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Increased systemic inflammation and an impaired immune response are features of adult chronic renal failure (CRF). These patients have increased rates of infection, cardiovascular disease, anemia, and malnutrition. We measured inflammatory and immunological markers in a group of children with pre-dialytic CRF. No prior studies have explored these markers even though children with non-dialysed CRF exhibit similar complications to those seen in adults with CRF. Blood was collected from children with mild, moderate, or severe CRF and an age-matched control group. Functional leukocyte subsets were determined using flow cytometry. Circulating levels of interleukin (IL)-1beta, IL-6, IL-8, IL-12, IL-10, and tumor necrosis factor-alpha were measured using a flow cytometric bead assay. Children with severe CRF showed significantly reduced total white cell count and absolute neutrophil and lymphocyte counts. Absolute numbers of CD3+/CD45RO+ memory T cells and CD3+/CD45RO+/CD62L+ memory Th2 cells were significantly reduced in all CRF groups versus controls. Children with severe CRF showed increased CD11b expression on neutrophils and monocytes. Some patients showed increases in pro-inflammatory cytokines that were not related to their level of residual renal function. As CD11b expression mediates leukocyte adhesion to vascular endothelium, upregulation may contribute to the increased endothelial dysfunction observed in children with CRF. L-selectin mediates extravasation of leukocytes into tissue and homing of peripheral blood lymphocytes to lymph nodes. The reduction in L-selectin may inhibit these actions and predispose patients to increased infection later in life. This is the first study to comprehensively investigate leukocyte functional molecules and inflammatory cytokine profiles in children with pre-dialytic CRF and provides new immunological evidence for the clinical manifestations associated with the disease.

The first thermally robust and air stable bent-sandwich chromocene complex with chromium in the +4 oxidation state has been isolated and fully characterized.

In order to test an alternative selectable marker system for the production of transgenic peanut plants (Arachis hypogaea), the bacterial mercuric ion reductase gene, merA, was introduced into embryogenic cultures via microprojectile bombardment. MerA reduces toxic Hg(II) to the volatile and less toxic metallic mercury molecule, Hg(0), and renders its source Gram-negative bacterium mercury resistant. A codon-modified version of the merA gene, MerApe9, was cloned into a plant expression cassette containing the ACT2 promoter from Arabidopsis thaliana and the NOS terminator. The expression cassette also was inserted into a second vector containing the hygromycin resistance gene driven by the UBI3 promoter from potato. Stable transgenic plants were recovered through hygromycin-based selection from somatic embryo tissues bombarded with the plasmid containing both genes. However, no transgenic somatic embryos were recovered from selection on 50-100 micromol/L HgCl2. Expression of merA as mRNA was detected by Northern blot analysis in leaf tissues of transgenic peanut, but not in somatic embryos. Western blot analysis showed the production of the mercuric ion reductase protein in leaf tissues. Differential responses to HgCl2 of embryo-derived explants from segregating R1 seeds of one transgenic line also were observed.

Intact soil-core microcosms were used to compare persistence of Pseudomonas chlororaphis 3732RN-L11 in fallow soil and on wheat roots with field releases at diverse sites. Parallel field and microcosm releases at four sites in 1996 were repeated with addition of one site in 1997. Microcosms were obtained fresh and maintained at 60% soil water holding capacity in a growth chamber at 70% relative humidity, a 12-hour photoperiod, and constant temperature. Persistence of 3732RN-L11 was measured at each site in field plots and microcosms at 7-21 day intervals, and in duplicate microcosms sampled at an independent laboratory. Linear regression slopes of field plot and microcosm persistence were compared for each site, and between identical microcosms sampled at different sites, using log10 transformed plate counts. Microcosm persistence closely matched field plots for wheat roots, but persistence in fallow soil differed significantly in several instances where persistence in field plots was lower than in microcosms. Analysis of weather variations at each site indicated that rainfall events of 30-40 mm caused decreased persistence in fallow soil. Cooler temperatures enhanced persistence in field plots at later time points. Inter-laboratory comparison of regression slopes showed good agreement for data generated at different sites, though in two instances, longer sampling periods at one site caused significant differences between the sites. Soil characteristics were compared and it was found that fertility, namely the carbon to nitrogen ratio, and the presence of expanding clays, were related to persistence. These microcosm protocols produced reliable data at low cost, and were useable for pre-release risk analyses for microorganisms.

