Influence of the rate and the share of water freezing on hydrogen and oxygen isotope separation

The influence of the rate v and the share of freezing water g on the separation of deuterium D and oxygen ¹⁸O was studied by mass spectrometry. Evidence was obtained supporting the well known facts that upon freezing of water, (1) the concentration of D in ice is higher than in water; (2) the degree of separation for D is higher than for ¹⁸O; (3) an increase in the concentration of D and ¹⁸O in ice takes place as the v value decreases. It was shown for the first time that, at g < 0.05, the concentrations of D at high v values are higher than at g > 0.05, and at low v values, they are less than at g > 0.05.     

Population dynamics of a Western corn rootworm (Coleoptera: Chrysomelidae) variant in east central Illinois commercial maize and soybean fields

Three on-farm sites in Iroquois County, IL, each containing an adjacent 16.2-ha commercial production maize, Zea mays L., and soybean, Glycine max (L.) Merr., field, were monitored for western corn rootworm, Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), adults from June through September 1999-2001. Mean captures of D. v. virgifera adults as measured with Pherocon AM yellow sticky traps were significantly greater in maize than in soybean. Overall mean numbers of D. v. virgifera adults captured with vial traps were significantly greater in soybean than in maize. Emergence cage data revealed that after 50% emergence of D. v. virgifera adults occurred, peak captures of D. v. virgifera adults occurred in maize as measured with vial and Pherocon AM traps. After maize reached the R2 (blister stage, 10-14 d after silking) stage of development and 90% emergence of D. v. virgifera adults had occurred, peak captures of D. v. virgifera adults were observed in soybean by using vial and Pherocon AM traps. Also, after maize reached the R2 stage of development, numbers of females significantly increased in soybean and decreased in maize. Captures of female D. v. virgifera adults frequently exceeded published economic thresholds in soybean, regardless of trap type used. Estimated survival of variant D. v. virgifera (egg to adult) in these commercial rotated maize fields was 10.7 and 9.4% from 1999 to 2000 and from 2000 to 2001, respectively. This compares with nonvariant D. v. virgifera survival estimates in continuous maize production systems in Iowa of 6.7 and 11% from 1983 to 1984 and from 1984 to 1985, respectively.     

A Reanalysis of Protein Polymorphism in Drosophila Melanogaster, D. Simulans, D. Sechellia and D. Mauritiana: Effects of Population Size and Selection

Comparison of synonymous and nonsynonymous variation/substitution within and between species at individual genes has become a widely used general approach to detect the effect of selection versus drift. The sibling species group comprised of two cosmopolitan (Drosophila melanogaster and Drosophila simulans) and two island (Drosophila mauritiana and Drosophila sechellia) species has become a model system for such studies. In the present study we reanalyzed the pattern of protein variation in these species, and the results were compared against the patterns of nucleotide variation obtained from the literature, mostly available for melanogaster and simulans. We have mainly focused on the contrasting patterns of variation between the cosmopolitan pair. The results can be summarized as follows: (1) As expected the island species D. mauritiana and D. sechellia showed much less variation than the cosmopolitan species D. melanogaster and D. simulans. (2) The chromosome 2 showed significantly less variation than chromosome 3 and X in all four species which may indicate effects of past selective sweeps. (3) In contrast to its overall low variation, D. mauritiana showed highest variation for X-linked loci which may indicate introgression from its sibling, D. simulans. (4) An average population of D. simulans was as heterozygous as that of D. melanogaster (14.4% v.s. 13.9%) but the difference was large and significant when considering only polymorphic loci (37.2% v.s. 26.1%). (5) The species-wise pooled populations of these two species showed similar results (all loci = 18.3% v.s. 20.0%, polymorphic loci = 47.2% v.s. 37.6%). (6) An average population of D. simulans had more low-frequency alleles than D. melanogaster, and the D. simulans alleles were found widely distributed in all populations whereas the D. melanogaster alleles were limited to local populations. As a results of this, pooled populations of D. melanogaster showed more polymorphic loci than those of D. simulans (48.0% v.s. 32.0%) but the difference was reduced when the comparison was made on the basis of an average population (29.1% v.s. 21.4%). (7) While the allele frequency distributions within populations were nonsignificant in both D. melanogaster and D. simulans, melanogaster had fewer than simulans, but more than expected from the neutral theory, low frequency alleles. (8) Diallelic loci with the second allele with a frequency less than 20% had similar frequencies in all four species but those with the second allele with a frequency higher than 20% were limited to only melanogaster the latter group of loci have clinal (latitudinal) patterns of variation indicative of balancing selection. (9) The comparison of D. simulans/D. melanogaster protein variation gave a ratio of 1.04 for all loci and 1.42 for polymorphic loci, against a ratio of approximately 2-fold difference for silent nucleotide sites. This suggests that the species ratios of protein and silent nucleotide polymorphism are too close to call for selective difference between silent and allozyme variation in D. simulans. In conclusion, the contrasting levels of allozyme polymorphism, distribution of rare alleles, number of diallelic loci and the patterns of geographic differentiation between the two species suggest the role of natural selection in D. melanogaster, and of possibly ancient population structure and recent worldwide migration in D. simulans. Population size differences alone are insufficient as an explanation for the patterns of variation between these two species.     

