The effects of different thymidine concentrations on DNA replication in pea-root cells synchronized by a protracted 5-fluorodeoxyuridine treatment

Single-cell and DNA fiber autoradiography, cytophotometry and velocity sedimentation in alkaline sucrose gradients were used to analyse DNA replication and nascent replicon maturation in 5-fluorodeoxyuridine (FUdR)-synchronized cells of Pisum sativum. The replicon size was not significantly changed by the protracted FUdR treatment. When the synchronized cells were released from the inhibitor, labeled with [³H]TdR for 30 min, and chased in medium containing 1 × 10⁻⁶ M or lower concentrations of cold thymidine, DNA replication stopped after approx. 25% of the genome had replicated, and the nascent strands failed to grow above 9–12 × 10⁶ D single-stranded (ss) DNA. When the cells were chased in medium with 1 × 10⁻⁵ M cold thymidine, the DNA content of the labeled cells steadily increased with time and the size of the nascent molecules grew continuously until replicon size was achieved; then they were accumulated at replicon size until the cells arrived in late S or G2. When the FUdR-synchronized cells were chased in medium containing 1 × 10⁻⁴ M cold thymidine, the size of the nascent strands increased continuously with time, indicating that some neighbouring nascent replicons were joined as soon as they completed their replication. These observations led us to postulate that in FUdR-synchronized cells the rates of chain elongation, cell progression through the S phase and nascent replicon maturation are controlled by thymidine availability.     

Effect of prostaglandin F2α on propulsive activity of the isolated segmental colon of the guinea-pig

The effects of prostaglandin F_2α (PGF_2α) on propulsive activity in segments of isolated colon and on isolated strips of guinea-pig colon were investigated.Using experimental conditions under which spontaneous propulsive activity was negligible, PGF_2α (5×10⁻⁸×1×10⁻⁶M), added to the bathing medium, increased propulsive activity in a concentration dependent manner. This increase of propulsive activity was abolished in the presence of atropine or tetrodotoxin (1×10⁻⁷g/ml).The contractions produced by PGF_2α(5×10⁻⁷ − 1×10⁻⁵M) in isolated longitudinal and circular smooth muscle strips of guinea-pig colon were unaffected in the presence of atropine or tetrodotoxin (1×10⁻⁷ g/ml).From these results it is concluded that under the conditions employed in this study propulsive activity stimulated by PGF_2α may depend on the contractions of both muscle layers and stimulation of the peristalic reflex.     

Control of flagellar motility in Euglena and Chlamydomonas : Microinjection of EDTA, EGTA, Mn, and Zn

When a Euglena, in a medium containing ATP, is microinjected with 7 × 10⁻¹⁴ l of 0.02 M EDTA, which binds Ca²⁺ and Mg²⁺, flagellar motility stops. Flagellar arrest in Chlamydomonas occurs with the injection of 2 × 10⁻¹⁴ l of 0.02 M EDTA. The injection of similar amounts (7 × 10⁻¹⁴ l in Euglena and 3 × 10⁻¹⁴ l in Chlamydomonas) of 0.02 M EGTA, which preferentially binds Ca²⁺, did not significantly alter flagellar motility. This suggests that a decrease in the internal Ca²⁺ concentration in Euglena or Chlamydomonas did not stimulate flagellar beating. Further, flagellar motility decreased when internal Mg²⁺ was chelated. The microinjection of Zn²⁺ into these cells caused a decrease in flagellar frequency analogous to the decrease in frequency caused by the injection of Ca²⁺ and EDTA. The microinjection of 7 × 10⁻¹⁴ l of 0.2 M Mn²⁺ caused an approx. 1.5-fold increase in Euglena flagellar motility. Chlamydomonas flagella, which cease to beat upon impalement in an Mg²⁺-free medium, resume a flagellar frequency of 18 Hz when injected with 3 × 10⁻¹⁴ l of 0.2 M Mn²⁺. In the experiments reported here, Mn²⁺ acts as an analog of Mg²⁺.     