The structure and backbone dynamics of a double labelled (15N,13C) monomeric, 23.7 kD phosphoglycerate mutase (PGAM) from Schizosaccharomyces pombe have been investigated in solution using NMR spectroscopy. A set of 3125 NOE-derived distance restraints, 148 restraints representing inferred hydrogen bonds and 149 values of (3)J(HNHalpha) were used in the structure calculation. The mean rmsd from the average structure for all backbone atoms from residues 6-205 in the best 21 calculated structures was 0.59 A. The core of the enzyme includes an open, twisted, six-stranded beta-sheet flanked by four alpha-helices and a short 3(10)-helix. An additional smaller domain contains two short antiparallel beta-strands and a further pair of alpha-helices. The C(alpha) atoms of the S. pombe PGAM may be superimposed on their equivalents in one of the four identical subunits of Saccharomyces cerevisiae PGAM with an rmsd of 1.34 A (0.92 A if only the beta-sheet is considered). Small differences between the two structures are attributable partly to the deletion in the S. pombe sequence of a 25 residue loop involved in stabilising the S. cerevisiae tetramer. Analysis of 15N relaxation parameters indicates that PGAM tumbles isotropically with a rotational correlation time of 8.7 ns and displays a range of dynamic features. Of 178 residues analysed, only 77 could be fitted without invoking terms for fast internal motion or chemical exchange, and out of the remainder, 77 required a chemical exchange term. Significantly, 46 of the slowly exchanging (milli- to microsecond) residues lie in helices, and these account for two-thirds of all analysed helix residues. On the contrary, only one beta-sheet residue required an exchange term. In contrast to other analyses of backbone dynamics reported previously, residues in slow exchange appeared to correlate with architectural features of the enzyme rather than congregating close to ligand binding sites.

The roles of a number of amino acids present at the active site of the monomeric phosphoglycerate mutase from the fission yeast Schizosaccharomyces pombe have been explored by site-directed mutagenesis. The amino acids examined could be divided broadly into those presumed from previous related structural studies to be important in the catalytic process (R14, S62 and E93) and those thought to be important in substrate binding (R94, R120 and R121). Most of these residues have not previously been studied by site-directed mutagenesis. All the mutants except R14 were expressed in an engineered null strain of Saccharomyces cerevisiae (S150-gpm:HIS) in good yield. The R14Q mutant was expressed in good yield in the transformed AH22 strain of S. cerevisiae. The S62A mutant was markedly unstable, preventing purification. The various mutants were purified to homogeneity and characterized in terms of kinetic parameters, CD and fluorescence spectra, stability towards denaturation by guanidinium chloride, and stability of phosphorylated enzyme intermediate. In addition, the binding of substrate (3-phosphoglycerate) to wild-type, E93D and R120,121Q enzymes was measured by isothermal titration calorimetry. The results provide evidence for the proposed roles of each of these amino acids in the catalytic cycle and in substrate binding, and will support the current investigation of the structure and dynamics of the enzyme using multidimensional NMR techniques.

Two 3D experiments, (H)CCH(3)-TOCSY and H(C)CH(3)-TOCSY, are proposed for resonance assignment of methyl-containing amino acid side chains. After the initial proton-carbon INEPT step, during which either carbon or proton chemical shift labeling is achieved (t(1)), the magnetization is spread along the amino acid side chains by a carbon spin lock. The chemical shifts of methyl carbons are labeled (t(2)) during the following constant time interval. Finally the magnetization is transferred, in a reversed INEPT step, to methyl protons for detection (t(3)). The proposed experiments are characterized by high digital resolution in the methyl carbon dimension (t(2max) = 28.6 ms), optimum sensitivity due to the use of proton decoupling during the long constant time interval, and an optional removal of CH(2), or CH(2) and CH, resonances from the F(2)F(3) planes. The building blocks used in these experiments can be implemented in a range of heteronuclear experiments focusing on methyl resonances in proteins. The techniques are illustrated using a (15)N, (13)C-labeled E93D mutant of Schizosacharomyces pombe phosphoglycerate mutase (23.7 kDa)., (Copyright 2000 Academic Press.)

Removal of the C-terminal seven residues from phosphoglycerate mutase from Saccharomyces cerevisiae by limited proteolysis is associated with loss of mutase activity, but no change in phosphatase activity. The presence of the cofactor 2, 3-bisphosphoglycerate, or of the cofactor and substrate 3-phosphoglycerate together, confers protection against proteolysis. The substrate alone offers no protection. Replacement of either or both of the two lysines at the C-terminus by glycines has only limited effects on the kinetic properties of phosphoglycerate mutase, indicating that these residues are unlikely to be involved in crucial electrostatic interactions with the substrate, intermediate or product in the reaction. However, the double-mutant form of the enzyme is more sensitive to proteolysis and is no longer protected against proteolysis by the presence of cofactor. The proteolysed wild-type and two of the mutated forms of the enzyme show a reduced response to 2-phosphoglycollate, which enhances the instability of the phospho form of the native enzyme. The phosphoglycerate mutase from Schizosaccharomyces pombe, which lacks the analogous C-terminal tail, has an inherently lower mutase activity and is also less responsive to stimulation by 2-phosphoglycollate. It is proposed that the C-terminal region of phosphoglycerate mutase helps to maintain the enzyme in its active phosphorylated form and assists in the retention of the bisphosphoglycerate intermediate at the active site. However, its role seems not to be to contribute directly to ligand binding, but rather to exert indirect effects on the transfer of the phospho group between substrate, enzyme, intermediate and product.

