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PubMedDate not supplied

Chemical analysis of processing of spiralin, the major lipoprotein of Spiroplasma melliferum.

The plasma membrane of Spiroplasma melliferum contains a major membrane-associated lipoprotein called spiralin. In this study, the processing pathway of spiralin was investigated by chemical analysis of the purified protein and by using [35S]cysteine, [35S]methionine, [14C]myristic acid (14C-14:0), [14C]palmitic acid (14C-16:0), and globomycin. SDS-PAGE analysis of membrane proteins showed the leader peptide cleavage of prospiralin and provided evidence for an apparent selectivity in the acylation: the unprocessed protein was labelled with 14C-16:0 only (O-ester-linked acyl chains), and the mature form with both 14C-labelled fatty acids (O-ester-linked + amide-linked chains). Chemical analysis of the purified protein revealed that spiralin contains S-glycerylcysteine and is covalently modified with two O-ester-linked acyl chains and one amide-linked fatty acid chain. However, a specific selectivity in the O- and the N-acylations was not confirmed; palmitate and stearate were the major components. The amounts of O-ester- and amide-linked acyl chains, the resistance to Edman degradation and the presence of S-glycerylcysteine together indicate that spiralin is a "classical" lipoprotein (i.e. is triacylated) and is probably processed by a mechanism similar to that described for gram-negative eubacteria. On the basis of these findings, a biogenesis pathway for spiralin is proposed.

Amino Acids↗
PubMed2005

The fatty acid profile of vegetative Azotobacter vinelandii ATCC 12837: growth phase-dependence.

Fatty acids of Azotobacter vinelandii ATCC 12837 were determined at various times during aerobic vegetative growth at 30 degrees C to provide baseline data for studying the effects of chemical agents on the organism's survival and fatty acid biosynthesis. Palmitate (16:0) was the highest at 36.7+/-4.3 mol% (mean+/-SD) after the first 5 h in fresh culture, decreasing slightly to 33.4+/-2.6 mol% at 49 h. The other fatty acids were therefore each normalized as a ratio of 16:0. At 5 h, as a ratio of 16:0, myristate (14:0) was 0.14+/-0.06, palmitoleate (16:1cDelta9-10) 0.13+/-0.06, oleate (18:1cDelta9-10) 0.21+/-0.12, cis-vaccenate (18:1cDelta11-12) 0.30+/-0.17 and stearate (18:0) 0.68+/-0.02. As the growth phase advanced to 49 h, 14:0 and 16:1cDelta9-10 increased, 18:1cDelta9-10 decreased and cis-vaccenate reciprocally increased, whereas 18:0 decreased. These suggest that the saturated fatty acid biosynthesis pathway yielded 16:0 and 18:0 in the 5-h lag period. By desaturation, 18:0 formed the unsaturated fatty acid (UFA) 18:1cDelta9-10. As the culture aged, the anaerobic UFA biosynthesis pathway formed 16:1cDelta9-10, which was elongated to 18:1cDelta11-12. These fatty acid alterations represent a homeoviscous adaptation, modulating the microbe's membrane lipid viscosity for optimal cellular function.

Azotobacter vinelandii↗
PubMed1999

Independent production of two molecular forms of a recombinant Rhizopus oryzae lipase by KEX2-engineered strains of Saccharomyces cerevisiae.