Effects of arsenic exposure on the frequency of HPRT-mutant lymphocytes in a population of copper roasters in Antofagasta, Chile: a pilot study

A pilot biomarker study was conducted to investigate the feasibility of using the hypoxanthine guanine phosphoribosyltransferase (HPRT) gene in peripheral blood lymphocytes as a biomarker for detecting genetic effects of arsenic exposure. Blood and urine samples were obtained from workers highly exposed to arsenic in a copper roasting plant in Antofagasta, Chile. Individuals were classified according to their job titles into three potential exposure groups: high, medium, and low. To confirm exposure, arsenic concentration was determined in urine samples. The HPRT mutant frequencies were measured in lymphocytes from 15 individuals ranging in age from 24 to 66 years. The mean mutant frequencies for the three exposure groups were: low (9×10⁻⁶), medium (11×10⁻⁶), and high (24×10⁻⁶). An increased mutant frequency was observed in the highly exposed group, but the response was so slight that it is not likely that this assay will be capable of providing dose–response information across a range of lower, more typical environmental arsenic levels.     

Prostaglandin I2 as a potentiator of acute inflammation in rats

Prostaglandin I₂ potentiated the paw swelling induced by carrageenin in rats. Prostaglandin I₂ (0.1 μg) showed similar activity to PGE₁ (0.01 μg). This potentiating property disappeared in 60 minutes and was completely abolished by diphenhydramine (25 mg kg⁻¹, i.p.). In vascular permeability tests, PGI₂ itself (2.5 × 10⁻¹⁰ mol, 88 ng) caused no dye leakage reaction, but PGE₁ (2.5 × 10⁻¹⁰ mol, 88.5 ng) caused a significant dye leakage. This effect of PGE₁ was statistically significant compared with vehicle- or PGI₂-treated group (p<0.05). Prostaglandin I₂ potentiated the increased vascular permeability induced by 5-hydroxytriptamine (2.5 × 10⁻¹⁰ mol), bradykinin (5 × 10⁻¹⁰ mol) and histamine (2 × 10⁻¹⁰ to 2 × 10⁻⁸ mol). The potentiation was the most evidence in the case of histamine.     

Cell cycle-dependent expression of surface antigens in murine T-lymphoma cells : II. Murine leukemia virus gp70 increases in S phase

Studies were undertaken to identify cell surface markers specific for different phases of the cell cycle. Antisera were prepared in rabbits against membrane protein preparations from synchronized BW 5147 cells, an AKR mouse T-lymphoma cell line, in the G1, S, G2 or M phases of the cell cycle. These antisera were used to precipitate radioiodinated surface proteins from synchronized cells in the different phases. The immunoprecipitates were quantitatively analyzed by sodiumdodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Cells in S phase had significantly higher concentrations of proteins weighing 70 × 10³ and 165 × 10³ D than cells in G1 or G2 phase. The other major labeled surface components did not vary. These results were confirmed by quantitative absorption of the antisera with synchronized cells. Comparative analysis of the antisera showed that the 165 × 10³ D peak contained at least two antigens, one recognized by both a-G1 and a-S and the other by a-G1 only. Though cells in S phase had large quantities of the 70 × 10³ D protein, intact and SDS-solubilized membrane preparations from S phase could not elicit in rabbits any antibody against that protein. These antisera did, however, have good antibody titers to the other major protein peaks and the antisera developed against cells in G1, G2 or M had good anti-70 × 10³ activity. The results suggest a qualitative molecular change in the 70 × 10³ protein during S phase.     

Mutagenicity of bisphenol A and its suppression by interferon-α in human RSa cells

Bisphenol A is used as a monomer in the production of polycarbonate plastic products. The widespread use of bisphenol A has raised concerns about its effects in humans. Since there is little information on the mutagenic potential of the chemical, the mutagenicity of bisphenol A was tested using human RSa cells, which has been utilized for identification of novel mutagens. In genomic DNA from cells treated with bisphenol A at concentrations ranging from 1×10⁻⁷ to 1×10⁻⁵ M, base substitution mutations at K-ras codon 12 were detected using PCR and differential dot-blot hybridization with mutant probes. Mutations were also detected using the method of peptide nucleic acid (PNA)-mediated PCR clamping. The latter method enabled us to detect the mutation in bisphenol A-treated cells at a dose (1×10⁻⁸ M) equivalent to that typically found in the environment. Induction of ouabain-resistant (Oua^R) phenotypic mutation was also found in cells treated with 1×10⁻⁷ and 1×10⁻⁵ M of bisphenol A. The induction of K-ras codon 12 mutations and Oua^R mutations was suppressed by pretreating RSa cells with human interferon (HuIFN)-α prior to bisphenol A treatment. The cells treated with bisphenol A at the concentration of 1×10⁻⁶ M elicited unscheduled DNA synthesis (UDS). These findings suggested that bisphenol A has mutagenicity in RSa cells as well as mutagens that have been tested in these cells, and furthermore, that a combination of the PNA-mediated PCR clamping method with the human RSa cell line may be used as an assay system for screening the mutagenic chemicals at very low doses.     

