(n-7) and (n-9) cis-Monounsaturated fatty acid contents of 12 Brassica species

cis-Vaccenic acid or cis-11-octadecenoic acid, a C18:1 (n-7) isomer of oleic acid (C18:1 (n-9)) has been found in several oilseeds. It is synthesized from palmitic acid (C16:0) via production of C16:1 (n-7) by a Delta9 desaturase and elongation by an elongase giving C18:1 (n-7). In this study, the fatty acid composition of 12 Brassica species was analyzed by GC-FID and confirmed by GC-MS. All species contained C18:1 (n-7), C20:1 (n-7) and C22:1 (n-7) fatty acid isomers, suggesting that C18:1 (n-7) was elongated. The levels of these fatty acids varied according to the species. C18:1(n-7)) represented from 0.4% to 3.3% of the total relative fatty acid contents of the seeds. The contents of C20:1(n-7) and C22:1(n-7) levels were lower than C18:1(n-7) contents; the relative fatty acid composition varied from 0.02% to 1.3% and from below the limit of detection to 1.3% for C20:1 (n-7) and C22:1 (n-7), respectively. The ratios of (n-7)/(n-9) ranged from 2.8% to 16.7%, 0.6% to 29.5% and 0% to 2.6% for C18:1, C20:1 and C22:2, respectively. Using statistical similarities or differences of the C18:1 (n-7)/(n-9) ratios for chemotaxonomy, the surveyed species could be arranged into three groups. The first group would include Brassica napus, B. rapa, and B. tournefortii with Eruca sativa branching only related to B. napus. The second group would include B. tournefortii, Raphanus sativus and Sinapis alba. The last group would include B. juncea, B. carinata and B. nigra with no similarity/relationship between them and between the other species. Results suggested that the level of C20:1 (n-7) influenced the levels of all monounsaturated fatty acids with chain length higher than 20 carbons. On the other hand, palmitoleic acid (C16:1) levels, C16:1 being the parent of all (n-7) fatty acids, had no statistically significant correlation with the content of any of the fatty acids of the (n-7) or (n-9) family.     

Polar lipid and fatty acid profiles – Re-vitalizing old approaches as a modern tool for the classification of mycoplasmas?

A set of 20 Mollicutes strains representing different lines of descent, including the type species of the genus Mycoplasma, Mycoplasma mycoides, Acholeplasma laidlawii and a strain of Mesoplasma, were subjected to polar lipid and fatty acid analyses in order to evaluate their suitability for classification purposes within members of this group. Complex polar lipid and fatty acid profiles were detected for each examined strain. All strains contained the polar lipids phosphocholine-6'-alpha-glucopyranosyl-(1'-3)-1, 2-diacyl-glycerol (MfGL-I), 1-O-alkyl/alkenyl-2-O-acyl-glycero-3-phosphocholine (MfEL), sphingomyelin (SphM), 1-O-alkyl/alkenyl-glycero-3-phosphocholine (lysoMfEL), the unknown aminophospholipid APL1 and the cholesterol Chol2. A total of 19 strains revealed the presence of phosphatidylethanolamine (PE) and/or phosphatidylglycerol (PG), and the presence of diphosphatidylglycerol (DPG) was detected in 13 strains. The unknown aminolipid AL1 was found in the extracts of 17 strains. Unbranched saturated and unsaturated compounds predominated in the fatty acid profiles. Major fatty acids were usually C16:0, C18:0, C18:1 omega9c and 'Summed feature 5' (C18:2 omega6, 9c/C18:0 anteiso). Our results demonstrated that members of the M. mycoides cluster showed rather homogenous polar lipid and fatty acid profiles. In contrast, each of the other strains was characterized by a unique polar lipid profile and significant quantitative differences in the presence of certain fatty acids. These results indicate that analyses of both polar lipid and fatty acid profiles could be a useful tool for classification of mycoplasmas.     

