the monomeric state, but drastically alters the hydrophobicity. In this study, BChl-a and BPhe-a in the six phylogenetically distinct classes of purple bacteria were analyzed in terms of their esterifying groups in the 17-propionate residues, using high-performance liquid chromatography. Both BChls-a and BPhes-a carrying GG, DHGG and THGG in addition to the usual Phy were found for all the bacterial species studied at measurable levels. In some of the species, the ratio of BPhes-a esterified with GG, DHGG and THGG over the total BPhe-a drastically decreased in comparison with that of the corresponding BChls-a. Especially, the relative content of BPhe-a with GG largely decreased. This observation might indicate that BPhe-a as a cofactor of reaction centers was preferentially esterified with partially reduced and flexible chains (THGG and Phy) rather than less reduced and rigid ones (GG and DHGG).

Role of Rhodobacter sphaeroides photosynthetic reaction center residue M214 in the composition, absorbance properties, and conformations of H(A) and B(A) cofactors.

In the native reaction center (RC) of Rhodobacter sphaeroides, the side chain of (M)L214 projects orthogonally toward the plane and into the center of the A branch bacteriopheophytin (BPhe) macrocycle. The possibility that this side chain is responsible for the dechelation of the central Mg(2+) of bacteriochlorophyll (BChl) was investigated by replacement of (M)214 with residues possessing small, nonpolar side chains that can neither coordinate nor block access to the central metal ion. The (M)L214 side chain was also replaced with Cys, Gln, and Asn to evaluate further the requirements for assembly of the RC with BChl in the HA pocket. Photoheterotrophic growth studies showed no difference in growth rates of the (M)214 nonpolar mutants at a low light intensity, but the growth of the amide-containing mutants was impaired. The absorbance spectra of purified RCs indicated that although absorbance changes are associated with the nonpolar mutations, the nonpolar mutant RC pigment compositions are the same as in the wild-type protein. Crystal structures of the (M)L214G, (M)L214A, and (M)L214N mutants were determined (determined to 2.2-2.85 Å resolution), confirming the presence of BPhe in the HA pocket and revealing alternative conformations of the phytyl tail of the accessory BChl in the BA site of these nonpolar mutants. Our results demonstrate that (i) BChl is converted to BPhe in a manner independent of the aliphatic side chain length of nonpolar residues replacing (M)214, (ii) BChl replaces BPhe if residue (M)214 has an amide-bearing side chain, (iii) (M)214 side chains containing sulfur are not sufficient to bind BChl in the HA pocket, and (iv) the (M)214 side chain influences the conformation of the phytyl tail of the BA BChl.

Determination of chlorophylls in Taraxacum formosanum by high-performance liquid chromatography-diode array detection-mass spectrometry and preparation by column chromatography.

Taraxacum formosanum, a well-known Chinese herb shown to be protective against hepatic cancer as well as liver and lung damage, may be attributed to the presence of abundant carotenoids and chlorophylls. However, the variety and content of chlorophylls remain uncertain. The objectives of this study were to develop an high-performance liquid chromatography-diode array detection-mass spectrometry method for determination of chlorophylls in T. formosanum and preparation by column chromatography. An HyPURITY C18 column and a gradient mobile phase of water (A), methanol (B), acetonitrile (C), and acetone (D) could resolve 10 chlorophylls and an internal standard Fast Green FCF within 30 min with a flow rate at 1 mL/min and detection at 660 nm. Both chlorophylls a and a' were present in the largest amount (1389.6 µg/g), followed by chlorophylls b and b' (561.2 µg/g), pheophytins a and a' (31.7 µg/g), hydroxychlorophyll b (26.5 µg/g), hydroxychlorophylls a and a' (9.8 µg/g), and chlorophyllides a and a' (0.35 µg/g). A glass column containing 52 g of magnesium oxide-diatomaceous earth (1:3, w/w) could elute chlorophylls with 800 mL of acetone containing 50% ethanol at a flow rate of 10 mL/min. Some new chlorophyll derivatives including chlorophyllide b, pyropheophorbide b, hydroxypheophytin a, and hydroxypheophytin a' were generated during column chromatography but accompanied by a 63% loss in total chlorophylls. Thus, the possibility of chlorophyll fraction prepared from T. formosanum as a raw material for future production of functional food needs further investigation.

Isolation and pigment composition of the reaction centers from purple photosynthetic bacterium Rhodopseudomonas palustris species.

The reaction centers (RCs) from several species of a purple photosynthetic bacterium, Rhodopseudomonas palustris, were first isolated by ammonium-sulfate fractionation of the isolated core complexes, and were successfully purified by anion-exchange and gel-filtration chromatography as well as sucrose-density gradient centrifugation. The RCs were characterized by spectroscopic and biochemical analyses, indicating that they were sufficiently pure and had conserved their redox activity. The pigment composition of the purified RCs was carefully analyzed by LCMS. Significant accumulation of both bacteriochlorophyll(BChl)-a and bacteriopheophytin(BPhe)-a esterified with various isoprenoid alcohols in the 17-propionate groups was shown in RCs for the first time. Moreover, a drastic decrease in BPhe-a with the most dehydrogenated and rigid geranylgeranyl(GG) ester was observed, indicating that BPhe-a in RC preferably took partially hydrogenated and flexible ester groups, i.e. dihydro-GG and tetrahydro-GG in addition to phytyl. Based on the reported X-ray crystal structures of purple bacterial RCs, the meaning of flexibility of the ester groups in BChl-a and BPhe-a as the cofactors of RCs is proposed.

A valuable visual colorimetric and electrochemical biosensor for porphyrin.