Primary colonic epithelial cell culture of the rabbit producing prostaglandins

We have established primary colonic epithelial cell culture from adult rabbits and examined effects of anti-inflammatory drugs on prostaglandin (PG) E₂ production. Colonic epithelium of adult rabbits was scraped and minced into small pieces. They were incubated for isolation in Hanks' balanced salt solution with 0.35 % collagenase and Earle's solution with 1 mM EDTA. Isolated cells were cultured in Coon's modified Ham's F-12 medium with 10 % fetal bovine serum and antibiotics on collagen coated cell wells. The medium was refed twice a week. The production of PGs was assessed by high pressure liquid chromatography (HPLC). PGE₂ and PGF_2α were measured by radioimmunoassay. Within 24 hours after inoculation, the cell clumps attached to the surface of the wells and cells began to spread out and grow. Monolayer cultures became confluent in 4 days. Phase contrast microscopy showed that these cells consisted of a homogeneous population of epithelial cells with large oval nuclei, polyhedral shape, and organized sheet-like growth pattern. HPLC profile showed synthesis of 6-keto-PGF_1α, thromboxane B₂, PGF_2α, PGE₂, and PGD₂ by cultured cells. Quantitatively, 117±7 ng/mg-protein/hour PGE₂ by 7.4±0.7 ng/mg-protein/hour PGF_2α were produced. While hydrocortisone (10⁻⁴-10⁻² M) did not show a significant effect on PGE₂ production, indomethacin (10⁻⁸-10⁻⁶ M), and 5-aminosalicylic acid (2×10⁻⁴-5×10^{−3 M}) inhibited PGE₂ production. We have established relatively convenient procedure for primary culture of colonic epithelial cells from adult rabbits. Different actions of anti-inflammatory drugs on PGE₂ synthesis suggest that these cultured cells might be a good tool for the various cellular functional studies of normal colonic epithelial cells.     

Influences of cell size and aging on l-[H]norepinephrine binding sites in rat epididymal fat cells

[³H]norepinephrine binding to isolated rat fat cells was studied as a function of adipose cell age and size. Rats aged from 4 to 78 weeks were used.Scatchard analysis of norepinephrine binding revealed in old fat cells like in young ones the existence of two orders of binding sites with respectively high and low affinity for norepinephrine. The apparent association constants Ka₁ and Ka₂ associated with these binding sites did not differ consistently in the different groups of fat cells studied (Ka₁ = 1.7 to 2.2 × 10⁶ × M⁻¹; Ka₂ = 1.9 to 2.5 × 10⁴ × M⁻¹), suggesting that age and cell size do not modify the apparent affinity of norepinephrine-binding sites in rat fat cells.On the contrary, the total amount of norepinephrine bound to each of these sites was dependent upon cell age and size. In fact, maximum binding of norepinephrine to the high affinity sites was 0.9 and 9 pmol/10⁵ cells in small (diameter: 35 μ) and large (diameter: 105 μ) adipocytes, respectively, the values found for the low affinity sites being 13 and 135 pmol/10⁵ cells. When expressed per unit of fat cell area, however, the total binding capacity for these sites appeared practically constant (2.4 — 2.8 pmol × 10⁻³/mm² and 34.2 — 38.2 pmol × 10⁻³/mm² for the high and low affinity sites respectively). These data suggest that the total norepinephrine binding capacity of the fat cell is directly proportional to its surface.     