Earlier attempts to purify and characterize nonrecombinant pyruvate kinase from Schizosaccharomyces pombe proved difficult due to problems associated with the instability of the protein. The enzyme has been overexpressed in Saccharomyces cerevisiae strain AH22, permitting studies to determine the conditions required to stabilize the enzyme during purification. Recombinant S. pombe pyruvate kinase was purified by a combination of ion-exchange chromatography and gel filtration. The purified enzyme showed sigmoidal kinetics with respect to PEP; in the presence of FBP, the kinetics were restored to Michaelis-Menten behavior. With respect to ADP, the Hill coefficient was not affected by FBP. Determination of the molecular mass of the purified enzyme by ultracentrifugation showed that it behaved as a dimer-tetramer system with a Kd of approximately 1 microM., (Copyright 1998 Academic Press.)

A series of homogeneous Eudragit RS100 matrix microspheres containing molecularly dispersed acylated esterified homologues of salicylic acid, (acetylsalicylic acid, valerylsalicylic acid, or caprylsalicylic acid) were prepared in order to investigate the effect of encapsulation on solid-state orientation of the encapsulated molecule. Electrostatic association of the drug with the charged quaternary residues in the polymer may be responsible for the previously observed stability of acetylsalicylic acid (ASA) in aqueous swollen ASA-loaded Eudragit RS100 microspheres. Evaluation of the 13C nuclear magnetic resonance spectra for evidence of structural association of the incorporated probe molecules indicated that alteration of the microenvironment of the incorporated solutes had occurred. For instance, increasing the aliphatic character of the acyl side chain resulted in an increase in the upfield shift of the acyl bearing aromatic ring carbon, (C2), in the incorporated probe molecule as compared to the unincorporated probe molecule. Similarly, a downfield perturbation in the chemical shift of the free acid bearing aromatic ring carbon, (C1), was also observed. This microenvironment electrostatic shielding in the proximity of the ester carbonyl is attributed to an increase in the association of the probe molecule with the polymer subunits. Thereby, it is postulated that the matrix incorporated probe molecule is essentially shielded from hydrolytic attack until it is liberated into the external aqueous environment.

We aimed to determine the natural history of borderline increases in albuminuria in adolescents with insulin-dependent (Type 1) diabetes mellitus (IDDM) and factors which are associated with progression to persistent microalbuminura. Fifty-five normotensive adolescents with IDDM and intermittent microalbuminura (overnight albumin excretion ratte of 20-200 micrograms min-1 on one of three consecutive timed collections, n = 29) or borderline albuminura (mean overnight albumin excretion rate of 7.2-20 micrograms min-1 on one of three consecutive timed collections, n = 30) were followed prospectively at 3 monthly intervals. The endpoint was persistent microalbuminuria defined as a minimum of three of four consecutive overnight albumin excretion rates of greater than 20 micrograms min-1. One hundred and forty-two adolescents with IDDM and normoalbuminura were also followed prospectively. Fifteen of the 59 patients (25.4%) with intermittent (9/29) or borderline (6/30) albuminura progressed to persistent microalbuminura (progressors) over 28 (15-50) months [median (range)] in comparison with two of the 142 patients with normoalbuminuria at entry (relative risk = 12.6; p = 0.001). Progressors to persistent microalbuminura were pubertal and had higher systolic (p = 0.02) and diastolic (p = 0.02) blood pressure, and HbA1c (p = 0.004) than non-progressors. All patients remained normotensive. Glomerular filtration rate, apolipoproteins, dietary phosphorus, protein and sodium intakes, and prevalence of smoking did not differ between progressors and non-progressors. Total renin was higher in the diabetic patients without a difference between progressors and non-progressors. In conclusion there is a relatively high rate of progression to persistent microalbuminuria in pubertal adolescents with borderline increases in albuminura and duration greater than 3 years. These patients require attention to minimize associated factors of poor metabolic control and higher blood pressure in the development of incipient nephropathy.