Differential regulation of fatty acid biosynthesis in two Chlorella species in response to nitrate treatments and the potential of binary blending microalgae oils for biodiesel application

This study was undertaken to investigate the effects of different nitrate concentrations in culture medium on oil content and fatty acid composition of Chlorella vulgaris (UMT-M1) and Chlorella sorokiniana (KS-MB2). Results showed that both species produced significant higher (p<0.05) oil content at nitrate ranging from 0.18 to 0.66 mM with C. vulgaris produced 10.20-11.34% dw, while C. sorokiniana produced 15.44-17.32% dw. The major fatty acids detected include C16:0, C18:0, C18:1, C18:2 and C18:3. It is interesting to note that both species displayed differentially regulated fatty acid accumulation patterns in response to nitrate treatments at early stationary growth phase. Their potential use for biodiesel application could be enhanced by exploring the concept of binary blending of the two microalgae oils using developed mathematical equations to calculate the oil mass blending ratio and simultaneously estimated the weight percentage (wt.%) of desirable fatty acid compositions.     

Optimization of growth and fatty acid composition of a unicellular marine picoplankton,Nannochloropsis sp., with enriched carbon sources

A unicellular marine picoplankton, Nannochloropsis sp., was grown under CO₂-enriched photoautotrophic or/and acetate-added mixotrophic conditions. Photoautotrophic conditions with enriched CO₂ of 2800 l CO₂ l^–1 and aeration gave the highest biomass yield (634 mg dry wt l^–1), the highest total lipid content (9% of dry wt), total fatty acids (64 mg g^–1 dry wt), polyunsaturated fatty acids (35% total fatty acids) and eicosapentaenoic acid (EPA, 20:53) (16 mg g^–1 dry wt or 25% of total fatty acids). Mixotrophic cultures gave a greater protein content but less carbohydrates. Adding sodium acetate (2 mM) decreased the amounts of the total fatty acids and EPA. Elevation of CO₂ in photoautotrophic culture thus enhances growth and raises the production of EPA in Nannochloropsis sp.     

Seasonal changes in lipid content and composition of the polychaete Nereis (Hediste) diversicolor

The lipid content and composition of Nereis (Hediste) diversicolor O. F. Müller (Annelida, Polychaeta, Nereidae) a mud-dwelling, intertidal errant polychaete in the Tagus estuary (Portugal), were examined on the monthly basis by lipid extraction, TLC and capillary GC. In this estuary, N. diversicolor is by far the dominant species among polychaeta and the main food item in the natural diet of several flatfishes. The biochemical elucidation of its lipid structure and distribution throughout the year, described in this study, provides information not only about the physiological role of lipids in the animal under consideration but also about dietary fatty acid requirements of some flatfishes in the wild and under laboratory conditions.The total lipid content varied between a maximum of 19.3% lyophilized dry weight in February (4.4% fresh weight) and a minimum of 6.6% in August (1.9% fresh weight). The major lipid classes were triacylglycerol, phospholipid, free sterol, free fatty acid, sterol ester/wax ester and alkyldiacylglycerol.The fatty acid composition was rather unsaturated with a 1:2 mean ratio of n-3: n-6. The major fatty acids were C160:0, C18:1n-9, C18:2n-6, and C20:5n-3; there were smaller amounts of C180:0, C18:1n-11, C18:1n-7, C18:3n-3, C20:1, C20:2n-6, C20:4n-6, C22:2, C22:5n-3, and many other fatty acids were detected at trace levels. The unsaturation ranged from 36.9 mg/g dry weight in summer to 107.4 mg/g in winter. An accumulation of fatty acids from plant origin was evident, in particular linoleic acid (C18:2n-6), which was quantitatively one of the major fatty acids throughout the year.     