A nano-Ni based ultrasensitive nonenzymatic electrochemical sensor for glucose: Enhancing sensitivity through a nanowire array strategy

Highly ordered Ni nanowire arrays (NiNWAs) were synthesized for the first time using a template-directed electropolymerization strategy with a nanopore polycarbonate (PC) membrane template, and their morphological characterization were examined by scanning electron microscopy (SEM) and transmission electron microscope (TEM). A NiNWAs based electrode shows very high electrochemical activity for electrocatalytic oxidation of glucose in alkaline medium, which has been utilized as the basis of the fabrication of a nonenzymatic biosensor for electrochemical detection of glucose. The biosensor can be applied to the quantification of glucose with a linear range covering from 5.0 × 10⁻⁷ to 7.0 × 10⁻³ M, a high sensitivity of 1043 μA mM⁻¹ cm⁻², and a low detection limit of 1 × 10⁻⁷ M. The experiment results also showed that the sensor exhibits good reproducibility and long-term stability, as well as high selectivity with no interference from other oxidable species.     

Biodegradation of lindane by a native bacterial consortium isolated from contaminated river sediment

The aerobic biodegradation of lindane (γ-hexachlorocyclohexane) by a consortium of acclimated bacteria from sediment at a polluted site on the Suquia River, Cordoba, Argentina, is reported. The bacteria were acclimated for 30 days under aerobic conditions, using a minimal culture medium containing lindane (0.034 mM) as sole carbon source. Growth of the bacterial consortium decreased at a lindane concentration of 1.03 mM and was totally inhibited at 2.41 mM. The consortium showed initial lindane degradation rates of 4.92×10⁻³, 11.0×10⁻³ and 34.8×10⁻³ mM h⁻¹ when exposed to lindane concentrations of 0.069, 0.137 and 0.412 mM, respectively. Chloride concentration increased during aerobic biodegradation, indicating lindane mineralization. A metabolite identified as γ-2,3,4,5,6-pentachlorocyclohexene appeared during the first 24 h of biodegradation. Four different bacteria, identified as Sphingobacterium spiritivorum, Ochrobactrum anthropi, Bosea thiooxidans and Sphingomonas paucimobilis, were isolated. Pure strains of B. thiooxidans and S. paucimobilis degraded lindane after 3 days of aerobic incubation. This is the first report of lindane biodegradation by B. thiooxidans.     

Involvement of prostaglandins in the response of frog adrenocortical cells to muscarinic receptor activation

The role of prostaglandins (PGs) in the mechanism of action of acetylcholine (ACh) on frog adrenocortical cells has been examined. Administration of a single dose of ACh (5 × 10⁻⁵ M) to perifused frog interrenal fragments, for 20 min, stimulated the production of corticosterone, aldosterone, PGE₂ and 6-keto-PGF_1α. In contrast ACh did not significantly alter TXB2 production. The effect of ACh could be mimicked by muscarine (10⁻⁵ M). Conversely, nicotine (10⁻⁶ to 10⁻⁴ M) was totally inactive. The increase in PG biosynthesis preceded the peak of corticosteroid release. Repeated 20-min pulses of ACh (5 × 10⁻⁵ M) or muscarine (10⁻⁵ M) given at 130-min intervales induced a desensitization phenomenon. In presence of indomethacin (5 × 10⁻⁶ M), the effect of ACh on PG and steroid secretion was totally abolished. In calcium-free medium, the effect of ACh on PG and corticosteroid production was completely blocked. These results indicated that, in the frog, ACh stimulates corticosteroid secretion through a PG-dependent mechanism.     

High-Frequency Phage-Mediated Gene Transfer among Escherichia coli Cells, Determined at the Single-Cell Level

