Species specificity in the biosynthesis of branched paraffins in leaves

Isobutyrate-1-(14)C and l-isoleucine-U-(14)C fed through the petiole labeled the surface lipids of broccoli leaves, but the incorporation was much less than from straight chain precursors. Not more than one-third of the (14)C incorporated into the surface lipids was found in the C(29) paraffin and derivatives, whereas more than two-thirds of the (14)C from straight chain precursors are usually found in these compounds. The small amount of (14)C incorporated into the paraffin fraction was found in the n-C(29) paraffin rather than branched paraffins showing that the (14)C in the paraffin must have come from degradation products. Radio gas-liquid chromatography of the saturated fatty acids showed that, in addition to the n-C(16) acid which was formed from both branched precursors, isoleucine-U-(14)C gave rise to branched C(15), C(17), and C(19) fatty acids, and isobutyrate-1-(14)C gave rise to branched C(16) and C(18) acids. Thus the reason for the failure of broccoli leaf to incorporate branched precursors into branched paraffins is not the unavailability of branched fatty acids, but the absolute specificity of the system that synthesizes paraffins, probably the elongation-decar-boxylation enzyme complex. Consistent with this view, no labeled branched fatty acids longer than C(19) could be found in the broccoli leaf. The branched fatty acids were also found in the surface lipids indicating that the epidermal layer of cells did have access to branched chains. Thus the paraffin synthesizing enzyme system is specific for straight chains in broccoli, but the fatty acid synthetase is not.     

Further evidence for an elongation-decarboxylation mechanism in the biosynthesis of paraffins in leaves

In isolated tobacco leaves l-valine-U-¹⁴C gave rise to labeled even-numbered isobranched fatty acids containing 16 to 26 carbon atoms and iso C₂₉, iso C₃₁, and iso C₃₃ paraffins. l-Isoleucine-U-¹⁴C on the other hand produced labeled odd-numbered anteiso C₁₇ to C₂₇ fatty acids and anteiso C₃₀ and C₃₂ paraffins. Trichloroacetic acid inhibited the incorporation of isobutyrate into C₂₀ and higher fatty acids and paraffins without affecting the synthesis of the C₁₆ and C₁₈ fatty acids. Thus the very long branched fatty acids are biosynthetically related to the paraffins. In Senecio odoris leaves acetate-1-¹⁴C was incorporated into the paraffins (mainly n-C₃₁) only in the epidermis although acetate was readily incorporated into fatty acids in the mesophyll tissue. Similarly only the epidermal tissue incorporated acetate into fatty acids longer than C₁₈ suggesting that the epidermis is the site of synthesis of both paraffins and the very long fatty acids. In broccoli leaves n-C₁₂ acid labeled with ¹⁴C in the carboxyl carbon and ³H in the methylene carbons was incorporated into C₂₉ paraffin without the loss of ¹⁴C relative to ³H. Since n-C₁₈ acid is known to be incorporated into the paraffin without loss of carboxyl carbon these results suggest that the condensation of C₁₂ acid with C₁₈ acid is not responsible for n-C₂₉ paraffin synthesis in this tissue. Thus all the experimental evidence thus far obtained strongly suggests that elongation of fatty acids followed by decarboxylation is the most likely pathway for paraffin biosynthesis in leaves.     

Branched Chain Amino Acid Metabolism in the Biosynthesis of Lycopersicon pennellii Glucose Esters

Lycopersicon pennellii Corr. (D'Arcy) an insect-resistant, wild tomato possesses high densities of glandular trichomes which exude a mixture of 2,3,4-tri-O-acylated glucose esters that function as a physical impediment and feeding deterrent to small arthropod pests. The acyl moieties are branched C₄ and C₅ acids, and branched and straight chain C₁₀, C₁₁, and C₁₂ acids. The structure of the branched acyl constituents suggests that the branched chain amino acid biosynthetic pathway participates in their biosynthesis. [¹⁴C]Valine and deuterated branched chain amino acids (and their oxo-acid derivatives) were incorporated into branched C₄ and C₅ acid groups of glucose esters by a process of transamination, oxidative decarboxylation and subsequent acylation. C₄ and C₅ branched acids were elongated by two carbon units to produce the branched C₁₀-C₁₂ groups. Norvaline, norleucine, allylglycine, and methionine also were processed into acyl moieties and secreted from the trichomes as glucose esters. Changes in the acyl composition of the glucose esters following sulfonylurea herbicide administration support the participation of acetohydroxyacid synthetase and the other enzymes of branched amino acid biosynthesis in the production of glucose esters.     