Homogeneous Eudragit RS100 matrix microspheres containing molecularly dispersed acetylsalicylic acid (ASA) were prepared in order to investigate the effect of encapsulation on the decomposition rate of a hydrolytically susceptible drug. ASA-loaded microspheres of this non-eroding polymer matrix were analysed at predetermined time points following immersion of the microspheres in temperature controlled buffer systems at pH 1.2 or pH 12.1 at 30, 40 or 50 degrees C. The mass balance of the total amount of solutes (ASA and SA) initially located within the microsphere interior was equal to the sum of the amount of solutes remaining in the microsphere interior and the amount of solutes in the aqueous phase at any time during the course of the study. Each analysis involved the quantitation of four species; the drug and decomposition product, salicylic acid (SA), in both the microspheres phase and the external aqueous phase. A simple model system using first-order rate approximations for the concurrent Fickian diffusion and hydrolysis decomposition of the drug resulted in a multiexponential expression which adequately described the time-course profile of the drug. SA-loaded microspheres were used as a control under similar conditions to determine the magnitude of the contribution of microsphere phase hydrolysis of ASA to the overall rate of drug loss from the microspheres. Results indicated that microspheres phase hydrolysis of ASA was minimal. Even after 900 h of immersion in pH 12.1 buffer some ASA remained within the microsphere. It is postulated that the matrix incorporated drug is essentially shielded from hydrolytic attack until it is liberated into the external aqueous environment. Electrostatic association of the drug with the charged quaternary residues in the polymer along with the limiting availability of water within the microsphere may be responsible for the observed stability of ASA in aqueous swollen ASA-loaded Eudragit microspheres.

p-Aminosalicylic acid, which exists in four ionic forms and whose decarboxylation is well known, was used as a model drug to investigate the effects of surfactants with different charges on its stability. The greatest reduction in the rate of decarboxylation at 50 degrees C occurred when the charge on the micelles was opposite to that of the charge on the drug or when both the drug and the micelle had a neutral charge. Thus, at the highest surfactant concentration, 3 or 5% (w/v), there was a 59% reduction in the rate at pH 2.68 in the presence of the nonionic surfactant polyoxyethylene 24 monocetyl ether, 69% reduction in the rate at pH 4.88 in the presence of the cationic surfactant hexadecyl trimethylammonium bromide, and a 43% reduction at pH 1.01 in the presence of the anionic surfactant sodium cetyl sulfate. The decrease in the rate of decarboxylation was attributed to the partitioning of the drug into the micelles, which provided a phase for improved stabilization. Partition coefficients and rate constants for decarboxylation of the drug inside the micelles were calculated.

The small, monomeric, phosphoglycerate mutase (PGAM) from Schizosaccharomyces pombe has been overexpressed in a strain of Saccharomyces cerevisiae in which the gene encoding PGAM has been deleted, with a yield of purified enzyme of 10-15 mg per litre cell culture. Three mutants in which histidine residues in S. pombe PGAM have been substituted by glutamine have been purified and characterised. Two mutants (H151Q and H196Q) have kinetic and structural properties very similar to wild-type enzyme, consistent with the proposed location of these (non-conserved) histidines on the surface of the enzyme. The third mutant (H163Q) involving a histidine thought to be part of the active site has greatly reduced mutase and phosphatase activities. Mass spectrometry shows that the phosphorylated form of the H163Q is several 100-times more stable towards hydrolysis than the phosphorylated form of wild-type enzyme. The H163Q mutant appears to be structurally quite distinct from wild-type enzyme. 600 MHz 1D proton NMR spectra of good quality have been obtained for wild-type enzyme and the H151Q and H196Q mutants.

A cDNA encoding pyruvate kinase from Schizosaccharomyces pombe has been isolated from a lambda ZAPII library. This cDNA was sequenced and found to contain an open reading frame of 1524 nucleotides, giving a predicted protein subunit M, of 55470. The sequence shows a high degree of identity with other pyruvate kinase sequences, with residues implicated in the binding of substrate and metal ion co-factors conserved. However, there are significant differences in the putative subunit interface and effector binding regions which may account for the unusual quaternary structure and regulatory properties of the S. pombe enzyme.

Electrospray mass spectrometry has been used to study the formation and hydrolysis of the phosphorylated forms of two phosphoglycerate mutases. The half-life of the enzyme from Saccharomyces cerevisiae was 35 min at 20 degrees C in 10 mM ammonium bicarbonate, pH 8.0. Addition of 1 mM 2-phosphoglycollate reduced this value by at least 100-fold. The phosphorylated form of the enzyme from Schizosaccharomyces pombe was much less stable with a half-life of less than 1 min. The results are discussed in terms of the kinetic properties of the enzymes. Mass spectrometry would appear to be a powerful method to study the formation and breakdown of phosphorylated proteins, processes which are of widespread significance in regulatory mechanisms.