Structure of the lipopolysaccharide core of Vibrio vulnificus type strain 27562

The structure of the lipopolysaccharide core of Vibrio vulnificus type strain 27562 is presented. LPS hydrolysis gave two oligosaccharides, OS-1 and OS-2, as well as lipid A. NMR spectroscopic data corresponded to the presence of one Kdo residue, one β-glucopyranose, three heptoses, one glyceric acid, one acetate, three PEtN, and one 5,7-diacylamido-3,5,7,9-tetradeoxynonulosonic acid residue (pseudaminic acid, Pse) in OS1. OS2 differed form OS 1 by the absence of glyceric acid, acetate, and Pse residues. Lipid A was analyzed for fatty acid composition and the following fatty acids were found: C14:0, C12:0-3OH, C16:0, C16:1, C14:0-3OH, C18:0, C18:1 in a ratio of 1:3:3:1:2.5:0.6:0.8.     

Isolation and mass spectrometry characterization of molecular species of lactosylceramides using liquid chromatography-electrospray ion trap mass spectrometry

Reverse-phase liquid chromatography/electrospray ion trap mass spectrometry (LC-ESI-MSn) was established for identification of the molecular species of lactosylceramides. Lactosylceramides derived from porcine blood cells were separated on a CapcellPak C8 column using a mixture of methanol and 1 mM ammonium formate from the C16 to C26 fatty acyl chains based on the length of total carbon chains and the nature of sphingoid bases (w') and fatty acyl chains (Y0'-w') was identified by MS3 as their [M+H]+ ions. The same number of fatty acyl moieties appeared in the order of unsaturated, (2-)hydroxylated, and saturated components. The molecular species of lactosylceramides derived from porcine blood cells totaled more than 33 and included mainly C24:0-d18:1, Ch24:0-d18:1, Ch24:1-d18:1, C24:1-d18:1, and C22:0-d18:1 in addition to 28 minor species from C16:0 to C26:0 fatty acyl moieties. The molecular species of lactosylceramides in the membrane microdomain fraction of HL-60 cells (70% were differentiated into macrophage-lineage cells) were identified as C24:0-d18:1, C24:1-d18:1, C22:0-d18:1, C16:0-d18:1, and more than 21 other minor species. Our results suggest that reverse-phase LC-ESI-MSn is a useful and simple method for identification of lactosylceramide molecular species.     

Feeding stimulants eliciting the probing behavior for Peregrinator blannulipes Montrouzier et Signore (Hemiptera: Ruduviidae) from Tribolium confusum (Jacquelin du Val)

Four fatty acid methyl esters identified in the solvent extract of Tribolium confusum (Jacquelin du Val) larvae as kairomones were individually and collectively tested for probing behavior of Peregrinator biannulipes Montrouzier et Signoret. All identified fatty acid methyl eaters, methyl palmitate, methyl linolate, methyl oleate and methyl stearate, exhibited characterisitic kairomonal probing behavior of P. biannulipe toward the lure. These fatty acid methyl ester were active at 0.2 microg/lure but a synergistic effect was not observed among them. Commercially available C8-C14 even-numbered fatty acid methyl esters that were not detected in the extract of T. confusum larvae also elicited a probing behavior but their activities were weaker than those of four fatty acid methyl ester (C16:0, C18:0, C18:1 and C18:2) identified in the extract. On the other hand, C17 and C19 odd-numbered fatty acid methyl esters did not show any activity at all.     

Synthesis in vitro of very long chain fatty acids in Vibrio sp. strain ABE-1

The activity of fatty acid synthetase (FAS) from Vibrio sp. strain ABE-1 required the presence of acyl carrier protein and was completely inhibited by thiolactomycin, an inhibitor specific for a type II FAS. These observations indicate that this enzyme is a type II FAS. Analysis by gas-liquid chromotography of the reaction products synthesized in vitro from [2-¹⁴C]malonyl-CoA by the partially purified FAS revealed, in addition to 16-and 18-carbon fatty acids which are normal constituents of this bacterium, the presence of fatty acids with very long chains. These fatty acids were identified as saturated and mono-unsaturated fatty acids with 20 up to as many as 30 carbon atoms. The longest fatty acids normally found in this bacterium contain 18-carbon atoms. These results suggest that the FAS from Vibrio sp. strain ABE-1 has potentially the ability to synthesize fatty acids with very long chains.Abbreviations ACP acyl carrier protein - FAME fatty acid methyl ester - FAS fatty acid synthetase - FID flame ionization detection - GLC gas-liquid chromatography - TLC thin-layer chromatography - In designations of fatty acids, such as 16:0, 16:1, etc the colon separates the number that denotes the number of carbon atoms and the number that denotes the number of double bonds, respectively, in the molecule - 16:0-CoA CoA ester of 16:0     