Emulsification using environmental compatible emulsifiers and de-emulsification using D.C. field and immobilized Nocardia amarae

As environmental compatible emulsifiers, various polysaccharides were investigated and de-emulsification methods were studied using filtration, direct current (D.C.) field, and also Nocardia amarae. 0.01% (v/v) of alginic acid, arabic gum, chitosan, and curdlan showed more than about 2 h of emulsion stability in the half-life of emulsion layer by a homogenizer and showed about twice synergic effects with other polysaccharides in emulsification activity at the ratio of 9:1 (v/v). De-emulsification by 110V D.C. field took about 1 h for the separation of oils from 1 l of oil-in-water emulsion in case of 0.01% arabic gum. Oil-water separator was designed using non-woven fabrics in filtration system and Nocardia amarae grown in n-hexadecane or kerosene was immobilized in the non-woven fabrics.     

The western corn rootworm diabrotica Virgifera virgifera en route to Germany

The western corn rootworm Diabrotica virgifera virgifera LeConte (Col.:Chrysomelidae) (D.v.v.) is one of the most important maize pests in North America. Ever since its invasion into Europe and its detection near Belgrade airport by BACA in 1993 it quickly spread all over southeastern Europe and is now advancing towards central Europe. Up until summer 2004 considered free of D.v.v., Germany is, with the exception of its northern and northeastern borders, surrounded by countries with proven D.v.v. infestations. In addition to simultaneous spot introductions by airplanes, three main routes for terrestrial introduction into Germany are likely: 1. from south to north via Lombardy (Italy) through Switzerland to the State of Baden-Wuerttemberg in the southwest; 2. from south east to northwest via Croatia, Slovenia, Austria into the State of Bavaria; and 3. from Belgium and the Netherlands in southeasterly direction to the state of Northrhine-Westfalia. From these, progress of D.v.v. along route 1 is so far the most advanced. It follows the well established network of road and rail connections through Switzerland and underscores the active role mankind and its technology plays as an active distribution vector for D.v.v. Mandatory crop rotation in the Swiss Canton of Ticino did slow down but could not prevent the northbound advance of D.v.v. in 2004. Considering the recent discovery of D.v.vu near the South German border, its introduction into German territory is only a matter of time and may be ecologically unavoidable. In Slovenla, another relatively small southern transit state, the D.v.v. population density is still much lower than in Switzerland but with significantly increasing trend during 2004 and with special emphasis in its southeastern provinces. Considering its relatively short distance to southeastern Bavaria and the well developed transalpine rail, road and tunnel system, Slovenia as a transit state may provide another access route for D.v.v. of lesser but still significant importance to Germany.     

Analysis of introgression of <Emphasis Type="Italic">Aegilops ventricosa</Emphasis> Tausch. genetic material in a common wheat background using C-banding