Signature Lipids and Stable Carbon Isotope Analyses of Octopus Spring Hyperthermophilic Communities Compared with Those of Aquificales Representatives

The molecular and isotopic compositions of lipid biomarkers of cultured Aquificales genera have been used to study the community and trophic structure of the hyperthermophilic pink streamers and vent biofilm from Octopus Spring. Thermocrinis ruber, Thermocrinis sp. strain HI 11/12, Hydrogenobacter thermophilus TK-6, Aquifex pyrophilus, and Aquifex aeolicus all contained glycerol-ether phospholipids as well as acyl glycerides. The n-C_20:1 and cy-C₂₁ fatty acids dominated all of the Aquificales, while the alkyl glycerol ethers were mainly C_18:0. These Aquificales biomarkers were major constituents of the lipid extracts of two Octopus Spring samples, a biofilm associated with the siliceous vent walls, and the well-known pink streamer community (PSC). Both the biofilm and the PSC contained mono- and dialkyl glycerol ethers in which C₁₈ and C₂₀ alkyl groups were prevalent. Phospholipid fatty acids included both the Aquificales n-C_20:1 and cy-C₂₁, plus a series of iso-branched fatty acids (i-C_15:0 to i-C_21:0), indicating an additional bacterial component. Biomass and lipids from the PSC were depleted in ¹³C relative to source water CO₂ by 10.9 and 17.2‰, respectively. The C_20–21 fatty acids of the PSC were less depleted than the iso-branched fatty acids, 18.4 and 22.6‰, respectively. The biomass of T. ruber grown on CO₂ was depleted in ¹³C by only 3.3‰ relative to C source. In contrast, biomass was depleted by 19.7‰ when formate was the C source. Independent of carbon source, T. ruber lipids were heavier than biomass (+1.3‰). The depletion in the C_20–21 fatty acids from the PSC indicates that Thermocrinis biomass must be similarly depleted and too light to be explained by growth on CO₂. Accordingly, Thermocrinis in the PSC is likely to have utilized formate, presumably generated in the spring source region.     

In vivo incorporation of acetate into the acyl moieties of polar lipids from etiolated leek seedlings

Seven-day-old leek seedlings actively synthesize lipids in vivo from [1-¹⁴C]acetate, both in the light and in the dark. In the dark, phospholipid synthesis is more effective than galactolipid synthesis. Whatever the time of acetate incorporation by the etiolated seedlings, very long chain fatty acids having from 20 to 26 carbon atoms are found in all the polar lipids, including the acyl-CoAs. All of the labelled very long chain fatty acids incorporated into the polar lipids are saturated. On the other hand, the labelled C₁₈-fatty acids are unsaturated in phospholipids and galactolipids and almost no label is found in the saturated or unsaturated C₁₈-fatty acids of the acyl-CoAs.     

In vitro synthesis of mycolic acids by the fluffy layer fraction of Bacterionema matruchotii

Biosynthetic activity for mycolic acid occurred in the fluffy layer fraction but not in the 5000g supernatant of Bacterionema matruchotii. With [1-¹⁴C]palmitic acid as precursor for the in vitro system, the predominant product was identified as C_32:0 mycolic acid by radio-gas-liquid chromatographie (radio-GLC) and gas chromatographic/mass spectroscopic analyses; if [1-¹⁴C]stearic acid was used, two major radioactive peaks appeared on GLC: one corresponding to the peak of (C_34:0 + C_34:1) mycolic acids and the other to (C_36:0 + C_36:1) mycolic acids. By pyrolysis/radio-GLC analysis, C_32:0 mycolic acid synthesized by [1-¹⁴C]palmitic acid was pyrolyzed at 300 °C to form palmitaldehyde (the mero moiety) and methyl palmitate (the branch moiety). The pH optimum for the incorporation of [1-¹⁴C]palmitate into bacterionema mycolic acids was 6.4 and the reaction required a divalent cation. The in vitro system utilized myristic, palmitic, stearic and oleic acids (probably via their activated forms) well as precursors, among which myristic and palmitic acids were more effective than the rest. Avidin showed no effect on the biosynthesis of mycolic acid from ¹⁴C-palmitate whereas cerulenin, a specific inhibitor of β-ketoacyl synthetase in de novo fatty acid synthesis, inhibited the reaction at a relatively higher concentration. Thin-layer chromatographic analysis of lipids extracted from the reacting mixture without alkaline hydrolysis showed that both exogenous [1-¹⁴] fatty acid and synthesized mycolic acids were bound to an unknown compound by an alkali-labile linkage and this association seemed to occur prior to the condensation of two molecules of fatty acid.     