A mixture of rProROL having the full-length prosequence (97 amino acids) for a recombinant lipase of Rhizopus oryzae (rROL) and r28ROL having 28 amino acids of the same prosequence has been produced as active forms by Saccharomyces cerevisiae [Takahashi et al. (1998) J Ferment Bioeng 86: 164-168]. However, the separation of rProROL and r28ROL has not been successful due to their identical behavior on column chromatographs, presumably because of the similarity of their surface properties. The independent production of two different molecular forms of rROL was carried out using KEX2-engineered strains of S. cerevisiae, since r28ROL was predicted to be a product from rProROL by a Kex2-like protease. rProROL was successfully obtained by expression of the ROL gene in the S. cerevisiae kex2 strain in which the KEX2 gene encoding Kex2p was disrupted, while r28ROL was obtained by co-expression of the gene (KEX2 delta 613) encoding the soluble form of the C-terminal truncated Kex2 protease (sKex2p). The specific lipase activities of rProROL and r28ROL were 92.9 U/mg and 140 U/mg, respectively. rProROL was stable at pH 2.2-8.0, and showed the optimal reaction temperature to be 30-35 degrees C with a T50 of 55 degrees C (T50 is the temperature resulting in 50% loss of activity). The values for r28ROL were pH 3.0-10.0, 25-30 degrees C, and 40 degrees C, respectively. rProROL was an N-linked glycosylated form, but r28ROL was not. The enhanced thermostability of rProROL did not seem to be due to the N-linked glycosylation, as judged by the results of the Endo H treatment. rProROL had the highest esterase activity toward p-nitrophenyl laurate (C12), whereas r28ROL had the highest esterase activity toward p-nitrophenyl caprylate (C8) and stearate (C18). These results suggest that the distinct properties of these two forms of lipase are caused by the different length of the ROL prosequence.

Bacterial Proteins↗
PubMed2005

Antimetastatic effect of a lipophilic ascorbic acid derivative with antioxidation through inhibition of tumor invasion.

PURPOSE: Ascorbic acid (AA), the natural antioxidant, has been demonstrated to exert an antimetastatic action; however, AA is quite unstable in physiological condition. The aim of the present study is to investigate the stability, the antioxidation and the antimetastatic effects of three lipophilic AA derivatives in vitro as well as in vivo. METHODS: The 95D cells were treated with ascorbic acid-2-O-phosphate-6-O-laureate (AA2P6L), ascorbic acid-2-O-phosphate-6-O-myristate (AA2P6M) and ascorbic acid-2-O-phosphate-6-O-stearate (AA2P6S). AA derivatives' stability in medium under cell culture condition, in the presence and in the absence of 95D cells, was assessed by high-performance liquid chromatography assay. Cell viability and intracellular oxidative stress were measured by MTT assay and CDCFH assay, respectively. Wound healing assay and cell adhesion assay were used to investigate the antimetastatic activities against 95D cells in vitro, and the C57BL/6 mice model was used to evaluate the antimetastatic action in vivo. RESULTS: All the three AA derivatives exhibited excellent stability, significantly different from AA. Results of MTT assay showed that IC(50) values of the cytotoxicity of those AA derivatives, namely AA2P6L, AA2P6M and AA2P6S, were 38.46, 28 and 22.97 microg/ml, while the CDCFH assay indicated that EC(50) values of antioxidant effects on 95D cells were 31.12, 33.51 and 38.31 microg/ml, respectively. Through the ratio of IC(50) vs EC(50) for AA derivatives, AA2P6L was demonstrated to be the most effective AA derivative, which retained the antioxidant ability as well as low cytotoxicity. AA2P6L dose-dependently inhibited 95D cells' migration and adhesion, by 50% at the concentration of 20 and 57 microg/ml, respectively. In the animal experiment, intraperitoneal administration of 75 mg/kg AA2P6L decreased the number of metastatic nodules by 62% and elevated the survival rate of C57BL/6 mice about onefold compared to the control group. CONCLUSION: AA2P6L, a lipophilic AA derivative with antioxidation, is shown to be a potent antimetastatic agent through the inhibition of tumor invasion. These results support future investigations on the feasibility of cancer chemotherapy with AA2P6L.

Animals↗
PubMed1997

Changes in natural killer cell subpopulations in lead workers.