Recent whole-genome analysis suggests that lateral gene transfer by bacteriophages has contributed significantly to the genetic diversity of bacteria. To accurately determine the frequency of phage-mediated gene transfer, we employed cycling primed in situ amplification-fluorescent in situ hybridization (CPRINS-FISH) and investigated the movement of the ampicillin resistance gene among Escherichia coli cells mediated by phage at the single-cell level. Phages P1 and T4 and the newly isolated E. coli phage EC10 were used as vectors. The transduction frequencies determined by conventional plating were 3 × 10⁻⁸ to 2 × 10⁻⁶, 1 × 10⁻⁸ to 4 × 10⁻⁸, and <4 × 10⁻⁹ to 4 × 10⁻⁸ per PFU for phages P1, T4, and EC10, respectively. The frequencies of DNA transfer determined by CPRINS-FISH were 7 × 10⁻⁴ to 1 × 10⁻³, 9 × 10⁻⁴ to 3 × 10⁻³, and 5 × 10⁻⁴ to 4 × 10⁻³ for phages P1, T4, and EC10, respectively. Direct viable counting combined with CPRINS-FISH revealed that more than 20% of the cells carrying the transferred gene retained their viabilities. These results revealed that the difference in the number of viable cells carrying the transferred gene and the number of cells capable of growth on the selective medium was 3 to 4 orders of magnitude, indicating that phage-mediated exchange of DNA sequences among bacteria occurs with unexpectedly high frequency.     

Cytochalasin B blocks sperm incorporation but allows activation of the sea urchin egg

Cytochalasin B (CB) (2 × 10⁻⁶ M) prevents the incorporation of sperm into the eggs of Lytechinus pictus and Strongylocentrotus purpuratus as judged by light and transmission electron microscopy (TEM). At lower concentrations of CB (2 × 10⁻⁷ M), sperm are successfully incorporated into the egg, but their migration in the area of the egg cortex is impaired. The site of action of CB on the sperm may be on the initial rotation of the sperm nucleus in the cortex; the subsequent migration is not affected by CB. Although sperm incorporation is prevented at the higher CB concentrations, the eggs become activated—as judged by cortical reaction, increased protein synthesis and increased respiration. These findings raise the concept that egg activation by sperm could result from some pre-fusion event and hence that sperm-egg fusion would not be a prerequisite for the triggering of development. An alternative hypothesis is that fusion occurs between the acrosome process membrane and egg membrane, but since CB has destroyed the integrity of the cortex actin, the fusion bridge is so weak that it cannot be maintained without some contractile or cytoskeletal support by the cortex. The sperm may activate the CB-treated egg in the same manner as pricking with a microelectrode sometimes does.     

Membrane Permeability Characteristics of Metaphase II Mouse Oocytes at Various Temperatures in the Presence of Me2SO

In this study, the hydraulic conductivity (L_p), Me₂SO permeability ( _Me2SO), and the reflection coefficients (ς) and their activation energies were determined for Metaphase II (MII) mouse oocytes by exposing them to 1.5 M Me₂SO at temperatures of 30, 20, 10, 3, 0, and −3°C. These data were then used to calculate the intracellular concentration of Me₂SO at given temperatures. Individual oocytes were immobilized using a holding pipette in 5 μl of an isosmotic PBS solution and perfused with precooled or prewarmed 1.5 M Me₂SO solutions. Oocyte images were video recorded. The cell volume changes were calculated from the measurement of the diameter of the oocytes, assuming a spherical shape. The initial volume of the oocytes in the isoosmotic solution was considered 100%, and relative changes in the volume of the oocytes after exposure to the Me₂SO were plotted against time. Mean (means ± SEM) L_pvalues in the presence of Me₂SO ( ^Me₂SO_p) at 30, 20, 10, 3, 0, and −3°C were determined to be 1.07 ± 0.03, 0.40 ± 0.02, 0.18 ± 0.01, 7.60 × 10⁻²± 0.60 × 10⁻², 5.29 × 10⁻²± 0.40 × 10⁻², and 3.69 × 10⁻²± 0.30 × 10⁻²μm/min/atm, respectively. The _Me2SOvalues were 3.69 × 10⁻³± 0.3 × 10⁻³, 1.07 × 10⁻³± 0.1 × 10⁻³, 2.75 × 10⁻⁴± 0.15 × 10⁻⁴, 7.83 × 10⁻⁵± 0.50 × 10⁻⁵, 5.24 × 10⁻⁵± 0.50 × 10⁻⁵, and 3.69 × 10⁻⁵± 0.40 × 10⁻⁵cm/min, respectively. The ς values were 0.70 ± 0.03, 0.77 ± 0.04, 0.81 ± 0.06, 0.91 ± 0.05, 0.97 ± 0.03, and 1 ± 0.04, respectively. The estimated activation energies (E_a) for ^Me₂SO_p, _Me2SO, and ς were 16.39, 23.24, and −1.75 Kcal/mol, respectively. These data may provide the fundamental basis for the development of more optimal cryopreservation protocols for MII mouse oocytes.     