Occurrence and Carbon Metabolism of Green Nonsulfur-like Bacteria in Californian and Nevada Hot Spring Microbial mats as Revealed by Wax Ester Lipid Analysis

Green nonsulfur-like bacteria (GNSLB) in Yellowstone hot spring microbial mats have been extensively studied and are thought to operate both as photoheterotrophs and photoautotrophs. Here we studied the occurrence and carbon metabolisms of GNSLB by analyzing the distribution and isotopic composition of their characteristic wax ester lipids in four Californian and Nevada hot spring microbial mats at a range of temperatures (37–96°C). The distribution of wax esters varied strongly with temperature. At temperatures between 50–60°C the wax ester composition in each of the four hot spring microbial mats was dominated by C₃₀ to C₃₆ wax esters, consisting of mixtures of C₁₅-C₁₈ n-alkyl and branched fatty acids and alcohols, typical for GNSLB. Stable carbon isotopic analysis showed that these wax esters were only depleted by 5 to 10‰ compared to dissolved inorganic carbon in the overlying water, suggesting that these GNSLB were mainly autotrophic. However, analysis of different depth layers of one microbial mat showed that these GNSLB wax esters were increasingly depleted in ¹³C with depth, suggesting that photoautotrophy mainly occurred in the top layer of the mat. ¹³C-depleted C₃₆-C₄₄ wax esters were found in one hot spring at high temperatures (77–96°C) and are likely derived from allochtonous plant waxes. At several lower temperature sites (35–40°C) the wax esters were predominantly composed of C₂₈, C₃₀ and C₃₂ wax esters consisting of mixtures of C₁₄-C₁₆ fatty acids and n-alkanols and were depleted in ¹³C by 15–20‰ relative to dissolved inorganic carbon, suggesting they may be derived from heterotrophic organisms. Our results indicate that autotrophic GNSLB occur widely in hot springs and that diverse groups of organisms contribute to the pool of wax ester lipids in hot spring environments.     

Glycerolipid synthesis in Chlorella kessleri 11h: I. Existence of a eukaryotic pathway

The fatty acid distributions at the sn-1 and sn-2 positions in major chloroplast lipids of Chlorella kessleri 11h, monogalactosyl diacylglycerol (MGDG) and digalactosyl diacylglycerol (DGDG), were determined to show the coexistence of both C₁₆ and C₁₈ acids at the sn-2 position, i.e. of prokaryotic and eukaryotic types in these galactolipids. For investigation of the biosynthetic pathway for glycerolipids in C. kessleri 11h, cells were fed with [¹⁴C]acetate for 30 min, and then the distribution of the radioactivity among glycerolipids and their constituent fatty acids during the subsequent chase period was determined. MGDG and DGDG were labeled predominantly as the sn-1-C₁₈-sn-2-C₁₆ (C₁₈/C₁₆) species as early as by the start of the chase, which suggested the synthesis of these lipids within chloroplasts via a prokaryotic pathway. On the other hand, the sn-1-C₁₈-sn-2-C₁₈ (C₁₈/C₁₈) species of these galactolipids gradually gained radioactivity at later times, concomitant with a decrease in the radioactivity of the C₁₈/C₁₈ species of phosphatidylcholine (PC). The change at later times can be explained by the conversion of the C₁₈/C₁₈ species of PC into galactolipids through a eukaryotic pathway. The results showed that C. kessleri 11h, distinct from most of other green algal species that were postulated mainly to use a prokaryotic pathway for the synthesis of chloroplast lipids, is similar to a group of higher plants designated as 16:3 plants in terms of the cooperation of prokaryotic and eukaryotic pathways to synthesize chloroplast lipids. We propose that the physiological function of the eukaryotic pathway in C. kessleri 11h is to supply chloroplast membranes with 18:3/18:3-MGDG for their functioning, and that the acquisition of a eukaryotic pathway by green algae was favorable for evolution into land plants.     