Seven Triticum aestivum (cv. Moisson)-Aegilops ventricosa addition lines and four VPM-1 lines were studied by C-banding, and compared with the parental common wheat cultivars Marne-Desprez (hereafter Marne), Moisson, and A. ventricosa lines 10 and 11. All of the VPM-1 lines had similar C-banding patterns and carried the same major 5B:7B translocation as the parental Marne cultivar. According to the C-banding analysis, the VPM-1 lines carry a complete 7D(7D(v)) chromosome substitution and a translocation involving the 5D and 5D(v) chromosomes. However, the translocation of the 2N(v)/6N(v) chromosome of A. ventricosa to the short arm of the 2A chromosome of wheat that had been identified in an earlier study using molecular analysis (Bonhomme A, Gale MD, Koebner RMD, Nicolas P, Jahier J, Bernard M in Theor Appl Genet 90:1042-1048, 1995; Jahier J, Abelard P, Tanguy AM, Dedryver F, Rivoal R, Khatkar S, Bariana HS Plant Breed 120:125-128, 2001) was not detected in our study. However, the appearance of a small pAs1 site at the tip of the chromosome 2A short arm in VPM-1 could be indicative of a minor translocation of the A. ventricosa chromosome. The 5B:7B translocation was also found in all seven T. aestivum-A. ventricosa addition lines, although it was not present in the parental common wheat cultivar Moisson. These lines showed different introgression patterns; besides the addition of the five N(v)-genome chromosomes, they also possessed different D(D(v)) genome substitutions or translocations. A whole arm translocation between chromosome 1N(v) and 3D(v) was identified in lines v86 and v137, and also in the A. ventricosa line 10. This observation lends further support to the idea that A. ventricosa line 10, rather than line 11, was used to develop a set of wheat A. ventricosa addition lines.     

Microbore high-performance liquid chromatographic determination of cisapride in rat serum samples using column switching

For the determination of cisapride from serum samples, an automated microbore high-performance liquid chromatographic method with column switching has been developed. After serum samples (100 μl) were directly injected onto a Capcell Pak MF Ph-1 pre-column (10×4 mm I.D.), the deproteinization and concentration were carried out by acetonitrile–phosphate buffer (20 mM, pH 7.0) (2:8, v/v) at valve position A. At 2.6 min, the valve was switched to position B and the concentrated analytes were transferred from MF Ph-1 pre-column to a C₁₈ intermediate column (35×2 mm I.D.) using washing solvent. By valve switching to position A at 4.3 min, the analytes were separated on a Capcell Pak C₁₈ UG 120 column (250×1.5 mm I.D.) with acetonitrile–phosphate buffer (20 mM, pH 7.0) (5:5, v/v) at a flow-rate of 0.1 ml/min. Total analysis time per sample was 18 min. The linearity of response was good (r=0.999) over the concentration range of 5–200 ng/ml. The within-day and day-to-day precision (CV) and inaccuracy were less than 3.7% and 3.8%, respectively. The mean recovery was 96.5±2.4% with the detection limit of 2 ng/ml.     

Enantioselectivity of debrisoquine 4-hydroxylation in Brazilian Caucasian hypertensive patients phenotyped as extensive metabolizers

Debrisoquine (D), an antihypertensive drug metabolized to 4-hydroxydebrisoquine (4-OHD) by CYP2D6, is commonly used as an in vivo probe of CYP2D6 activity and can be used to phenotype individuals as either extensive (EMs) or poor metabolizers (PMs) of such drugs as β-adrenergic blockers, tricyclic antidepressants, and class 1C antiarrhythmics. This report describes reversed-phase HPLC systems by which D and 4-OHD or S-(+) and R-(−)-4-OHD in urine are more selectively quantified without the need for derivatization techniques. We also studied the urinary excretion of R-(−)- and S-(+)-4-hydroxydebrisoquine in EM hypertensive patients in order to determine weather 4-OHD formation exhibits enantioselectivity. Twelve patients with mild to severe essential hypertension were admitted to the study. They received a single tablet of Declinax containing 10 mg debrisoquine sulfate. All the urine excreted during the following 8 h was collected. The debrisoquine metabolic ratio (DMR) was calculated as % of dose excreted as D/% of dose excreted as 4-OHD and the debrisoquine recovery ratio (DRR) was calculated as % of dose excreted as 4-OHD/% of dose excreted as D+4-OHD. Debrisoquine and its metabolite were determined in urine by HPLC using a reversed-phase Select B LiChrospher column, a mobile phase of 0.25 N acetate buffer, pH 5–acetonitrile (9:1, v/v) and a fluorescence detector. The limit of quantitation was determined to be 25.0 ng/ml for D and 18.75 ng/ml for 4-OHD. Intra- and inter-day relative standard deviations (RSDs) were less than 10%. All hypertensive patients studied showed a DMR of less than 12.6 or a DRR higher than 0.12 and were classified as EMs. Direct enantioselective separation on chiral stationary phase involved resolution of S-(+)-4-OHD and R-(−)-4-OHD on a Chiralcel OD-R column with a mobile phase of 0.125 N sodium perchlorate, pH 5–acetonitrile–methanol (85:12:3, v/v/v). The quantitation limit of each enantiomer was 3.75 ng/ml of urine. Intra- and inter-day RSDs were less than 10% for each enantiomer. A high degree of enantioselectivity in the 4-hydroxylation of D favouring the S-(+) enantiomer was observed, resulting in R-(−)-4-OHD not detected in the urine of the EM hypertensive patients studied.     