Deep-water shrimp (Pandalus borealis, Krøyer 1838) as indicator organism for fish-farm waste

A feeding experiment was carried out with deep-water shrimp (Pandalus borealis), a common benthic inhabitant of Norwegian fjords. Shrimp were reared in tanks for three months, fed either salmon feed or cod, and the fatty acid profile of their muscle tissue was monitored. The salmon feed pellets and cod had completely different fatty acid compositions, with significant differences in most of the analysed fatty acids, eight of them, 14:0, iso16:0, 16:1n7, 16:2n11, 18:3n3, 20:0, 22:0 and 22:1n11, differing between the two diets by a factor of 10 or more. The levels of the fatty acids in the diets differed also substantially from the levels in the shrimp tissue at the start of the experiment. The shrimp were fed ad libitum and both groups increased significantly in total fatty acid content in the course of the experiment. Nevertheless, the fatty acids made up no more than 6.5 mg g^− 1 muscle tissue, with 88 ± 1% of them bound in polar, membrane lipids. Their composition in the tissue did also change to a certain extent, but the resulting tissue composition did not reflect the fatty acid composition of the diets. Only two fatty acids, 18:2n6 and 18:3n3, clearly followed the dietary level. They are abundant in commercial salmon feed since they are rich in plant oils used as additives to the commercial feed. Determination of fatty acid levels in shrimp tissue may enable shrimp to be used as indicator organisms for the influence of organic fish-farm waste on the food web in adjacent areas.     

Structural elucidation of fungal polysaccharides isolated from the cell wall of Plectosphaerella cucumerina and Verticillium spp

The structure of acidic fungal polysaccharides isolated from the cell wall of Plectosphaerella cucumerina, Verticillium dahliae, and V. albo-atrum has been investigated by chemical analysis, methylation analysis, and 1D and 2D 1H and 13C NMR spectroscopy. The polysaccharides have an idealized repeating block of the type: [carbohydrates: see text] linked to a small mannan core (<15%), where n=13, m=13, p=5, and q=8 for P. cucumerina, and n=16, m=16, p=6, and q <1 for both Verticillium species.     

Structural analysis of an acidic, fatty acid ester-bonded extracellular polysaccharide produced by a pristane-assimilating marine bacterium, Rhodococcus erythropolis PR4

Rhodococcus erythropolis PR4 is a marine bacterium that can degrade various alkanes including pristane, a C(19) branched alkane. This strain produces a large quantity of extracellular polysaccharides, which are assumed to play an important role in the hydrocarbon tolerance of this bacterium. The strain produced two acidic extracellular polysaccharides, FR1 and FR2, and the latter showed emulsifying activity toward clove oil, whereas the former did not. FR2 was composed of D-galactose, D-glucose, D-mannose, D-glucuronic acid, and pyruvic acid at a molar ratio of 1:1:1:1:1, and contained 2.9% (w/w) stearic acid and 4.3% (w/w) palmitic acid attached via ester bonds. Therefore, we designated FR2 as a PR4 fatty acid-containing extracellular polysaccharide or FACEPS. The chemical structure of the PR4 FACEPS polysaccharide chain was determined by 1D (1)H and (13)C NMR spectroscopies as well as by 2D DQF-COSY, TOCSY, HMQC, HMBC, and NOESY experiments. The sugar chain of PR4 FACEPS was shown to consist of tetrasaccharide repeating units having the following structure: [structure: see text].     