To investigate the effects of lead on human immune system, we analyzed T cell subpopulations (CD4+, CD8+ and CD3+ cells), natural killer (NK) cell subpopulations (CD16+ and CD57+ cells) and B (CD19+) cells in peripheral blood in 29 male lead workers. All were engaged in manufacturing lead stearate in a chemical factory. They were aged 23-74 (mean 49) years. Their blood lead concentrations (PbB) were between 7 and 35 (mean 18) micrograms/dl. They were divided into two groups according to their PbB: a high-PbB group (> or = 20 micrograms/dl), and a low-PbB group (< 20 micrograms/dl). The control group consisted of 19 "healthy" male workers without a history of occupational exposure to lead or to other hazardous substances, aged 48-67 (mean 58) years. The number and percentage of CD16+ cells in the high-PbB group were significantly lower than those in the controls and in the low-PbB group. There was significant negative correlation between the number of CD16+ cells and PbB in the lead workers. The percentage of CD8+ cells in the high-PbB group was larger than that in the controls and in the low-PbB group. It is suggested that the CD16+ NK cell should be a major site of the effects of lead on lymphocyte subpopulations.

Adult↗
PubMed2006

The extracellular pollen coat in members of the Brassicaceae: composition, biosynthesis, and functions in pollination.

I have used cellular and molecular genetic and bioinformatic approaches to characterise the components of the pollen coat in plants of the family Brassicaceae, including Arabidopsis thaliana and several brassicas including Brassica napus, B. oleracea, and B. rapa. The pollen coat in these species is mostly made up of a unique mixture of lipids that is highly enriched in acylated compounds, such as sterol esters and phospholipids. These acyl lipids are characterised by an unusually high degree of saturation. The fatty acids typically contain 70-90% saturated acyl residues such as myristate, palmitate, and stearate. The major sterol components of the pollen coat are saturated fatty acyl esters of stigmasterol, campesterol, and campestdienol. In addition to lipids, the second major component of the pollen coat is a specific group of proteins that is dominated by a family of proteins that we term pollenins. Although pollenins are by far the major protein components of the pollen coat of members of the Brassicaceae, proteomic analysis reveals that there are several additional protein components, including lipases, protein kinases, a pectin esterase, and a caleosin. The biosynthesis of these lipids and proteins and their significance for overall pollen function are reviewed and discussed.

Brassicaceae↗
PubMed2006

Metabolism of polyunsaturated fatty acids and their toxicity to the microflora of the rumen.

Ruminal microorganisms hydrogenate polyunsaturated fatty acids (PUFA) present in forages and thereby restrict the availability of health-promoting PUFA in meat and milk. The aim of this study was to investigate PUFA metabolism and the influence of PUFA on members of the ruminal microflora. Eleven of 26 predominant species of ruminal bacteria metabolised linoleic acid (LA; cis-9,cis-12-18:2) substantially. The most common product was vaccenic acid (trans-11-18:1), produced by species related to Butyrivibrio fibrisolvens. alpha-Linolenic acid (LNA; cis-9,cis-12,cis-15-18:3) was metabolised mostly by the same species. The fish oil fatty acids, eicosapentaenoic acid (EPA; 20:5(n - 3)) and docosahexaenoic acid (DHA; 22:6(n - 3)) were not metabolised. Cellulolytic bacteria did not grow in the presence of any PUFA at 50 microg ml(-1), nor did some butyrate-producing bacteria, including the stearate producer Clostridium proteoclasticum, Butyrivibrio hungatei and Eubacterium ruminantium. Toxicity to growth was ranked EPA > DHA > LNA > LA. Cell integrity, as measured using propidium iodide, was damaged by LA in all 26 bacteria, but to different extents. Correlations between its effects on growth and apparent effects on cell integrity in different bacteria were low. Combined effects of LA and sodium lactate in E. ruminantium and C. proteoclasticum indicated that LA toxicity is linked to metabolism in butyrate-producing bacteria. PUFA also inhibited the growth of the cellulolytic ruminal fungi, with Neocallimastix frontalis producing small amounts of cis-9,trans-11-18:2 (CLA) from LA. Thus, while dietary PUFA might be useful in suppressing the numbers of biohydrogenating ruminal bacteria, particularly C. proteoclasticum, care should be taken to avoid unwanted effects in suppressing cellulolysis.

