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Sex dimorphism and gene expression were studied in developing catkins in 159 F2 individuals from the bioenergy crop Salix purpurea, and potential mechanisms and pathways for regulating sex development were explored. Differential expression, eQTL, bisulfite sequencing, and network analysis were used to characterize sex dimorphism, detect candidate master regulator genes, and identify pathways through which the sex determination region (SDR) may mediate sex dimorphism. Eleven genes are presented as candidates for master regulators of sex, supported by gene expression and network analyses. These include genes putatively involved in hormone signaling, epigenetic modification, and regulation of transcription. eQTL analysis revealed a suite of transcription factors and genes involved in secondary metabolism and floral development that were predicted to be under direct control of the sex determination region. Furthermore, data from bisulfite sequencing and small RNA sequencing revealed strong differences in expression between males and females that would implicate both of these processes in sex dimorphism pathways. These data indicate that the mechanism of sex determination in Salix purpurea is likely different from that observed in the related genus Populus. This further demonstrates the dynamic nature of SDRs in plants, which involves a multitude of mechanisms of sex determination and a high rate of turnover.

Short rotation woody biomass cultivars developed from fast-growing shrub species of willow (Salix spp.) have superior properties as perennial energy crops for the Northeast and Midwest US. However, the insect pest potato leafhopper (PLH) Empoasca fabae (Harris) can cause serious damage and reduce yield of susceptible genotypes. Currently, the willow cultivars in use display varying levels of susceptibility under PLH infestation. However, genes and markers for resistance to PLH are not yet available for marker-assisted selection in breeding. In this study, transcriptome differences between a resistant genotype 94006 (S. purpurea) and a susceptible cultivar 'Jorr' (S. viminalis), and their hybrid progeny were determined. Over 600 million RNA-Seq reads were generated and mapped to the Salix purpurea reference transcriptome. Gene expression analyses revealed the unique defense mechanism in resistant genotype 94006 that involves PLH-induced secondary cell wall modification. In the susceptible genotypes, genes involved in programed cell death were highly expressed, explaining the necrosis symptoms after PLH feeding. Overall, the discovery of resistance genes and defense mechanisms provides new resources for shrub willow breeding and research in the future.

a b s t r a c t Studies of arsenate and phosphate uptake by plants in hydroponic and soil systems indicate a common transport mechanism via the phosphate transporters (PHTs) due to structural similarity of the anions. Typically, the presence of phosphate decreases plant uptake and translocation of arsenate in hydroponic solution. This study quantified arsenic (As) uptake related to the presence of phosphorus in an As-tolerant willow (Salix viminalis × Salix miyabeana) and an As-sensitive willow (Salix eriocephala). Addition of phos- phate resulted in greater As accumulation than in treatments without phosphate in both genotypes, although the tolerant genotype accumulated more As than the sensitive one. Expression of genes for two putative high-affinity phosphate transporters, PHT1;3 and PHT1;12, were up-regulated in both wil- low genotypes upon addition of As, but to a greater extent in the As-sensitive genotype. Expression of a third putative transporter, PHT1;4, was greater in the As-sensitive genotype but was not up-regulated as a result of As addition in either genotype. Leaves of the As-tolerant willow genotype contained greater concentrations of -glutamylcysteine (EC) than the sensitive genotype, although this was not due to dif- ferential expression of the � -glutamylcysteine synthetase (� ECS) gene. The results suggest that increased expression of PHT1 upon exposure to As in an As-sensitive genotype contributes to rapid toxicity. Our data suggest that although detoxification capacity may be different between genotypes, the differences are not due to up-regulation of ECS or phytochelatin synthase.

The leaf cuticular waxes of six Salix clones (one Salix miyabeana, one Salix dasyclados, one Salix eriocephala, two Salix purpurea, and one interspecific hybrid of Salix eriocephala x interior) with different biomass productivities were characterized by gas chromatography-mass spectrometry. Total wax content ranged from 6.3 to 16.8 microg cm(-2), and two distinct patterns of wax were measured. The wax from leaves of S. dasyclados 'SV1' differed from all other clones and was dominated by fatty acids (42%), high concentrations of n-alkanes (25%) and n-alcohols (28%), with low n-aldehyde content (4%). All other clones produced cuticular wax dominated by n-alcohols (32-51%), particularly 1-hexacosanol, with fatty acids (14-37%) and n-aldehydes (19-26%) present in lower abundances. Clones of Salix grown under identical environmental conditions produce noticeably different amounts of cuticular wax. In contrast to previous studies of Salix, total wax content was independent of biomass productivity, measured as basal area, suggesting that wax production is not directly linked with woody biomass production by shrub willows under these site conditions.