Porphyrin is able to specifically combine with phosphorus, thus a novel bifunctional sensing platform for determination of porphyrin by visual colorimetry and electrochemistry was demonstrated. A pretreated gold sheet (or electrode) with 2-mercatpoethanol (2-ME) was chemically modified by POCl(3) to obtain the surface phosphate active sites. The different stages of modified electrode were characterized by electrochemical impedance spectroscopy (EIS). The 1:1 cationic sitting-atop (SAT) complex P(V)-porphyrin was formed due to the high affinity of the modified gold sheet (or electrode) towards the porphyrin, resulting in electron transfer resistance increase of the electrode surface. Meanwhile, a dramatic color changing from burgundy to dark green of porphyrin solution was observed with the naked-eye within 3s. What's more, this was reflected by the notable change of the Soret band of porphyrin when using UV-vis. Two sensing systems provide different sensitivity for porphyrin analysis. With visual colorimetry, porphyrin can be detected at a level of 1.0×10(-6) M, whereas the detection limit of porphyrin is 3.0×10(-8) M using the EIS method. The practical application of the sensor to determination of pheophytin which was obtained from fresh spinach leaves has been accomplished. The results demonstrate the facility and effectivity of our introduced bifunctional biosensor for quantitative analysis of porphyrin.

Color measurements as a reliable method for estimating chlorophyll degradation to phaeopigments.

The application of biocides is a traditional method of controlling biodecay of outdoor cultural heritage. Chlorophyll degradation to phaeopigments is used to test the biocidal efficacy of the antimicrobial agents. In the present study, the usefulness of color measurements in estimating chlorophyll degradation was investigated. An aeroterrestrial stone biofilm-forming cyanobacterium of the genus Nostoc was chosen as test organism, comparing its different behaviour in both planktonic and biofilm mode of growth against the isothiazoline biocide Biotin T®. Changes in A(435 nm)/A(415 nm) and A(665 nm)/A(665a nm) and in the chlorophyll a and adenosine triphosphate (ATP) cell content were compared with the variations in the CIELAB color parameters (L*, a*, b*, C*(ab) and h(ab)). Our findings showed that both the phaeophytination indexes are useful in describing degradation of chlorophyl a to phaeopigments. Moreover, the CIELAB color parameters represented an effective tool in describing chlorophyll degradation. L* CIELAB parameter appeared to be the most informative parameter in describing the biocidal activity of Biotin T® against Nostoc sp. in both planktonic and biofilm mode of growth.

Defect in non-yellow coloring 3, an alpha/beta hydrolase-fold family protein, causes a stay-green phenotype during leaf senescence in rice.

Chlorophyll degradation is an important phenomenon in the senescence process. It is necessary for the degradation of certain chlorophyll-protein complexes and thylakoid membranes during leaf senescence. Mutants retaining greenness during leaf senescence are known as 'stay-green' mutants. Non-functional type stay-green mutants, which possess defects in chlorophyll degradation, retain greenness but not leaf functionality during senescence. Here, we report a new stay-green mutant in rice, nyc3. nyc3 retained a higher chlorophyll a and chlorophyll b content than the wild-type but showed a decrease in other senescence parameters during dark incubation, suggesting that it is a non-functional stay-green mutant. In addition, a small amount of pheophytin a, a chlorophyll a-derivative without Mg(2+) ions in its tetrapyrrole ring, accumulated in the senescent leaves of nyc3. nyc3 shows a similar but weaker phenotype to stay green (sgr), another non-functional stay-green mutant in rice. The chlorophyll content of nyc3 sgr double mutants at the late stage of leaf senescence was also similar to that of sgr. Linkage analysis revealed that NYC3 is located near the centromere region of chromosome 6. Map-based cloning of genes near the centromere is very difficult because of the low recombination rate; however, we overcame this problem by using ionizing radiation-induced mutant alleles harboring deletions of hundreds of kilobases. Thus, it was revealed that NYC3 encodes a plastid-localizing alpha/beta hydrolase-fold family protein with an esterase/lipase motif. The possible function of NYC3 in the regulation of chlorophyll degradation is discussed.

Spatial patterns of macrobenthic communities in shallow-water tidal embayments and their association with environmental factors.

This study examines the distribution, composition, and structure of benthic communities in nine shallow-water semienclosed embayments on Cape Cod, southeastern Massachusetts. The spatial patterns were used to associate benthic characteristics with local environmental factors. Field data from multiple years were collected to measure macrofaunal abundance, community composition, and environmental characteristics. Multivariate statistics were used to analyze the spatial variations of species composition and the abundance of benthic macrofauna. Canonical ordination, specifically redundancy analysis, was used to determine the relative importance of the environmental factors (nutrients and habitat factors) being studied. The analyses demonstrate that the benthic communities in the shallow tidal embayments are clearly clustered in association with the local environments. Species composition and community structure of the benthic macrofauna are significantly correlated with water column parameters, especially with phytoplankton biomass, total nitrogen, and dissolved oxygen.

Carotenoids in sea buckthorn ( Hippophae rhamnoides L.) berries during ripening and use of pheophytin a as a maturity marker.

Four cultivars of sea buckthorn berries were analyzed for their carotenoid and chlorophyll contents during ripening in three consecutive years. The different carotenoids generally increased in concentration during ripening and comprised from 120 to 1425 microg/g of DW of total carotenoids (1.5-18.5 mg/100 g of FW) depending on cultivar, harvest time, and year. GLM analyses revealed the effect of cultivar to be considerably larger than that of year and harvest time. The content of pheophytin a, a chlorophyll a derivate, steadily decreased during berry ripening. Pheophytin a therefore acted as a marker of the degree of ripening of sea buckthorn berries and was used here to convert harvest date into an estimated ripening time.