Animals↗
PubMed2006

Regulation of osteoblast gene expression and phenotype by polylactide-fatty acid surfaces.

Cell function is influenced by surface structure and molecules. Molecules that enhance cellular differentiation can be applied to tissue scaffold surfaces to stimulate endogenous tissue regeneration. The application of this approach to bone implants yields surfaces coated with factors (proteins, peptides, etc...) that promote the differentiation of osteoblasts, the cells that make bone. Increased bone formation leads to increased healing and union of the implant with endogenous bone. To obtain better control over surface coating we developed PLLA copolymers with allyl (PLLA-co-DAG) and 3-hydroxypropyl (PLLA-co-HP) side chains to which we can attach functional groups. Given the potential of fatty acids being able to incorporate into lipid bilayers and/or influence gene expression, we grafted different fatty acid side chains to PLLA-co-HP by esterifying the corresponding fatty acids with the PLLA-co-HP 3-hydroxypropyl side chains. The effects of the polymer modifications on osteoblasts were then evaluated. While cellular morphology differed between surface coatings, they did not reflect changes in cellular phenotype. Changes in gene expression were most evident with arachidonate and 3-hydroxypropyl side-chains which exhibited osteoblast differentiating capabilities. Linoleate, myristate, oleate, and stearate ester side-chains did not have a significant influence on osteoblast phenotype. Growth characteristics of osteoblasts did not differ between the fatty acid copolymer films, although cells grown on PLLA-co-HP exhibited a trend toward increased growth. Taken together our findings demonstrate that surface fatty acid composition can impact osteoblast phenotype.

Animals↗
PubMed2002

Dietary cholesterol modulates delta6 and delta9 desaturase mRNAs and enzymatic activity in rats fed a low-eFA diet.

The effects of a 1% addition of cholesterol to a diet low in EFA on FA desaturases were examined. The administration of cholesterol markedly increased the esterified cholesterol content in microsomes and total liver lipids from the first day, whereas the proportion of free cholesterol remained unaltered throughout the treatment. An excellent homeostasis in the free cholesterol content was apparently evoked by the acyl-CoA cholesterol acyltransferase. The cholesterol esters were mainly oleate, palmitate, and stearate, and the addition of cholesterol increased the relative proportions of cholesterol palmitoleate and oleate. The addition of cholesterol to a low-EFA diet induced, as in animals fed a high-EFA diet, a marked increase in liver stearoyl-CoA desaturase-1 mRNA and enzyme activity. This increased activity apparently evoked a similar enhancement of palmitoleic and oleic acids in total and microsomal liver lipids. The cholesterol-rich diet depressed the liver A6 and delta5 desaturase activity. However, the abundance of delta6 desaturase mRNA was not modified throughout the treatment. This indicates that the depressive effect is evoked at a step beyond that controlled by the mRNA level. The depression of both enzymatic activities was consistent with the decrease in the percentages of arachidonic acid and DHA in total and microsomal liver lipids. Taken together, these results indicate that through its modulating effect on the desaturases, dietary cholesterol may lead an animal or human fed low-EFA diet to a true deficiency by the decreased synthesis of the highly polyunsaturated acids derived from linoleic and alpha-linolenic acids.

Animals↗
PubMed2002

Pancreatic beta-cell alpha2A adrenoceptor and phospholipid changes in hyperlipidemic rats.

We previously showed that a 48-h intravenous lipid infusion in rats induces pancreatic beta-cell hypersensitivity to catecholamines. Our aim was to study the lipid-related changes that may account for such hypersensitivity in pancreatic islets. We show here that a 48-h increase in plasma FFA alters the binding characteristics of beta-cell alpha2 adrenoceptors in rats. Lipid infusion decreases pancreatic norepinephrine (NE) turnover rate by 28%, reflecting a reduction of pancreatic NE stores. Following lipid infusion, the density of alpha2 adrenoceptor binding sites is significantly lower and receptor affinity higher, both in islet homogenates (by three- and fivefold, respectively) and isolated whole beta-cells (by two- and sixfold, respectively). These changes correlate with the elevated insulin response to glucose found in lipid-infused rats. We also found a modification of islet phospholipid content, particularly in phosphoethanolamine species containing infused FA such as palmitate, oleate, stearate, and linoleate. This may account for the modifications in receptor affinity. These results suggest that hyperlipidemia-associated pathologies such as diabetes and obesity not only may result from alterations of metabolic pathways but also may be a consequence of early modifications in nervous firing rates and signal transduction pathways.