RADIOLABELING STUDIES OF LIPIDS AND FATTY ACIDS IN NANNOCHLOROPSIS (EUSTIGMATOPHYCEAE), AN OLEAGINOUS MARINE ALGA1

The synthesis of fatty acids and lipids in Nannochloropsis sp. was investigated by labeling cells in vivo with [¹⁴C]-bicarbonate or [¹⁴C]-acetate. [¹⁴C]-bicarbonate was incorporated to the greatest extent into 16:0, 16:1, and 14:0 fatty acids, which are the predominant fatty acids of triacylglycerols. However, more than half of the [¹⁴C]-acetate was incorporated into longer and more desaturated fatty acids, which are constituents of membrane lipids. [¹⁴C]-acetate was incorporated most strongly into phosphatidylcholine, which rapidly lost label during a 5-h chase period. The label associated with phosphatidylethanolamine also decreased during the chase period, whereas label in other membrane lipids and triacylglycerol increased. The dynamics of labeling, along with information regarding the acyl compositions of various lipids, suggests that 1) the primary products of chloroplast fatty acid synthesis are 14:0, 16:0, and 16:1; 2) C₂₀ fatty acids are formed by an elongation reaction that can utilize externally supplied acetate; 3) phosphatidylcholine is a site for desaturation of C₁₈ fatty acids; and 4) phosphatidylethanolamine may be a site for desaturation of C₂₀ fatty acids.     

Fat Metabolism in Higher Plants: XL. Synthesis of Fatty Acids in the Initial Stage of Seed Germination

To understand more fully organelle membrane assemblage, the synthesis of the first fatty acids by the germinating pea, Pisum sativum, was studied by the incorporation of either tritiated water or acetate-1-¹⁴C into lipids by the intact, initially dry seed. After a lag phase, labeling proceeded linearly. This lag phase ended when uptake of water had increased the seed weight to 185% of its original weight. The first fatty acids synthesized were palmitic and stearic followed shortly after by long chain saturated fatty acids (C₂₀-C₂₆). The synthesis of very long chain acids was consistently characteristic of several other seeds in early stages of germination. The majority of the radioactive acids were present in phospholipids and were localized in particulate fractions. The acyl components of phosphatidyl glycerol were highly labeled. The very long chain acids were found predominantly in the waxes. Pulse labeling indicated little turnover of the labeled fatty acids. Evidence is presented indicating that the enzymes for fatty acid synthesis are already present in the dry seed and participate in the synthesis of fatty acids once a critical water content of the seed is achieved.     

Influence of dietary sources of fat on lipid synthesis in mink (Mustela vison) mammary tissue

• 1.1. The fatty acid composition of the triglyceride fraction of mink milk sampled during mid-lactation (day 28 post partum) from two nursing mink was compared to that of plasma samples and to the fatty acid composition of the feed rations used.
• 2.2. Chemical analysis of the triglyceride composition of mink milk demonstrated only minute concentrations of fatty acids with a chain length below C₁₄.
• 3.3. The saturated C_16:0- and C_18:0-unit fatty acids in mink milk made up for 24–40% of the total amount of fatty acids extracted, the remainder being represented by mono and polyunsaturated long-chain (C₁₆-C₂₄) fatty acids.
• 4.4. Preliminary in vitro experiments proved the incorporation of¹⁴C-labelled glucose, acetate or palmitate into triacylglycerols in cultures of mink mammary tissue to be linear for at least 2 hr.
• 5.5. The in vitro capacity for de novo fatty acid synthesis in mink mammary tissue using 14C-labelled glucose or acetate was low, i.e. ranging from 0.096–0.109 nmol/g (fresh tissue)/min, and amounted to only about 5% of that obtained in the case of [¹⁴C]palmitic acid incubation.
• 6.6. Following ¹⁴C-labeIled acetic or palmitic acid incubation of mink mammary tissue neither desaturation nor chain elongation was observed.
• 7.7. In response to long-term feeding on rations with two different sources of animal fat (F = fish oil or L = lard) the influence of compositional changes in dietary neutral lipids on the fatty acid composition of the lipids of mink milk is discussed.