Laccase-catalyzed oxidation of phenolic compounds in organic media

Rhus vernificera laccase-catalyzed oxidation of phenolic compounds, i.e., (+)-catechin, (−)-epicatechin and catechol, was carried out in selected organic solvents to search for the favorable reaction medium. The investigation on reaction parameters showed that optimal laccase activity was obtained in hexane at 30 °C, pH 7.75 for the oxidation of (+)-catechin as well as for (−)-epicatechin, and in toluene at 35 °C, pH 7.25 for the oxidation of catechol. E_a and Q₁₀ values of the biocatalysis in the reaction media of the larger log p solvents like isooctane and hexane were relatively higher than those in the reaction media of lower log p solvents like toluene and dichloromethane. Maximum laccase activity in the organic media was found with 6.5% of buffer as co-solvent. A wider range of 0–28 μg protein/ml in hexane than that of 0–16.7 μg protein/ml in aqueous medium was observed for the linear increasing conversion of (+)-catechin. The kinetic studies revealed that in the presence of isooctane, hexane, toluene and dichloromethane, the K_m values were 0.77, 0.97, 0.53 and 2.9 mmol/L for the substrate of (+)-catechin; 0.43, 0.34, 0.14 and 3.4 mmol/L for (−)-epicatechin; 2.9, 1.8, 0.61 and 1.1 mmol/L for catechol, respectively, while the corresponding V_max values were 2.1 × 10⁻², 2.3 × 10⁻², 0.65 × 10⁻² and 0.71 × 10⁻² δA/μg protein min); 1.8 × 10⁻², 0.88 × 10⁻², 0.19 × 10⁻² and 1.0 × 10⁻² δA/μg protein min); 0.48 × 10⁻², 0.59 × 10⁻², 0.67 × 10⁻² and 0.54 × 10⁻² δA/μg protein min), respectively. FT-IR indicated the formation of probable dimer from (+)-catechin in organic solvent. These results suggest that this laccase has higher catalytic oxidation capacity of phenolic compounds in suitable organic media and favorite oligomers could be obtained.     

Clonal selection by fluorescence redistribution after photobleaching (FRAP)—A “fast” lateral mobility fibroblast mutant (E7G1)

Fluorescence redistribution after photobleaching (FRAP) was utilized to select a “fast” lateral mobility clone from Kirsten murine sarcoma virus-transformed 3T3 (KMSV-3T3) fibroblasts. The clone, E7G1, demonstrated a lateral mobility for membrane wheat germ agglutinin (WGA) and succinylated concanavalin A (sCon A) receptors of (2.1 ± 1.6) × 10⁻⁹ cm²/s and (2.7 ± 2.3) × 10⁻⁹ cm²/s, respectively. These mobilities were approximately equivalent to phospholipid mobility (2.8 ± 1.9 × 10⁻⁹ cm²/s). The fast mobility phenotype is observed when the cells are unattached and spherical. Upon attachment, the mobility decreases to (0.19 ± 0.19) × 10⁻¹⁰ cm²/s. In addition, the ability of Con A to initiate global modulation was completely lost in spread as well as spherical cells in the E7G1 fast mobility clone. A comparison of F-actin patterns between untransformed Balb/c fibroblasts and the E7G1-transformed line suggests a correlation between well-developed stress fiber assemblies and the ability to induce global modulation. The fast mobility clone was stable for at least 23 passages.     