Animals↗
PubMed2003

Kinetic analysis of the selectivity of acylcarnitine synthesis in rat mitochondria.

Mitochondrial acylcarnitine synthesis is an obligatory step in the transport of cytosolic long-chain FA into the mitochondria. It is an important control point in the partitioning of cytosolic fatty acids to synthetic pathways or to mitochondrial beta-oxidation. Mitochondrial carnitine palmitoyltransferase I (CPT I; EC 2.3.1.21) is the enzyme that catalyzes the transformation of long-chain fatty acylCoA esters to acylcarnitine. Additionally, the isoform of acylCoA synthetase (EC 6.2.1.3) found in mitochondria, which is in close proximity to CPT I on the outer membrane, may act in concert with CPT I to form acylcarnitines from cytosolic nonesterified FA (NEFA). The mitochondrial acylcarnitine synthesis pathway is exposed to multiple fatty acid substrates present simultaneously in the cell milieu, with each fatty acid present at varying pool sizes. The selectivity of this pathway for any particular fatty acid substrate under conditions of multisubstrate availability has not yet been tested experimentally. Our objective was to develop mathematical equations that make use of kinetic constants derived from single-substrate experiments to predict the selectivity of the acylcarnitine synthesis pathway under conditions in which two or more substrates are present simultaneously. In addition, the derived equations must be verifiable by experiment. Our approach was to begin with a Michaelis-Menten model that describes the initial rates of an enzyme system acting on multiple and mutually competitive substrates. From this, we derived equations expressing ratios of reaction rates and fractional turnover rates for pairs of substrates. The derived equations do not require assumptions concerning the degree of enzyme saturation. Using rat mitochondrial preparations and the NEFA substrate pairs, linolenic-oleic acids and palmitic-linoleic acids, we showed that the shape of the experimentally derived data on acylcarnitine synthesis fits the predictions of the derived model equations. We further validated the derived equations by showing that their predictions calculated from previously published kinetic constants were consistent with data from actual experiments. Thus, we are able to conclude that with respect to acylcarnitine synthesis, the fractional turnover rate of the linolenic acid pool would always be 2.9-fold faster than that of the oleate pool regardless of the pool size of either fatty acid. Similarly, the fractional turnover rate of the palmitate pool would always be 1.8-fold faster than that of the linoleate pool regardless of pool size. We extended our kinetic model to more than two mutually competitive substrates. Using previously published rate constants for eight physiologically relevant fatty acids, the derived model predicts that regardless of pool size of any of the fatty acids, the linolenate pool, whether as NEFA or as a CoA ester, would always have the highest fractional turnover rate with respect to acylcarnitine synthesis. Conversely, the stearate pool whether as NEFA or as CoA ester will have the lowest fractional turnover rate relative to all the other fatty acids.

Animals↗
PubMed2005

Influence of stearic acid on postprandial lipemia and hemostatic function.

It has been suggested that fats rich in stearic acid may result in exaggerated postprandial lipemia and have adverse effects on hemostatic function. The effects of test meals containing different saturated and monounsaturated FA were compared in healthy subjects in a series of studies to investigate this hypothesis. Stearic acid, when present as cocoa butter, resulted in similar postprandial lipemia and factor VII activation compared with a meal containing high-oleic sunflower oil. Stearic acid when presented as shea butter or as randomized stearate-rich TAG resulted in decreased postprandial lipemia and decreased postprandial activation of factor VII. Stearic acid-rich test meals did not result in impaired fibrinolytic activity compared with either a low-fat meal or a meal high in oleate. The difference in responses between the different stearic acid-rich fats appears to be due to varying solid fat contents of the fats at 37 degrees C.