A multifaceted hemolymph defense against predation in Diabrotica virgifera virgifera larvae

The physical and chemical aspects of Diabrotica virgifera virgifera larval hemolymph were quantitatively assessed against two predatory beetle species in the laboratory. Adult Poecilus cupreus and Harpalus pensylvanicus (Coleoptera: Carabidae) were fed pupae, second or third instar D. v. virgifera or a palatable surrogate prey, i.e., Calliphora vicina or Sarcophaga bullata larvae (Diptera: Calliphoridae, Sarcophagidae, respectively) of equivalent size. The ethanol-soluble fraction of third instar D. v. virgifera hemolymph was extracted and suspended in a 0.24 M sucrose solution and offered to H. pensylvanicus (using a sucrose only control for comparison). The mean duration until first consumption was recorded for each predator, as was the amount of time spent eating, cleaning, resting, or walking for 2 min post-attack (or 5 min for the sugar assay). Maggots and D. virgifera larvae and pupae were attacked equally by both predators. But upon attack, D. v. virgifera larval hemolymph coagulated onto the mouthparts of the predators, which they began vigorously cleaning. Predators ate the sucrose solution for significantly longer than hemolymph + sucrose solution, indicating the presence of deterrent chemicals in the hemolymph. This research suggests that D. v. virgifera larvae are defended from predation by sticky and repellent hemolymph. We hypothesize that this defense partially explains the widespread success of D. v. virgifera as an invasive pest.     

Carboxylated ε-poly-L-lysine,a cryoprotective agent,is an effective partner of ethylene glycol for the vitrification of embryos at various preimplantation stages

It has been known that different protocols are used for embryo preservation at different stages due to different sensitivity to the physical and physiological stress caused by vitrification. In this study, we developed a common vitrification protocol using carboxlated ε-poly-l-lysine (COOH-PLL), a new cryoprotective agent for the vitrification of mouse embryos at different stages. The IVF-derived Crl:CD1(ICR) x B6D2F1/Crl pronuclear, 2-cell, 4-cell, and 8-cell, morula and blastocyst stage embryos were vitrified with 15% (v/v) ethylene glycol (EG) and 10% (w/v) COOH-PLL (E15P15) or 15% (v/v) EG and 15% (v/v) dimethyl sulfoxide (E15D15) using the minimal volume cooling method. The survival of vitrified embryos from pronuclear to blastocyst stages was equivalent between E15P15 and E15D15 groups. However, the rate of development to blastocysts was significantly lower in E15P15 than E15D15. The rates of survival and development to blastocysts were dramatically improved by a slight modification of EG and COOH-PLL concentrations (E20P10). After transferring 17 (E20P10) and 15 (E15D15) vitrified/warmed blastocysts, 8 and 7 pups were obtained (47.1% and 46.7%, respectively). Taken together, these results indicate that our vitrification protocol is appropriate for the vitrification of mouse embryos at different stages.     

High-performance liquid chromatographic assay for xanomeline, a specific M-1 agonist, and its metabolite in human plasma

A reversed-phase high-performance liquid chromatographic assay (HPLC) was utilized for monitoring xanomeline (LY246708/NNC 11–0232) and a metabolite, desmethylxanomeline, in human plasma. Xanomeline, desmethylxanomeline and internal standard were extracted from plasma with hexane at basic pH. The organic solvent extract was evaporated to dryness with nitrogen and the dried residue was reconstituted with 0.2 M HCl-methanol (50:50, v/v). A Zorbax CN 150 × 4.6 mm I.D., 5-μm column and mobile phase consisting of 0.5% (5 ml/l) triethylamine (TEA) adjusted to pH 3.0 with concentrated orthophosphoric acid-tetrahydrofuran (THF) (70:30, v/v) produced consistent resolution of analytes from endogenous co-extracted plasma components. Column effluent was monitored at 296 nm/0.008 a.u.f.s. and the assay limit of quantification was 1.5 ng/ml. A linear response of 1.5 to 20 ng/ml was sufficient to monitor plasma drug/metabolite concentrations during clinical trials. HPLC assay validation as well as routine assay quality control (QC) samples indicated assay precision/accuracy was better than ±15%.     