Water flux across cell membranes has been shown to occur not only through the lipid bilayer, but also through aquaporins, which are members of the major intrinsic protein (MIP) super-family of channel proteins. Aquaporins greatly increase the membrane permeability for water, but may also be regulated, allowing cellular control over the rate of water influx/efflux. Water flux is crucial for stomatal opening and closing, but little is known about the role that aquaporins play in stomatal physiology. Our initial goal was to isolate and characterize the MIP genes expressed in guard cells of the model plant, Nicotiana glauca. Degenerate oligonucleotides corresponding to amino acid sequences conserved in tonoplast intrinsic proteins (TIPs) or plasma membrane intrinsic proteins (PIPs) were used to amplify portions of MIP genes by RT-PCR. These PCR products were used as probes in screening a N. glauca guard cell cDNA library. We isolated three clones (NgMIP1, NgMIP2 and NgMIP3) homologous to TIPs and two clones (NgMIP4 and NgMIP5) homologous to PIPs. All of the MIP genes we characterized displayed highest levels of mRNA accumulation in roots or stems, with lower levels of expression in mesophyll cells and whole leaves, and lowest transcript accumulation in guard cell RNA. Interestingly, the accumulation of transcripts arising from NgMIP2, NgMIP3 and NgMIP4 diminished dramatically in drought-stressed plants. This down-regulation of MIP gene expression may result in reduced membrane water permeability and may encourage cellular water conservation during periods of dehydration stress.

Two probiotic strains of bacteria Bifidobacterium lactis DR10 and Lactobacillus rhamnosus DR20 were tested for their ability to utilise and grow on galacto-oligosaccharides present in a commercial hydrolysed lactose milk powder. The results clearly demonstrated that B. lactis DR10 preferentially utilises tri- and tetra-saccharides whereas Lb. rhamnosus DR20 prefers sugars with a lower degree of polymerisation, i.e., disaccharides and monosaccharides. Since galacto-oligosaccharides are non-digestible oligosaccharides, this in vitro data suggest that galacto-oligosaccharides present in milk powders are likely to promote growth of DR10 and DR20 in vivo if these strains are consumed in combination with the milk powder. Fifty four strains of lactic acid bacteria, including members from the genera Bifidobacterium and Lactobacillus, were studied for their ability to utilise lactose derived oligosaccharides. A perfect correlation was observed between the ability of a strain to utilise oligosaccharide and the presence of the lactose hydrolysing enzyme β-galactosidase. Based on these observations, a mechanism for the utilisation of galacto-oligosaccharides in genus bifidobacteria is proposed that may help to explain the ability of these organisms to out-compete other bacteria in the ecosystem of the human gastro-intestinal tract.

Vanadium was complexed with 2,3-dihydroxynaphthalene (DHN), absorbed onto a hanging mercury drop electrode, and measured using square wave stripping voltammetry. The reduction current of the adsorbed complex was greatly increased (100 times) by the addition of bromate ion. Preconcentration of the vanadium complex was done at a potential of −0.1 V vs. Ag AgCl at pH 4.8 in the presence of 0.1 M DHN and 5 mM NaBrO 3 . The detection limit for a 1 min adsorption time was 1.5 × 10 −11 M, and the reduction current was linear over the range of 5 × 10 −11 to 4 × 10 −9 M. The relative standard deviation was calculated as 1% ( n = 7) for 1 × 10 −9 M. The method was applied to the determination of vanadium in natural water samples.

A method is presented for the measurement of arsenic(III) in natural waters using square wave cathodic stripping voltammetry at a hanging mercury drop electrode. Pre-concentration was carried out in 2 M HC1 in, the presence of 0.8 mM CuCl2 at a potential of −0.4 V vs.AgAgCl. The deposited intermetallic compound was reduced at a potential of about − 0.8 V vs. AgAgCl. The square wave frequency was 70 Hz with a staircase step of 2 mV and a pulse height of 25 mV. The peak current decreased with scan number, however, this effect can be minimized by the addition of hydrazine. The detection limit is approximately 0.06 nM (5 parts per trillion) when a 10 min deposition time was used. The relative standard deviation was calculated as 8% (n = 11) at 1 × 10−9 M arsenic for l min period of deposition. The method was tested by the measurement of arsenic(III) in both river and sea water.