Genetic variation at the FADS1-FADS2 gene locus influences delta-5 desaturase activity and LC-PUFA proportions after fish oil supplement

Delta-5 and delta-6 desaturases (D5D and D6D) are key enzymes in endogenous synthesis of long-chain PUFAs. In this sample of healthy subjects (n = 310), genotypes of single nucleotide polymorphisms (SNPs) rs174537, rs174561, and rs3834458 in the FADS1-FADS2 gene cluster were strongly associated with proportions of LC-PUFAs and desaturase activities estimated in plasma and erythrocytes. Minor allele carriage associated with decreased activities of D5D (FADS1) (5.84 × 10⁻¹⁹ ≤ P ≤ 4.5 × 10⁻¹⁸) and D6D (FADS2) (6.05 × 10⁻⁸ ≤ P ≤ 4.20 × 10⁻⁷) was accompanied by increased substrate and decreased product proportions (0.05 ≤ P ≤ 2.49 × 10⁻¹⁶). The significance of haplotype association with D5D activity (P = 2.19 × 10⁻¹⁷) was comparable to that of single SNPs, but haplotype association with D6D activity (P = 3.39 × 10⁻²⁸) was much stronger. In a randomized controlled dietary intervention, increasing eicosapentaenoic acid (EPA, 20:5n-3) and docosahexaenoic acid (DHA, 22:6n-3) intake significantly increased D5D (P = 4.0 × 10⁻⁹) and decreased D6D activity (P = 9.16 × 10⁻⁶) after doses of 0.45, 0.9, and 1.8 g/day for six months. Interaction of rs174537 genotype with treatment was a determinant of D5D activity estimated in plasma (P = 0.05). In conclusion, different sites at the FADS1-FADS2 locus appear to influence D5D and D6D activity, and rs174537 genotype interacts with dietary EPA+DHA to modulate D5D.     

Calculation of Cell Production from [3H]Thymidine Incorporation with Freshwater Bacteria

The conversion factor for the calculation of bacterial production from rates of [³H]thymidine incorporation was examined with diluted batch cultures of freshwater bacteria. Natural bacterial assemblages were grown in aged, normal, and enriched media at 10 to 20°C. The generation time during 101 growth cycles covered a range from 4 to >200 h. The average conversion factor was 2.15 × 10¹⁸ cells mol^-1 of thymidine incorporated into the trichloroacetic acid (TCA) precipitate (standard error = 0.29 × 10¹⁸; n = 54), when the generation time exceeded 20 h. At generation times of <20 h, the average conversion factor was 11.8 × 10¹⁸ cells mol^-1 of thymidine incorporated into TCA precipitate (standard error = 1.72 × 10¹⁸; n = 47). The amount of radioactivity in purified DNA increased with decreasing generation time and increasing conversion factor (calculated from the TCA precipitate), corresponding to a decrease in the percentage in protein. The conversion factors calculated from purified DNA or from the TCA precipitate gave the same variability. Conversion factors did not change significantly with the medium, but were significantly higher at 20°C than at 15 and 10°C. A detailed examination of the [³H]thymidine concentrations that were needed to achieve maximum labeling in DNA was carried out 6 times during a complete growth cycle. During periods with low generation times and high conversion factors, 15 nM [³H]thymidine was enough for the maximum labeling of the TCA precipitate. This suggests that incorporation of [³H]thymidine into DNA is probably limited by uptake during periods with generation times of <20 h and that freshwater bacterioplankton cell production sometimes is underestimated when a conversion factor of 2.15 × 10¹⁸ cells mol^-1 of thymidine incorporated is used.     

Graft copolymerization of N-vinylformamide onto sodium carboxymethylcellulose and study of its swelling, metal ion sorption and flocculation behaviour

The present paper reports the graft copolymerization of N-vinylformamide onto sodium carboxymethylcellulose by free radical polymerization using potassium peroxymonosulphate/thiourea redox system in an inert atmosphere. The reaction conditions for maximum grafting have been optimized by varying the reaction variables, including the concentration of N-vinylformamide (12.0 × 10⁻²–28.0 × 10⁻² mol dm⁻³), potassium peroxymonosulphate (4.0 × 10⁻³–12.0 × 10⁻³ mol dm⁻³), thiourea (1.2 × 10⁻³–4.4 × 10⁻³ mol dm⁻³), sulphuric acid (2.0 × 10⁻³–10.0 × 10⁻³ mol dm⁻³), sodium carboxymethylcellulose (0.2–1.8 g dm⁻³) along with time duration (60–180 min) and temperature (25–45° C). Water swelling capacity, metal ion sorption and flocculation studies of synthesized graft copolymer have been performed with respect to the parent polymer. The graft copolymer has been characterized by FTIR spectroscopy and thermogravimetric analysis.