Quantitative relationships between the measurable transport resistances of membrane carriers: theory and experiment

A theoretical analysis is made of the possible quantitative relationships between the transport resistances that characterise membrane carrier systems. It is shown that there exist only five possible patterns in which to rank the four transport resistances. Symbolising these as A, B, C and D, the five possible patterns are (i) A = B = C = D; (ii) A = B much greater than C, D; (iii) A = B much greater than C = D; (iv) A = B = 2C = 2D; (v) A = 2B = 2C much greater than D. A survey of the available experimental data shows that pattern (ii) is the most prevalent, pattern (v) is often found and pattern (iii) has been identified. None of the ten transport systems so far analysed experimentally failed to fit one of the predicted patterns.     

Use of electrospray ionization liquid chromatography-mass spectrometry to study the role of CYP2D^ in the in vitro metabolism of 5-hydroxytryptamine receptor antagonists

An electrospray ionization liquid chromatographic-mass spectrometric (ESI-LC-MS) method has been developed to study the involvement of the cytochrome P450 isoenzyme CYP2D6 in the in vitro metabolism of the indole containing 5-hydroxytryptamine (5-HT₃) receptor antagonists tropisetron, ondansetron and dolasetron in human liver microsomes. Compounds were eluted using linear gradients of acetonitrile-20 mM ammonium acetate, solvent A, (10:90, v/v) (ph 6.0) and solvent B, (60:40, v/v) (pH 6.0) and a Nucleosil C₄ column. Microsomal incubations were analysed using selected ion monitoring of the molecular ion of parent drug and the molecular ion of hydroxylated metabolites. The involvement of CYP2D6 in drug metabolism was assessed by inhibition studies using quinidine (5 μM), a specific inhibitor of human CYP2D6, as well as by incubating compounds with microsomes prepared from celss transfected with cDNA encoding human CYP2D6. Results showed that the oxidation of all three compounds involved CYP2D6, but only that of tropisetron was inhibited by over 90% in the presence of quinidine. The present method can be applied to pre-clinical compounds, at an early stage of drug discovery, to assess the involvement of CYP2D6 in their metabolism and to screen for those compounds where CYP2D6 is the only isoenzyme implicated in the formation of major metabolites.     

异细胞质小麦与黑麦、小黑麦杂交对性状和减数分裂行为的影响

本文利用17种异细胞质“中国春”小麦与黑麦,小黑麦杂交,回交,研究其性状与减数分裂行为的表现。首次观察到D2型Ae.crassa.4x细胞质对同源染色体配对有抑制作用,对部分同源染色体配对有促进作用,SV型Ae.kotschyi细胞质对同源染色体,部分同源配对均有抑制作用,S1型Ae.sharonesis细胞质对部分同源染色体配对有促进,还观察到G型细胞质T.timopheevi,T.zhukovskyi,D2型细胞质,Ae.crassa 可提高产生有功能雌配子数,首次合成G型,SV型,D2型细胞质雄性不育的八倍体小黑麦,D2型细胞质八倍体小黑麦是光敏性雄性不育,在15小时以上的长光照条件下表现不育,这对进一步了解异细胞质的作用,小黑麦杂种优势的利用和小黑麦的改良均有重要意义。     

Immunocytochemical studies on the LHRH system of the Japanese quail: influence by photoperiod and aspects of sexual differentiation

Summary Immunocytochemistry was used to determine if photoperiod and/or sex have any effect on the pattern of the luteinizing hormone-releasing hormone (LHRH) system in the brain of the Japanese quail. Immunopositive perikarya were found within three major areas of the brain: the rostral paraolfactory lobe, the preoptic, and the septal region. A quantitative analysis of LHRH cell numbers was performed on male and female quail after two photoperiodic treatments: sexually mature birds exposed to 24 weeks of 20 h light: 4 h darkness (20L4D), and birds with a regressed reproductive system (induced by transfer from a photoregime of 20L4D to 25 short days of 8L16D). Two-way analysis of variance showed that short-day males display significantly (p < 0.05) more immunopositive perikarya (607 + 134) than long-day males (291 + 114), short-day females (293 + 103) or long-day females (330 + 92). The density of LHRH-immunoreactive nerve fibres and the intensity of the immunostaining in the median eminence were always greater in long-day sexually mature quail (male and female) than in animals exposed to 25 days of 8L16D. These results demonstrate that the LHRH system of the quail is influenced by photoperiod and mirrors sexual differentiation.     