In photosystem I (PSI) of oxygenic photosynthetic organisms the psaC polypeptide, encoded by the psaC gene, provides the ligands for two [4Fe-4S] clusters, FA and FB. Unlike other cyanobacteria, two different psaC genes have been reported in the cyanobacterium Synechocystis 6803, one (copy 1) with a deduced amino acid sequence identical to that of tobacco and another (copy 2) with a deduced amino acid sequence similar to those reported for other cyanobacteria. Insertion of a gene encoding kanamycin resistance into copy 2 resulted in a photosynthesis-deficient strain, CDK25, lacking the PsaC, PsaD and PsaE polypeptides in isolated thylakoid membranes, while the PsaA/PsaB and PsaF subunits were found. Growth of the mutant cells was indistinguishable from that of wild-type cells under light-activated heterotrophic growth (LAHG). A reversible P700+ signal was detected by EPR spectroscopy in the isolated thylakoids during illumination at low temperature. Under these conditions, the EPR signals attributed to FA and FB were absent in the mutant strain, but a reversible Fx signal was present with broad resonances at g = 2.079, 1.903, and 1.784. Addition of PsaC and PsaD proteins to the thylakoids gave rise to resonances at g = 2.046, 1.936, 1.922, and 1.880; these values are characteristic of an interaction-type spectrum of FA- and FB-. In room-temperature optical spectroscopic analysis, addition of PsaC and PsaD to the thylakoids also restored a 30 ms kinetic transient which is characteristic of the P700+ [FA/FB]- backreaction. Expression of copy 1 was not detected in cells grown under LAHG and under mixotrophic conditions. These results demonstrate that copy 2 encodes the PsaC polypeptide in PSI in Synechocystis 6803, while copy 1 is not involved in PSI; that the PsaC polypeptide is necessary for stable assembly of PsaD and PsaE into PSI complex in vivo; and that PsaC, PsaD and PsaE are not needed for assembly of PsaA-PsaB dimer and electron transport from P700 to Fx.

We have taken a genetic approach to eliminating the presence of photosystem I (PSI) in site-directed mutants of photosystem II (PSII) in the cyanobacterium Synechocystis sp. PCC 6803. By selecting under light-activated heterotrophic conditions, we have inactivated the psaA-psaB operon encoding the PSI reaction center proteins in cells containing deletions of the three psbA genes. We have also introduced deletions into both copies of psbD in a strain containing a mutation that inactivates psaA (ADK9). These strains, designated D1-/PSI- and D2-/PSI-, may serve as recipient strains for the incorporation of site-directed mutations in either psbA2 or psbD1. The characterization of these cells, which lack both PSI and PSII, is described.

SummarySpectrophotometric assays ofβ–galactosidase (EC 3.2.1.23) and phospho-β–galactosidase (EC 3.2.1.85) activity were used to survey the lactose utilization pathways of lactic acid bacteria and bifidobacteria.β–Galactosidase activity was found in all six genera represented (Lactococcus, Streptococcus, Leuconostoc, Lactobacillus, PediococcusandBifidobacterium) while phospho-β–galactosidase was restricted to the lactococci, twoLactobacillusand twoLeuconostocspecies. A number of strains ofLactococcus lactis, Lactobacillus caseiandLeuconostocspp. contained both enzymes. Enzyme activities varied when cells were grown on different sugars, but in general were low or absent for cells grown on glucose compared with lactose. Two lactose-related compounds, lactulose and galactosyl lactose, believed to be specific growth factors for bifidobacteria, supported growth amongst a wide range of lactic acid bacteria in addition to bifidobacteria. Growth on galactosyl lactose was restricted to some but not all strains containingβ–galactosidase, implying that the presence ofβ–galactosidase is insufficient by itself to ensure utilization of galactosyl lactose. DNA fragments that encoded theLactococcus lactissubsp.cremorisphospho-β–galactosidase gene or theβ–galactosidase genes ofStreptococcus salivariussubsp.thermophilusorLactobacillus delbrueckiisubsp.bulgaricuswere isolated and used as probes in DNA-DNA hybridizations. Little or no hybridization was detected between these probes and plasmid or genomic DNA isolated from heterologous species, despite the presence of the corresponding enzyme activity in the strains probed.