Adult↗
PubMed1997

Lipid composition of cultured endothelial cells in relation to their growth.

Human endothelial cells in culture were examined in different growth conditions. The human endothelial cell line, EA.hy 926 cell line, was used and cells were studied either in exponential growth phase, at confluence, or growth-arrested by serum deprivation. Phospholipids were separated and analyzed by high-performance thin-layer chromatography, and their fatty acids were quantified by gas-liquid chromatography. No significant differences in the phospholipid distributions were found between exponentially growing and confluent endothelial cells in which phosphatidylcholine (PC) represented the major phospholipid. In comparison, serum-deprived cells exhibited higher proportions of sphingomyelin and lower content of PC. We also found that among the total lipids, cholesterol level for dividing endothelial cells was lower than for cells growth-arrested either by serum deprivation or by contact inhibition at confluence. The global fatty acid distribution was not affected by the growth conditions. Thus, oleate (18:1 n-9 and 18:1 n-7), palmitate (C16:0), and stearate (C18:0) were the main components of endothelial cell membranes. However, the fatty acid distributions obtained from each phospholipid species differed with the growth status. Altogether, the data indicated that subtle modulations of endothelial cell metabolism appear upon cell growth. The resulting membrane-dependent cellular functions such as cholesterol transport and receptor activities can be expected to be relevant for lipid trafficking within the vessel wall in vitro and in vivo.

Cell Division↗
PubMed1999

Dietary supplementation with arachidonic and docosahexaenoic acids has no effect on pulmonary surfactant in artificially reared infant rats.

Despite the potential use of long chain polyunsaturated fatty acid (LCPUFA) supplementation to promote growth and neural development of the infant, little is known about potential harmful effects of the supplementation. The present study determined whether supplementation with arachidonic acid (AA) and/or docosahexaenoic acid (DHA) in rat milk formula (RMF) affects saturation of pulmonary surfactant phospholipids (PL). Beginning at 7 d of age, infant rats were artificially fed for 10 d with RMF supplemented with AA at 0, 0.5, and 1.0% of total fatty acid, or supplemented with DHA at 0, 0.5, and 1.0%, or cosupplemented with AA and DHA at levels of 0:0, 0.5:0.3, and 1.0:0.6% of the fat blend. Lung tissue PL contained 43 weight percent palmitate (16:0) of total fatty acids in infant rats fed the unsupplemented RMF. The supplementation with AA at both 0.5 and 1.0% decreased the weight percentage of 16:0 and stearate (18:0), indicating a decrease in saturation of PL. The observed decreases were accompanied by increases in AA and linoleic acid (18:2n-6). Surfactant phosphatidylcholine (PC) consisted of 71 weight percent 16:0 in the unsupplemented group, and this highly saturated PC was not altered by the cosupplementation with AA and DHA although there was a slight increase in DHA. Similarly, the cosupplementation did not change fatty acid composition of surfactant PL when compared with the unsupplemented group. The cosupplementation slightly decreased the weight percentage of 16:0 with a proportional increase in 18:0 leading to an unchanged weight percentage of total saturated fatty acids. These results suggest that, unlike lung tissue PL, the composition of saturated fatty acids in surfactant PL, particularly PC, is resistant to change by dietary AA and DHA supplementation. This, together with the unchanged concentration of total fatty acids in surfactant PC, indicates that LCPUFA cosupplementation causes no effect on pulmonary surfactant.

Animals↗
PubMed1983

Efficacy of antibiotic therapy in preventing spontaneous pregnancy loss among couples colonized with genital mycoplasmas.