Cryopreservation of Bangladeshi ram semen using different diluents and manual freezing techniques

A study was conducted to establish a sustainable and effective manual freezing technique for cryopreservation of Bangladeshi ram semen. Three diluents and freezing techniques were tested, both as treatment combinations (diluent × freezing technique) and fixed effects (diluent or freezing technique) on post-thaw sperm motility (SM), viability (SV), plasma membrane integrity (SPMI) and acrosome integrity (SAI). Ten rams were selected, based on semen evaluation. Eight ejaculates were used for each treatment combination. Semen samples were diluted using a two-step protocol for home-made Tris-based egg yolk (20%, v/v) diluents: D1 (7% glycerol, v/v) and D2 (5% glycerol, v/v), and one-step for commercial diluent: D3 (Triladyl^®, consists of bi-distilled water, glycerol, tris, citric acid, fructose, spectinomycin, lincomycin, tylosin and gentamycin) at 35 °C. Fraction-A (without glycerol) was added at 35 °C, and following cooling of sample to 5 °C (−0.30 °C/min), Fraction-B (with glycerol) was added. The diluted semen samples were aspirated into 0.25 ml French straws, sealed, and equilibrated at 5 °C for 2 h. The straws were frozen in liquid nitrogen (LN) vapour, in a Styrofoam box. The freezing techniques were; One-step (F1): at −15.26 °C/min from +5 °C to −140 °C; Two-step (F2): at −11.33 °C/min from +5 °C to −80 °C, and −30 °C/min from −80 °C-140 °C; and Three-step (F3): at −11.33 °C/min from +5 °C to −80 °C, at −26.66 °C/min from to −80 °C to −120 °C, and at −13.33 °C/min from −120 °C to −140 °C. Two semen straws from each batch were evaluated before and after freezing. The group F3D3 exhibited significantly higher (p < 0.05) post-thaw SM 63.1 ± 2.5%, SV 79.0 ± 2.1% and SPMI 72.9 ± 1.7%, whereas SAI 72.9 ± 1.7% was significantly higher (p < 0.05) in group F3D2. The freezing technique F2 and F3 had significantly higher (p < 0.05) post-thaw sperm values compared to F1. The post-thaw SM and SV were above 50% and 65% with the freezing technique F2 and F3 but differed non-significant. The SPMI 67.6 ± 2.0% and SAI 76.1 ± 1.4% were significantly higher (p < 0.05) with F3. Likewise, the diluent D2 and D3 had significantly higher (p < 0.05) post-thaw sperm values compared to D1. The post-thaw SM, SV and SPMI were above 50%, 65% and 55% with the diluents D2 and D3 but differed non-significant. The SAI 76.1 ± 1.1% was significantly higher (p < 0.05) with D3. We concluded that the use of a simple home-made Tris-based diluent containing 20% (v/v) egg yolk and 5% glycerol (v/v), two-step dilution and a three-step freezing technique is a sustainable and effective method for freezing ram semen. For further validation, the fertility of ewes artificially inseminated with the frozen semen will be observed.     

Energetics of kayaking

The metabolic cost of paddling at low speeds (v) was measured from oxygen uptake (VO2) and anaerobic glycolysis in an annular pool or calculated from submaximal VO2 measured at higher speeds when the kayaker was assisted in overcoming water resistance. Also calculated were the total drag (D) and the net mechanical efficiency (e). Each of the above variables was determined in male (n = 17) and female (n = 7) kayakers ranging in experience from beginners to elite. The VO2 increased with v to a peak of approximately 3.4 l.min-1 (80%-100% of peak VO2 during running) in men and of approximately 2.8 l.min-1 in women, while at higher speeds the additional energy was accounted for by anaerobic glycolysis. In all subjects the energy cost to paddle a given distance (C) increased according to a power function with increasing v. The C was lower for the elite male paddlers than for the unskilled group, while that for elite women was slightly less than that for the elite men. Also the rates of increase of C appeared to be inversely proportional to the subjects' skill. Total D for elite men increased from approximately 15 to 60 N over a range of speeds from 1 to 2.2 m.s-1 while those of unskilled men and skilled women for the same speed range were 10-20 N greater and slightly less, respectively. The e increased linearly, but at a different rate, with increases in v for the unskilled and the elite kayakers (males and females) being 4.2% and 6%, respectively, at v = 1.2 m.s-1.     