The distribution of chromogranin/secretogranin (Cg/Sg) mRNAs, determined by Northern and in situ hybridization, was analyzed in 14 cultured pituitary adenomas characterized by immunohistochemistry and hormone secretion in a defined medium in vitro. There were 5 functional GH adenomas, 1 silent GH adenoma, 7 null cell adenomas, and 1 oncocytoma. The null cell adenomas, oncocytoma, and silent GH adenomas were also analyzed by electron microscopy. Most null cell adenomas and the oncocytoma secreted FSH and LH into the culture medium. GH adenomas, which are examples of well differentiated tumors based on morphological examination, expressed significantly more SgIII mRNA compared to the null cell adenomas and oncocytoma (70 +/- 6% vs. 22 +/- 5%; P0.001). GH adenomas also expressed significantly less CgA mRNA compared to the less well differentiated null cell adenomas and oncocytoma (27 +/- 6% vs. 67 +/- 4%; P0.001), which could be considered less well differentiated based on ultrastructural morphological features. After treatment with phorbol 12-myristate 13-acetate (10(-7) M) for 7 days, there was an increase in the mRNA for CgB and SgII mRNAs in GH and null cell tumors, while dexamethasone treatment for 7 days increased CgA mRNA in GH and null cell adenomas. GnRH treatment for 7 days increased CgB mRNA in null cell adenomas. Phorbol 12-myristate 13-acetate also decreased the percentage of immunoreactive GH cells and GHm RNA, determined by in situ and Northern hybridization analyses. These results indicate that pituitary adenomas have a distinct pattern of Cg/Sg mRNA expression, which appears to be related to the degree of morphological differentiation of these neoplasms, and suggest that the effects of secretagogues on various Cg/Sg mRNA levels may be related to the stimulation of hormone secretion.

We have utilized the unicellular cyanobacterium Synechocystis sp. PCC 6803 to incorporate site-directed amino acid substitutions into the photosystem I (PSI) reactioncenter protein PsaB. A cysteine residue (position 565 of PsaB) proposed to serve as a ligand to the [4Fe-4S] center Fx was changed to serine, histidine, and aspartate. These three mutants--C565S, C565H, and C565D--all exhibited greatly reduced accumulation of PSI reaction-center proteins and failed to grow autotrophically, indicating that this cysteine most likely does coordinate Fx, which is crucial for PSI biogenesis. Interestingly, the strain C565S accumulated significantly more PSI than the other two cysteine mutants and displayed photoreduction of the [4Fe-4S] terminal electron acceptors FA and FB. Mutations were also introduced into a leucine zipper motif of PsaB, proposed to participate in reaction-center dimerization. The mutants L522V, L536M, and L522V/L536M all exhibited wild-type characteristics and grew autotrophically, whereas the L522P mutation prevented PSI accumulation. These data do not provide support for a major structural role of the leucine zipper in reaction-center dimerization or in assembly of Fx. However, the amino acid substitutions incorporated were conservative and might not have perturbed the leucine zipper.

The leaf cuticular waxes of three Salix species and two Populus species hybrids, selected for their ability to produce high amounts of biomass, were characterized. Samples were extracted in CH(2)Cl(2) three times over the growing season. Low kV SEM was utilized to observe differences in the ultrastructure of leaf surfaces from each clone. Homologous series of wax components were classified into organic groups, and the variation in wax components due to clone, sample time, and their interaction was identified. All Salix species and Populus species hybrids showed differences in total wax load at each sampling period, whereas the pattern of wax deposition over time differed only between the Salix species. A strong positive relationship was identified between the entire homologous series of alcohols and total wax load in all clones. Similarly strong relationships were observed between fatty acids and total wax load as well as fatty acids and alcohols in two Salix species and one Populus species hybrid. One Salix species, S. dasyclados, also displayed a strong positive relationship between alcohols and alkanes. These data indicate that species grown under the same environmental conditions produce measurably different cuticular waxes and that regulation of wax production appears to be different in each species. The important roles cuticular waxes play in drought tolerance, pest, and pathogen resistance, as well as the ease of wax extraction and analysis, strongly suggest that the characteristics of the cuticular wax may prove to be useful selectable traits in a breeding program.