     

Phospholipid and acid composition of Nocardia and nocardoid bacteria as criteria of classification

Phospholipid and acid composition of 5 strains of ‘true’ Nocardia and 4 strains of nocardoid bacteria have been studied. A great homogeneity was found in all the Nocardia species: phospholipids consist of cardiolipin, phosphatidyl ethanolamine, phosphatidylinositol and phosphatidylinositol mannoside. Streptomyces (Nocardia) mediterranei did not contain phosphatidylinositol and Oerskovia (Nocardia) turbata had no phosphatidyl ethanolamine. The fatty acid composition of these phospholipids was determined and was found different in Nocardia and nocardoid species. Nocardia were rich in straight chain fatty acids and tuberculostearic acid while the phospholipids of nocardoid bacteria contained greater amounts of branched fatty acids. The fatty acids from acetone soluble lipids consisted of hydroxy and non-hydroxy compounds. Hydroxy acids were found in Nocardia which contained nocardic acids: high MW β-hydroxy α-branched acids and in S. mediterranei which contained β-hydroxy acids with 15–17 carbon atoms. Non-hydroxy acids were essentially palmitic and tuberculostearic acids in Nocardia species while S. mediterranei and O. turbata contained great amounts of iso acids from C₁₄ to C₁₇. Phospholipid and acid composition are discussed as criteria of taxonomic classification of Nocardia and related Actinomycetes.     

Spinach Leaves Desaturate Exogenous [C]Palmitate to Hexadecatrienoate : Evidence that de Novo Glycerolipid Synthesis in Chloroplasts Can Utilize Free Fatty Acids Imported from Other Cellular Compartments

Long-chain ¹⁴C-fatty acids applied to the surface of expanding spinach leaves were incorporated into all major lipid classes. When applied in diethyleneglycol monomethyl ether solution, as done by previous workers, [¹⁴C]palmitic acid uptake was much lower than that of [¹⁴C] oleic acid. However, when applied in a thin film of liquid paraffin the rate of [¹⁴C] palmitic acid metabolism was rapid and virtually complete. Considerable radioactivity from [¹⁴C]palmitate incorporated into lipids following either application method gradually appeared in polyunsaturated C₁₆ fatty acids esterified to those molecular species of galactolipids previously thought to be made using only fatty acids synthesized and retained within the chloroplast. Evidence for the incorporation of radioactivity from exogenous [¹⁴C]oleate into those same molecular species of galactolipids was less compelling. The unexpected availability of fatty acids bound to extrachloroplastidal lipids for incorporation into galactolipids characteristically assembled entirely within the chloroplast emphasizes the need to reassess interrelations between the “prokaryotic” and “eukaryotic” pathways of galactolipid formation.     

Photosynthesis,lipids and proteins in the cyanobacteriumSynechocystis PCC 6803 as affected by temperature

The cyanobacteriumSynechocystis PCC 6803 was grown photoautotrophically in an inorganic medium at constant growth temperatures of 20, 38 (control) or 43°C for 9 h. The up and down-shift of cultivation temperature decreased the growth as measured by culture absorbance and chlorophylla content. However, high temperature slightly increased the oxygen evolution while temperature lower than control inhibited oxygen evolution during the whole incubation period. The protein synthesis studied by¹⁴C-labeled protein declined under low temperature by about 50%. The fatty acid pattern is characterized as lacking in C₂₀/C₂₂ acids but containing large amounts of C₁₆ and C₁₈ polyunsaturated fatty acids, 16:2 and 18:3 in particular. The lower temperature increased the percentage of monounsaturated fatty acids while higher temperature increased the saturated fatty acid content in total lipids and lipid classes studied.     