Enhancement of butanol tolerance and butanol yield in Clostridium acetobutylicum mutant NT642 obtained by nitrogen ion beam implantation

As a promising alternative biofuel, biobutanol can be produced through acetone/butanol/ethanol (ABE) fermentation. Currently, ABE fermentation is still a small-scale industry due to its low production and high input cost. Moreover, butanol toxicity to the Clostridium fermentation host limits the accumulation of butanol in the fermentation broth. The wild-type Clostridium acetobutylicum D64 can only produce about 13 g butanol/L and tolerates less than 2% (v/v) butanol. To improve the tolerance of C. acetobutylicum D64 for enhancing the production of butanol, nitrogen ion beam implantation was employed and finally five mutants with enhanced butanol tolerance were obtained. Among these, the most butanol tolerant mutant C. acetobutylicum NT642 can tolerate above 3% (v/v) butanol while the wide-type strain can only withstand 2% (v/v). In batch fermentation, the production of butanol and ABE yield of C. acetobutylicum NT642 was 15.4 g/L and 22.3 g/L, respectively, which were both higher than those of its parental strain and the other mutants using corn or cassava as substrate. Enhancing butanol tolerance is a great precondition for obtaining a hyper-yield producer. Nitrogen ion beam implantation could be a promising biotechnology to improve butanol tolerance and production of the host strain C. acetobutylicum.     

Sensitive isocratic liquid chromatographic assay for the determination of 5, 10, 15, 20-tetra(m-hydroxyphenyl)chlorin in plasma and tissue with electrochemical detection

A simple extraction procedure and a sensitive high-performance liquid chromatographic (HPLC) method are described for the determination of the photodynamic therapeutic agent 5, 10, 15, 20-tetra(m-hydroxyphenyl)chlorin (mTHPC) in plasma and tumour tissue. Reversed-phase high-performance liquid chromatography was performed on a C₁₈ column (70×4.6 mm I.D.) with a mobile phase of 0.01 M potassium dihydrogenphosphate buffer, pH 2.5-acetonitrile (55:45, v/v) and a coulometric detection (+0.80 V). The mean recoveries of mTHPC in the concentration ranges (5–2000 and 10–1000 ng/ml) were 90 and 89% for plasma and tumour samples, respectively. The procedure for plasma and tissue preparation involved solvent precipitation using methanol combined with ammonia solution and dimethyl sulphoxide (4, 0.2, 0.1, v/v/v) and (2, 0.1, 0.1, v/v/v) for plasma and tissue, respectively. For mTHPC at concentrations ranging from 5 to 2000 ng/ml, the within-day relative standard deviations, based on triplicate determinations were less than 8% and the between-day relative standard deviations calculated by performing extraction procedure of plasma samples on three different days ranged from 3 to 18%. This highly sensitive method, 5 and 10 ng/ml for plasma and tissue respectively, was applied successfully to the determination of mTHPC in mouse tumours for pharmacokinetic studies.     

Identification and characterization of CYP79D6v4, a cytochrome P450 enzyme producing aldoximes in black poplar (Populus nigra)

After herbivore feeding, poplar trees produce complex volatile blends containing terpenes, green leaf volatiles, aromatics, and nitrogen-containing compounds such as aldoximes and nitriles. It has been shown recently that volatile aldoximes released from gypsy moth (Lymantria dispar) caterpillar-damaged black poplar (Populus nigra) trees attract parasitoids that are caterpillar enemies. In western balsam poplar (P. trichocarpa), volatile aldoximes are produced by 2 P450 monooxygenases, CYP79D6v3 and CYP79D7v2. A gene fragment with high similarity to CYP79D6/7 was recently shown to be upregulated in herbivore-damaged leaves of P. nigra. In the present study we report the cloning and characterization of this gene, designated as CYP79D6v4. Recombinant CYP79D6v4 was able to convert different amino acids into the corresponding aldoximes, which were also found in the volatile blend of P. nigra. Thus, CYP79D6v4 is most likely involved in herbivore-induced aldoxime formation in black poplar.