Cotton (Gossypium hirsutum L.) fibers are single-celled trichomes that synchronously undergo a phase of rapid cell expansion, then a phase including secondary cell wall deposition, and finally maturation. To determine if there is coordinated regulation of gene expression during fiber expansion, we analyzed the expression of components involved in turgor regulation and a cytoskeletal protein by measuring levels of mRNA and protein accumulation and enzyme activity. Fragments of the genes for the plasma membrane proton-translocating ATPase, vacuole-ATPase, proton-translocating pyrophosphatase (PPase), phosphoenolpyruvate carboxylase, major intrinsic protein, and α-tubulin were amplified by polymerase chain reaction and used as probes in ribonuclease protection assays of RNA from a fiber developmental series, revealing two discrete patterns of mRNA accumulation. Transcripts of all but the PPase accumulated to highest levels during the period of peak expansion (+12–15 d postanthesis [dpa]), then declined with the onset of secondary cell wall synthesis. The PPase was constitutively expressed through fiber development. Activity of the two proton-translocating-ATPases peaked at +15 dpa, whereas PPase activity peaked at +20 dpa, suggesting that all are involved in the process of cell expansion but with varying roles. Patterns of protein accumulation and enzyme activity for some of the proteins examined suggest posttranslational regulation through fiber development.

Chromogranin-A-positive pituitary adenomas include glycoprotein hormone-producing adenomas, null cell adenomas, and a few other pituitary adenomas. We studied the effects of GnRH, CRF, dexamethasone, and phorbol 12-myristate 13-acetate on FSH and LH secretion and on FSH beta and chromogranin-A and -B mRNA expression in 10 chromogranin-A-positive adenomas in vitro to analyze the regulation of FSH and chromogranin-A and -B expression in these neoplasms. Most adenomas responded to GnRH stimulation during 7 days in culture with a 2- to 10-fold increase in FSH and LH secretion and a 2- to 7-fold increase in FSH beta mRNA compared to control values. CRF and phorbol 12-myristate 13-acetate also stimulated FSH and LH secretion 2- to 5-fold in five of seven and three of three cases, respectively, during 7 days in culture. Dexamethasone stimulated both FSH and LH secretion in two of three cases as well as FSH beta mRNA in vitro in the one case examined. GnRH treatment consistently produced a 2-fold increase in chromogranin-B mRNA, but not in chromogranin-A mRNA, after 7 days of culture. These results indicate that many chromogranin-A-positive adenomas respond to GnRH and CRF in vitro by increased hormone secretion and that GnRH stimulation leads to increased amounts of FSH beta and chromogranin-B mRNAs. The differential response of chromogranin-A and -B mRNAs after GnRH stimulation indicates that the chromogranin genes are highly regulated in these tumors.

Strains of Clostridium acetobutylicum were tested for the presence of β-galactosidase and phospho-β-galactosidase activities when grown on lactose. All strains, except C. acetobutylicum ATCC 824, showed both enzyme activities. Only phospho-β-galactosidase activity was detected with C. acetobutylicum ATCC 824. C. acetobutylicum strains P262 and ATCC 824 showed no detectable β-galactosidase or phospho-β-galactosidase activities when grown on glucose. In the fermentation of whey permeate C. acetobutylicum P262 showed an early induction of phospho-β-galactosidase associated with the acidogenic phase. The β-galactosidase activity peaked at a later stage of the fermentation (22 h) coinciding with the solvent production phase. Similar induction of phospho-β-galactosidase at the early stages (13 h) of fermentation of whey permeate by C. acetobutylicum ATCC 824 was also shown. No β-galactosidase activity was detected during the entire course of fermentation by strain ATCC 824.

Non-steady state voltammetry has been applied to membrane electrodes to provide improved sensitivity as well as independence of stirring. This paper describes a new instrumentation system based on complementary metal oxide—silicon technology for the characterization of a pulsed oxygen electrode. The system provides the necessary timing, analog-to-digital conversion, and digital display of the transient current.

With continuous cultures in a fully defined minimal salts medium steady states were achieved at both limiting and non-limiting concentrations of phosphate in the inflowing medium for Rhizobium trifolii WU95, cowpea Rhizobium NGR234, and Bradyrhizobium CB756.