Cryptophyceae and rhodophyceae; chemotaxonomy, phylogeny, and application

The biochemical compositions of seven strains of marine cryptomonad and a rhodophyte were determined in logarithmic phase batch (1.4 L flask) and semi-continuous (10 L carboy) culture. Lipid ranged from 13% to 28%, protein ranged from 53% to 68%, and carbohydrate ranged from 9% to 24% of the organic weight. The major lipid classes in the species examined were polar lipids (78-88% of total lipid). The major sterol in the Cryptophyceae and the Rhodophyceae was 24-methylcholesta-5,22E-dien-3beta-ol (62-99% of total sterols); which is also the major sterol in some diatoms and haptophytes. Smaller proportions of cholest-5-en-3beta-ol (1-17.7%) were also found in the Cryptophyceae. Most cryptomonads contained high proportions of the n-3 polyunsaturated fatty acids (PUFA), 18:3n-3 (20.7-29.9% of the total fatty acids), 18:4n-3 (12.5-30.2%), 20:5n-3 (7.6-13.2%) and 22:6n-3 (6.4-10.8%). However, the blue-green cryptomonad Chroomonas placoidea was characterized by a low proportion of 22:6n-3 (0.2% of total fatty acids), and a significant proportion of 22:5n-6 (4.5%), and the presence of 24-ethylcholesta-5,22E-dien-3beta-ol (35.5% of total sterols). The fatty acid composition of the rhodophyte Rhodosorus sp. was similar to those of the Cryptophyceae except for lower proportions of 18:4n-3 and lack of C21 and C22 PUFA. It is postulated that the primary endosymbiosis of a photosynthetic n-3 C18 PUFA-producing prokaryote and a eukaryotic host capable of chain elongation and desaturation of exogenous PUFA, resulted in the Rhodophyceae capable of producing n-3 C20 PUFA. The secondary endosymbiosis of a photosynthetic n-3 C20 PUFA-producing eukaryote (such as a Rhodosorus sp. like-rhodophyte) and a eukaryotic host capable of further chain elongation and desaturation, resulted in the Cryptophyceae being capable of producing n-3 C20 and C22 PUFA de novo. Selected isolates were examined further in feeding trials with juvenile Pacific oysters (Crassostrea gigas). Rhodomonas salina CS-24(containing elevated 22:6n-3) produced high growth rates in oysters; equivalent to the microalga commonly used in aquaculture, Isochrysis sp. (T.ISO).     

The desaturase from Bacillus subtilis, a promising tool for the selective olefination of phospholipids

The Delta5-desaturase from Bacillus subtilis has been cloned in Escherichia coli BL21 cells and its enzyme activity has been investigated as a function of temperature and oxygenation by analyzing methyl ester adducts from the total lipid extract in GC-MS measurements. The present data bring out that the activity of recombinant Delta5-desaturase, at 20-22 degrees C and 20% oxygen, is surprisingly high yielding 22% of C16:1,Delta5 (5-cis-palmitoleic acid) and 13% C18:2, Delta5 Delta11 (efedrenic acid). Lower amounts of other mono- and doubly-Delta5-unsaturated fatty acids were also detected. These findings demonstrate that Delta5-desaturase can accept a multiplicity of substrates and is endowed with an unprecedented activity among other acyl-lipid desaturases thus representing a unique tool for the production of rare Delta5 unsaturated fatty acid derivatives.     

Composition of fatty acids triacylglycerols and unsaponifiable matter in Calophyllum calaba L. oil from Guadeloupe