Cellular fatty acid composition ofCorallococcus coralloides

The fatty acid composition of five strains ofCorallococcus coralloides and three reference species ofMyxococcus were determined by gas-liquid chromatography. Methyl esters of fatty acid containing from 12 to 22 carbon atoms were identified. The major fatty acids present were C₁₅ and C₁₇ saturated branched chain, and both C₁₆ saturated and unsaturated straight chain acids. The C₁₇ saturated branched and straight chain acids, which were in valuable concentration in species ofMyxococcus, were not, however, detected in all strains ofC. coralloides. The application of these results in the distinction ofC. coralloides from the genusMyxococcus is discussed.     

The Metabolism of the Germinating Oil Palm (Elaeis guineensis) Seedling : In Vivo Studies

The metabolism of ¹⁴C-labeled fatty acids and triacylglycerols was followed in intact germinating oil palm seedlings as well as in tissue slices. In the germinating seedling, the shoot contained a normal pattern of membrane fatty acids (mainly C₁₆, C_18:1, C_18:2) but the kernel contained about 68% C₁₂ and C₁₄ fatty acids. Haustorium fatty acids were intermediate between the two. [¹⁴C]Acetate was actively metabolized by shoot and haustorium slices but not so actively by the kernel. Approximately 9% to 17% was converted to water-soluble substances, 4% to 6% to CO₂, and 0.5% to 5.9% to lipids. The fatty acids synthesized in the shoot and haustorium were mainly C₁₆, C₁₈, and C_18:1 fatty acids but in the kernel about 18% to 32% of the ¹⁴C-fatty acids were C₁₂ fatty acids.

[¹⁴C]Lauric acid was absorbed and metabolized by haustorium slices and by the haustorium in intact seedlings; it was partly esterified to triacylglycerols and also converted to water-soluble substances and insoluble tissue material. In contrast, tri-[¹⁴C]laurin was absorbed but not metabolized. The haustorium also absorbed other fatty acids but the longer chain (C₁₆ and C₁₈) fatty acids were not esterified or metabolized further. Preincubation of the haustorium with plant hormones or in the presence of kernel tissue did not alter its inactivity towards tri-[¹⁴C]laurin.

When tri-[¹⁴C]laurin or [¹⁴C]lauric acid were injected into the seed or the shoot, there was no movement or radioactivity to other parts of the seedling. When injected into the shoot, but not into the seed, tri-[¹⁴C] laurin was hydrolyzed and partly metabolized to water-soluble substances.

     

Biosynthesis of methyl branched hydrocarbons of the German cockroach Blattella germanica (L.) (orthoptera,blattellidae)

The precursors and directionality of synthesis of the methyl branched cuticular hydrocarbons and the female contact sex pheromone, 3,11-dimethyl-2-nonacosanone, of the German cockroach, Blattella germanica, were investigated by radiotracer and carbon-13 NMR techniques. The amino acids [G-³H]valine, [4,5-³H]isoleucine and [3,4-¹⁴C₂]methionine labeled the hydrocarbon fraction in a manner indicating that the carbon skeletons of all three amino acids serve as the methyl branch group donor. The incorporation of [1,4-¹⁴C₂]- and [2,3-¹⁴C₂]succinates into the hydrocarbon and acylglycerol/polar lipid fractions indicated that succinate also served as a precursor to methylmalonyl-CoA. Carbon-13 NMR analyses showed that [1-¹³C]propionate labeled the carbon adjacent to the tertiary carbon, and, for the 3,x-dimethylalkanes, that carbon-4 and not carbon-2 was enriched. [1-¹³C]Acetate labeled carbon-2 of these hydrocarbons. This indicates that the methyl branching groups of the 3,x-dimethylalkanes were inserted early in the chain elongation process. [3,4,5-¹³C₃]Valine labeled the methyl, tertiary and carbon adjacent to the tertiary carbon of the methyl branched alkanes. Thus, the methyl branched hydrocarbon was formed by the insertion of methylmalonyl units derived from propionate, isoleucine, valine, methionine and succinate early in chain elongation.     