Sterol hydroperoxides 3 beta-hydroxy-5 alpha-cholest-6-ene-5-hydroperoxide and 3 beta-hydroxycholest-5-ene-7 alpha-hydroperoxide show weak dose-response direct mutagenicity towards Salmonella typhimurium strain TA 1537 in a liquid medium incubation bioassay. Responses were compromised by metabolism of the sterol hydroperoxides and by phase separation during the incubation period. Mutagenicity responses were increased by added superoxide dismutase but diminished by added rat liver S9 enzymes and abolished by added catalase. Catalase also abolished the stimulatory effect of superoxide dismutase. These results indicate that superoxide and peroxide be implicated in the mutagenicity responses.

SUMMARY: The concentrations of glycolytic intermediates and ATP and the activities of certain glycolytic and gluconeogenic enzymes were determined in Propionibacterium shermanii cultures grown on a fully defined medium with glucose, glycerol or lactate as energy source. On all three energy sources, enzyme activities were similar and pyruvate kinase was considerably more active than the gluconeogenic enzyme pyruvate, orthophosphate dikinase, indicating the need for regulation of pyruvate kinase activity. The intracellular concentration of glucose 6-phosphate, a specific activator of pyruvate kinase in this organism, changed markedly according to both the nature and the concentration of the growth substrate: the concentration (7-10 mM) during growth with excess glucose or glycerol was higher than that (1-2 mM) during growth with lactate or at growth-limiting concentrations of glycerol or glucose. Other glycolytic intermediates, apart from pyruvate, were present at concentrations below 2 mM. Glucose 6-phosphate overcame inhibition of pyruvate kinase activity by ATP and inorganic phosphate. With 1 mM-ATP and more than 10 mM inorganic phosphate, a change in glucose 6-phosphate concentration from 1-2 mM to 7-10 mM was sufficient to switch pyruvate kinase from a strongly inhibited to a fully active state. The results provide a plausible mechanism for the regulation of glycolysis and gluconeogenesis in P. shermanii.

Lysozyme/EDTA treatment of four fast-growing rhizobia released repeatable protein profiles after polyacrylamide slab gel electrophoresis. Similar treatment of slow-growing rhizobia failed to release such periplasmic proteins. For the four-fast-growing rhizobia, both P-repressible and P-inducible protein bands occurred. The only P-repressible protein identified was alkaline phosphatase, which showed strain differences in both electrophoretic mobility and activation by Mg2+. The derepression of the P-repressible periplasmic proteins in cowpea Rhizobium NGR234 correlated with derepression of both phosphate and glycerol 1-phosphate uptake.

Pyruvate kinase from Propionibacterium shermanii was shown to be activated by glucose-6-phosphate (G-6-P) at non-saturating phosphoenol pyruvate (PEP) concentrations but other glycolytic and hexose monophosphate pathway intermediates and AMP were without effect. Half-maximal activation was obtained at 1 mM G-6-P. The presence of G-6-P decreased both the PEP0.5V and ADP0.5V values and the slope of the Hill plots for both substrates. The enzyme was strongly inhibited by ATP and inorganic phosphate (Pi) at all PEP concentrations. At non-saturating (0.5 mM) PEP, half-maximal inhibition was obtained at 1.8 mM ATP or 1.4 mM Pi. The inhibition by both Pi and ATP was largely overcome by 4 mM G-6-P. The specific activity of pyruvate kinase was considerably higher in lactate-, glucose- and glycerol-grown cultures than that of the enzyme catalysing the reverse reaction, pyruvate, phosphate dikinase. It is suggested that the activity of pyruvate kinase in vivo is determined by the balance between activators and inhibitors such that it is inhibited during gluconeogenesis while, during glycolysis, the inhibition is relieved by G-6-P.

Three strains of Streptococcus lactis , two of Streptococcus cremoris , and one of Streptococcus thermophilus metabolized oxygen in the presence of added carbohydrate primarily via a closely coupled NADH oxidase/NADH peroxidase system. No buildup of the toxic intermediate H 2 O 2 was detected with the three S. lactis strains. All six strains contained significant superoxide dismutase activity and are clearly aerotolerant. Lactose- or glucose-driven oxygen consumption was biphasic, with a rapid initial rate followed by a slower secondary rate which correlated with factors affecting the in vivo activation of lactate dehydrogenase. The rate of oxygen consumption was rapid under conditions that led to a reduction in lactate dehydrogenase activity (low intracellular fructose 1,6-bisphosphate concentration). These conditions could be achieved with nongrowing cells by adding lactose at a constant but limiting rate. When the rate of lactose fermentation was limited to 5% of its maximum, nongrowing cells of S. lactis strains ML3 and ML8 carried out an essentially homoacetic fermentation under aerobic conditions. These same cells carried out the expected homolactic fermentation when presented with excess lactose under anaerobic conditions. Homoacetic fermentation leads to the generation of more energy, by substrate-level phosphorylation via acetate kinase, than the homolactic fermentation. However, it was not observed in growing cells and was restricted to slow fermentation rates with nongrowing cells.