The composition of the kernel oils of two Calophyllum species (Calophyllum calaba L. and Calophyllum inophyllum L.) was investigated. The physico-chemical properties and fatty acid composition of the kernel oils were examined. In two species, oleic acid C18:1 (39.1-50%) is the dominating fatty acid followed by linoleic acid C18:2 (21.7-31.1%) as the second major fatty acid. Stearic C18:0 (13.4-14.3%) and palmitic C16:0 (11-13.7%) acids are the major saturates. The oils contains an appreciable amount of unsaturated fatty acids (70.8-73.10%). Most of the fatty acids are present as triacylglycerol (76.7-84%), twenty one triacylglycerols are detected with predominantly unsaturated triacylglycerols. The total unsaponifiable content, its general composition and the identity of the components of the sterol and tocopherol fractions are presented. In both species, analysis of the unsaponifiable fractions revealed the preponderance of phytosterols, mainly stigmasterol (35.8-45.1%) and beta-sitosterol (41.1-43.1%). Among the eight tocopherols and tocotrienols present in two species, variations exist; alpha-tocopherol (183 mg/kg) is the main tocopherol in Calophyllum calaba L. and Delta-tocotrienol (236 mg/kg) is the dominant tocotrienol in Calophyllum inophyllum L.     

Changes in fatty acid profiles of thermo-intolerant and thermo-tolerant marine diatoms during temperature stress

Fatty acid composition and degree of fatty acid saturation during temperature stress in thermo-intolerant (Phaeodactylum tricornutum) and thermo-tolerant (Chaetoceros muelleri) marine diatoms were investigated. A greater number of fatty acids were observed in C. muelleri than in P. tricornutum regardless of treatment. The major fatty acids detected were 14:0, 16:0, 16:1, 16:2, 16:3, 18:0, 18:1(n-9)c, 18:2(n-6) and 20:5(n-3) with additional fatty acids 18:1(n-9)t and 20:4(n-6) detected in C. muelleri. Short duration (2 h) temperature increase above optimal growth temperature had a greater effect on fatty acid composition in C. muelleri than in P. tricornutum and the degree of fatty acid saturation was affected more by temperature in C. muelleri than in P. tricornutum during both short and long duration (24 h) treatments. Total protein assay results suggest that P. tricornutum, but not C. muelleri, was undergoing stress under our growing conditions although lipids in both diatoms were affected by increased temperature. Immunodetection of proteins with anti-rubisco indicates that the rubisco large subunit was undergoing greater turnover in C. muelleri than in P. tricornutum. However, the integrity of rubisco as a suitable indicator of lipid status needs further study. This work supports the hypothesis that a particular temperature, and not treatment duration, has the greater effect on changes in fatty acid composition. Furthermore, changes in fatty acid composition and degree of fatty acid saturation occurred more quickly in the diatoms in response to increased temperature than previously observed in nutrient starvation studies. Since diatom lipids represent an important resource for growth and reproduction of marine animals, the rapid alteration of their lipid composition under temperatures normally encountered in marine environments warrants further study.     

The effect of phosphate starvation on the lipid and fatty acid composition of the fresh water eustigmatophyte Monodus subterraneus

Phosphate limitation caused significant changes in the fatty acid and lipid composition of Monodus subterraneus. With decreasing phosphate availability from 175 to 52.5, 17.5 and 0 microM (K2HPO4), the proportion of the major VLC-PUFA, eicosapentaenoic acid (EPA), gradually decreased from 28.2 to 20.8, 19.4 and 15.5 mol% (of total fatty acids), respectively. The cellular total lipid content of starved cells increased, mainly due to the dramatic increase in triacylglycerols (TAG) levels. Among polar lipids, cellular contents of digalactosyldiacylglycerol (DGDG) and diacylglyceroltrimethylhomoserine (DGTS) increased sharply from 0.29 and 0.19 to 0.60 and 0.38 fg cell(-1), respectively, while that of monogalactosyldiacylglycerol (MGDG) was not significantly changed. In the absence of phosphate, the proportion of phospholipids was significantly reduced from 8.3% to 1.4% of total lipids, and the proportion of triacylglycerols (TAG) increased from 6.5% up to 39.3% of total lipids. The share of MGDG was substantially reduced, from 35.7% to 13.3%, while that of DGDG and DGTS reduced less from 18.3% to 15.1%, and 12.2% to 8.6%, respectively. The most distinctive change in the fatty acid composition was noted in that of DGDG, where the proportion of EPA, located exclusively at the sn-1 position, increased from 11.3% to 21.5% at the expense of 16:0, 16:1 and 18:1. In MGDG, however, the proportion of EPA did not change appreciably. In contrast to higher plants, DGDG accumulated under P-deprivation in M. subterraneus, did not resemble PC and the positional distribution of its fatty acids was not altered, preserving the C20/C16 structure of its molecular species. We suggest that under phosphate starvation DGTS is a likely source of C20 acyl groups that can be exported to the sn-1 position of DGDG and can partially compensate for the decrease in PE, the apparent source of C20 acyl-containing diacylglycerols in this alga. Moreover, accumulation of non-esterified 18:0 indicates that no polar lipid can replace PC, which appears to be the only lipid capable of C18 desaturation in this alga.     