An approach to improve methanogenesis through the use of mixed cultures isolated from biogas digester

Biogas production has been shown to be inhibited by branched chain fatty acids (isobutyric, isovaleric) produced in the digester by cellulolytic organisms. Performance of these mixed cellulolytic cultures isolated at 25°C (C₂₅) and at 35° (C₃₅) in a batch digester using cattle manure confirmed that C₃₅, which forms mainly straight chain fatty acids from cellulose was more suitable for use as an inoculum than C₂₅ which formed predominantly branched chain fatty acids. Reconstitution of cellulolytic culture C₃₅ and mixed methanogens M₃₅ almost doubled both the amount and rate of methane production. Cellulolytic culture was useful in pretreatment of water hyacinth prior to its use as a substrate for methane generation A method for preservation and transportation of mixed cellulolytic culture for use as an inoculum in the digester is described.     

Labelling of lipids by D-[1-14C]glucose, D-[6-14C]glucose and D-[3-3H]glucose in pancreatic islets from normal and GK rats

In pancreatic islets prepared from either normal or GK rats and incubated at either low (2.8 mM) or high (16.7 mM) D-glucose concentration, the labelling of both lipids and their glycerol moiety is higher in the presence of D-[1-¹⁴C]glucose than D-[6-¹⁴C]glucose. The rise in D-glucose concentration augments the labelling of lipids, the paired ¹⁴C/³H ratio found in islets exposed to both D-[1-¹⁴C]glucose or D-[6-¹⁴C]glucose and D-[3-³H]glucose being even slightly higher at 16.7 mM D-glucose than that found, under otherwise identical conditions, at 2.8 mM D-glucose. Such a paired ratio exceeds unity in islets exposed to D-[1-¹⁴C]glucose. The labelling of islet lipids by D-[6-¹⁴C]glucose is about 30 times lower than the generation of acidic metabolites from the same tracer. These findings indicate (i) that the labelling of islet lipids accounts for only a minor fraction of D-glucose catabolism in pancreatic islets, (ii) a greater escape to L-glycerol-3-phosphate of glycerone-3-phosphate generated from the C₁-C₂-C₃ moiety of D-glucose than D-glyceraldehyde-3-phosphate produced from the C₄-C₅-C₆ moiety of the hexose, (iii) that only a limited amount of [3-³H]glycerone 3-phosphate generated from D-[3-³H]glucose is detritiated at the triose phosphate isomerase level before being converted to L-glycerol-3-phosphate, and (iv) that a rise in D-glucose concentration results in an increased labelling of islet lipids, this phenomenon being somewhat more pronounced in the case of D-[1-¹⁴C]glucose or D-[6-¹⁴C]glucose rather than D-[3-³H]glucose.     

Salinicoccus halitifaciens sp. nov., a novel bacterium participating in halite formation

Strain JC90^T was isolated from a soda lake in Lonar, India. Strain JC90^T maintains its external pH to 8.5 and participates in halite formation. Based on 16S rRNA gene sequence similarity studies, strain JC90^T was found to belong to the genus Salinicoccus and is most closely related to “Salinicoccus kekensis” K164^T (99.3 %), Salinicoccus alkaliphilus T8^T (98.4 %) and other members of the genus Salinicoccus (<96.5 %). However Strain JC90^T is <36 % related (based on DNA–DNA hybridization) with the type strains of “S. kekensis” K164^T and S. alkaliphilus T8^T. The DNA G+C content of strain JC90^T was determined to be 46 mol %. The cell-wall amino acids were identified as lysine and glycine. Polar lipids were found to include diphosphatidylglycerol, phosphatidylglycerol, phosphatidyl ethanolamine, an unidentified glycolipid and unidentified lipids (L1,2). Major hopanoids of strain JC90^T were determined to be bacterial hopane derivatives (BHD1,2), diplopterol, diploptene and two unidentified hopanoids (UH1,2). The predominant isoprenoid quinone was identified as menaquinone (MK-6). Anteiso-C_15:0 was determined to be the predominant fatty acid and significant proportions of iso-C_14:0, C_14:0, iso-C_15:0, C_16:0, iso-C_16:0, iso-C_17:0, anteiso-C_17:0 and C_18:02OH were also detected. The results of physiological and biochemical tests support the molecular evidence and allowed a clear phenotypic differentiation of strain JC90^T from all other members of the genus Salinicoccus. Strain JC90^T is therefore considered to represent a novel species, for which the name Salinicoccus halitifaciens sp. nov. is proposed. The type strain is JC90^T (=KCTC 13894^T =DSM 25286^T).