The differential pulse polarography of phenyl arsenoxide (C6H5AsO) and its use in the indirect determination of chlorine and ozone is described.

Optimum conditions for the use of the bare glassy carbon electrode (GCE) are reported. Linear calibration graphs are obtained in the range 5 × 10 -7 –3.5 × 10 -5 M copper(II). The detection limit for copper(II) is 5.9 × 10 -9 M at pH 4.5 and 3.3 × 10 -8 M at pH 6.5.

3-Butenyllithium was prepared in an ether- and halide-free state by cleavage of di(3-butenyl)mercury with lithium metal. This product was shown to be hexameric in hydrocarbon solvents. A series of studies were carried out using 1 H and 7 Li NMR, infrared, and ultraviolet spectroscopy. The results of these studies indicate a perturbation of the π system of the butenyl moiety by the lithium aggregate, an interaction which also was supported by a calculation of the total energy of the system as a function of the conformation. This effect can be accounted for in terms of an intramolecular interaction. This direct observation of a lithium—olefin interaction in a non-dissociated lithium aggregate supports earlier postulations of a similar interaction based on kinetics for ethylenation reactions in ether. Also discussed are the role of aggregation states in accounting for differences in stereospecific addition in hydrocarbon vs. ether solvents.

A membrane separated chemiluminescence (CL) flow cell was constructed and used for the measurement of chlorine dioxide. The membrane separated the luminol-peroxide reagent from the chlorine dioxide and provided a barrier to possible interferences. CL intensity was measured as a function of analyte and reagent flow rate. Sub part per million measurement of dissolved chlorine dioxide was easily obtained.

The effect of P nutrition on phosphate uptake and alkaline phosphatase activity was studied in chemostat culture for four rhizobial and three bradyrhizobial species. Phosphate-limited cells took up phosphate 10- to 180-fold faster than phosphate-rich cells. The four fast-growing rhizobial strains contained high levels of alkaline phosphatase activity under P-limited conditions compared to the repressed levels found in P-rich cells; alkaline phosphatase activity could not be detected in three slow-growing rhizobial strains, regardless of their P-status. Glycerol 1-phosphate-uptake in the cowpea Rhizobium NGR234 was derepressed over 50-fold under P-limited conditions, and appeared to be co-regulated with phosphate uptake. The phosphate-uptake system appeared similar in all strains with apparent K m values ranging from 1.6 μM to 6.0 μM phosphate and maximum activities from 17.2 to 126 nmol · min-1 · (mg dry weight of cells)-1. Carbonyl cyanide m-chlorophenyl hydrazone strongly inhibited phosphate uptake in all strains and a number of other metabolic inhibitors also decreased phosphate uptake in the cowpea Rhizobium NGR234. The phosphate uptake system in all strains failed to catalyse exchange of 32P label in preloaded cells or efflux of phosphate. The results suggest a single, repressible, unidirectional and energy-dependent system for the transport of phosphate into rhizobia.

The previously demonstrated mutagenicity of naturally air-aged (autoxidized) USP cholesterol in test strains of Salmonella typhimurium has been confirmed. In contrast, autoxidized brassicasterol, 7-dehydrocholesterol, ergosterol, lanosterol, sitosterol, and stigmasterol were non-mutagenic in the same assay, and 38 individual cholesterol derivatives, including many cholesterol autoxidation products, were also non-mutagenic. The mutagenic species from mutagenic cholesterol preparations were shown to be neutral steroids that are very much more polar than the cholesterol autoxidation products so far identified. High-performance liquid chromatography resolved mutagenic cholesterol preparations into several mutagenic and non-mutagenic fractions. The chemical nature of the mutagens is suggested as involving oxidation of the sterol B-ring and of the side-chain.

Appraisal of fluorimetric assay of aryl hydrocarbon (benzo(α)pyrene) hydroxylase in cultured human lymphocytes