Neoscardovia arbecensis gen. nov., sp. nov., isolated from porcine slurries

Three Gram-positive, anaerobic, pleomorphic strains (PG10(T), PG18 and PG22), were selected among five strains isolated from pig slurries while searching for host specific bifidobacteria to track the source of fecal pollution in water. Analysis of the 16S rRNA gene sequence showed a maximum identity of 94% to various species of the family Bifidobacteriaceae. However, phylogenetic analyses of 16S rRNA and HSP60 gene sequences revealed a closer relationship of these strains to members of the recently described Aeriscardovia, Parascardovia and Scardovia genera, than to other Bifidobacterium species. The names Neoscardovia gen. nov. and Neoscardovia arbecensis sp. nov. are proposed for a new genus and for the first species belonging to this genus, respectively, and for which PG10(T) (CECT 8111(T), DSM 25737(T)) was designated as the type strain. This new species should be placed in the Bifidobacteriaceae family within the class Actinobacteria, with Aeriscardovia aeriphila being the closest relative. The prevailing cellular fatty acids were C(16:0) and C(18:1)ω9c, and the major polar lipids consisted of a variety of glycolipids, diphosphatidyl glycerol, two unidentified phospholipids, and phosphatidyl glycerol. The peptidoglycan structure was A1γmeso-Dpm-direct. The GenBank accession numbers for the 16S rRNA gene and HSP60 gene sequences of strains PG10(T), PG18 and PG22 are JF519691, JF519693, JQ767128 and JQ767130, JQ767131, JQ767133, respectively.     

Identification of the steroid fatty acid ester conjugates formed in vivo in Mytilus edulis as a result of exposure to estrogens

Vertebrate-type sex steroids have been detected in a number of mollusk species and may play a role in the reproductive physiology of the animal. Mollusks are also exposed to exogenous estrogenic steroids that are present in sewage effluents, and these may add to the estrogenic burden of exposed animals. We investigated the uptake of estrogens in the blue mussel, Mytlius edulis and report for the first time the identity of estrogen fatty acid ester metabolites formed in vivo in an invertebrate. We exposed mussels to waterborne radiolabeled [(14)C]-17beta-estradiol (E2) or estrone (E1) and determined the nature of their metabolites using radio-HPLC and mass spectrometry (MS). After 13 days of exposure to 10ng/L E2, concentrations of radiolabeled residues were 2428-fold higher in M. edulis soft tissues compared with the ambient water concentration of E2. All the E2 residues in the mussel were present as a lipophilic ester which, in depuration studies, had a half-life of 8.3 days. Exposure of mussels to [(14)C]-E1 (70ng/L) resulted in formation of a similar lipophilic metabolite that after hydrolysis released [(14)C]-E2. Tandem MSMS analyses of the purified steroid ester fraction isolated from mussels exposed to either E2 or E1 revealed that they had the same composition and comprised C16:0, C16:1 and C16:2 esters of E2. This work reveals that in vivo E1 is rapidly metabolized to E2 in mussels prior to conjugation to C16 fatty acid esters, proving that C17-ketoreductase and C16 fatty acid acyl-CoA:E2 acyltransferase are important enzymes for the metabolism of estrogens in M. edulis.