Evidence suggests that genital mycoplasmas play a role in spontaneous abortion, prematurity, and perinatal morbidity and mortality rate. Since these organisms are sensitive to antibiotics, three treatment regimens were assessed for efficacy in preventing spontaneous pregnancy loss. The pregnancy loss rate was significantly reduced from 96% to 47.4% among those treated with doxycycline prior to conception and to less than 20% among those treated with erythromycin stearate only during pregnancy or with both regimens. The reduction in pregnancy loss rate was independent of maternal age, the number of previous abortions, gestational age at abortion, or other minor abnormalities. Diagnosis and treatment of mycoplasma infection following a first or later pregnancy loss equally prevented spontaneous loss in the next pregnancy. These observations suggest that appropriate treatment of mycoplasma infection could also prevent recurrent spontaneous abortion syndrome in couples with positive cultures.

Abortion, Habitual↗
PubMed1986

Quantitation of biological retinoids by high-pressure liquid chromatography: primary internal standardization using tritiated retinoids.

A single method is described for quantitation of 14 retinoids found in biological material. The method consists of reversed-phase HPLC, internal standardization, and carrier extraction procedures with three synthetic retinoids. Primary standardization of HPLC uv detector is achieved using tritiated all-trans-retinoic acid, all-trans-retinol, all-trans-retinyl palmitate, and all-trans-retinyl acetate. Extraction methods are standardized by correlating the uv absorbance of retinoids at 340 nm with radioactivity of tritiated retinoids of known specific activity. Quantitation of 10 pg of tritiated or 5 ng of nonradioactive retinoid per 0.1 g sample in a polarity range from 4-oxo-retinoic acid to retinyl stearate can be achieved in a single, 50-min chromatographic run. A single HPLC pump, a C18 reversed-phase analytical column, a multistep three-solvent gradient, and inexpensive solvents based on methanol, water, and chloroform comprise this cost-effective chromatographic system. Our primary standardization method allows investigators employing different procedures to compare results between laboratories by standardizing the HPLC uv detector with commercially available tritiated retinoids. With this method we were able to quantitate nanomolar amounts of endogenous retinoic acids and retinyl esters, that "HPLC uv only" conditions usually would not detect in the circulation and liver of rats under physiological conditions.

Animals↗
PubMed1986

Technique for injection into the sciatic nerve of the mouse for quantitative in vivo metabolic studies.

In this paper we describe a technique for intraneural injections, applicable to mouse peripheral nerves, which, compared with previous techniques, reduces trauma to the nerves and increases the level and reduces the variability of label recovery. Our technique employs glass needles (tip diameter, 50 micron) linked to a peristaltic pump by polyethylene tubing to inject small volumes (in the microliter range) of radiolabeled substrate solutions into mouse sciatic nerves, and allows the recovery of 20.9 +/- 1.9% (mean +/- standard deviation) and 30.5 +/- 4.8% of the injected radioactivity for 2 microliter [3H]acetate and 0.5 microliter of [3H]stearate, respectively.

Animals↗
PubMed1986

Temperature- and growth-phase-dependent changes in membrane fatty acid compositions of Brevibacterium ammoniagenes.

Fatty acids newly synthesized by Brevibacterium ammoniagenes grown at different temperatures were analyzed. The assay temperature, not the growth temperature, was found to be the major factor affecting the unsaturated/saturated ratio of newly synthesized fatty acids in logarithmic-phase cells. However, in the stationary-phase cells the growth temperature also affected the product profile significantly; cells grown at 7 degrees C produced relatively more oleate and stearate and less palmitate and hexadecenoate when shifted up to 37 degrees C than did cells grown and assayed at 37 degrees C. The unsaturated/saturated ratio as well as average chain length of fatty acids also varied along with the progress of isothermal growth phase. These changes in fatty acid product profiles observed in vivo could be mimicked in vitro assays of the fatty acid synthetase by changing malonyl-CoA concentrations. Our results suggest that the malonyl-CoA concentration is a factor which, in addition to temperature, determines growth-phase-dependent and growth-temperature-dependent changes in the unsaturated/saturated ratios of fatty acids.

Acetates↗