Two pure, homogeneous xyloglucan-hydrolyzing enzymes from germinated nasturtium seeds have been used to localize xyloglucans specifically in seed cell walls. The enzymes, a novel endo (1----4)-beta-D-glucanase which shows absolute specificity towards xyloglucans and a beta-D-galactosidase which is capable of removing galactosyl residues from polymeric xyloglucans, were used to stabilize gold sols. The complexes were applied to ultrathin sections of nasturtium (Tropaeolum majus L) and tamarind (Tamarindus indica L) seeds. The gold complexes prepared from the active enzyme proteins retained enzyme activity, and such complexes gave extremely weak section-labelling or no labelling at all. When the enzymes were subjected to heat-deactivation before being used to stabilize the gold sols, gold complexes were obtained which lacked enzyme activity, but which gave strong, specific labelling of xyloglucans in ultrathin sections. The specificity of the labelling was checked by substrate-competition, by pretreatment of sections with the active and heat-denaturated enzymes and by comparing the labelling of xyloglucan-containing storage cells with other cell types in the same section. The labelling was maximal at the pH which was optimal for the active enzyme. We conclude that the enzyme-gold complexes which retain high activity against the substrate to be localized are likely to be unsuitable as cytochemical probes because they may cause in situ substrate modification. In the case of the enzyme complexes described here the specific localization obtained with the gold complexes prepared from heat deactivated enzymes may be attributable to the retention by the heat-treated enzymatically-inactive proteins of substrate recognition.(ABSTRACT TRUNCATED AT 250 WORDS)

A method of microencapsulating theophylline ion-exchange resins with ethylcellulose was developed to produce smooth and uniform coats which were predominantly mononucleated. This was achieved by controlling the amount of ethylcellulose and the particle size, and through the use of a protective colloid, polyisobutylene. The rate of release of theophylline was influenced by the ion-exchange resin crosslinking, the amount of ethylcellulose, and the smoothness of the coat. Mesh size and polyisobutylene did not appear to affect the rate in a regular manner. It was found that the release rate from coated resins with low crosslinking followed a logarithmic plot, indicating membrane-controlled release, whereas coated resins with high crosslinking fitted a t1/2 plot, suggesting particle diffusion control.

The hydrolysis kinetics of hydralazine hydrochloride was studied at pH 1 to 12 at 35 degrees, 50 degrees, and 70 degrees C. The hydrolysis takes place by water attack on the dicationic and the cationic forms of the drug. In addition there is hydroxyl attack on the cationic and the neutral forms of the drug. The drug is not subject to attack by acetate and carbonate buffers, but its decomposition is catalyzed by HPO4 = of the H2PO4-, HPO4 = buffer system. The hydrolysis rate follows first order kinetics under nitrogen, at constant pH, temperature, and buffer concentration. The pH profile indicates that hydralazine has maximum stability near pH 3.5. The drug decomposes to phthalazine and other products. The Arrhenius activation energy was found to be 17.9 kcal/mol at pH 3.5 and the time required for 10% loss (t0.9) was estimated from the Arrhenius parameters to be 1.56 years at 25 degrees C.

Ion-exchange resin beads in the benzoate form were coated by several microencapsulation techniques to alter and improve characteristics, especially the control of drug release, of this type of drug delivery system. The most successful techniques included polymer-polymer interaction, temperature change, and nonsolvent addition. The microencapsulated beads then were studied with respect to the release rate of the organic anion to determine the effects of microencapsulation. The release rate of the organic anion could be controlled over a wide range, depending on the encapsulating material characteristics. Factors affecting the extent and rate of release as result of microencapsulation are discussed.

A retrospective cohort study of mortality was conducted to assess whether the military service of young Australian men in Vietnam has influenced their overall death rates, or those from specific causes. The study was of all national servicemen who were conscripted during the conflict and served in the Army for at least 12 months. Of these, 19 205 served in Vietnam (veterans) while 25 677 served only in Australia (non-veterans). These men were traced from the end of their national service (between 1966 and 1973) until the beginning of 1982. For most causes of death, the observed number of deaths of veterans and of non-veterans was less than expected from Australian population death rates, and for no cause was there a statistically significant excess of deaths compared with that of the Australian population. Similarly, when veterans were compared with non-veterans, there was no statistically significant difference in deaths for all causes combined, for diseases of the circulatory system, for deaths in motor vehicle accidents, for suicide and for all external causes after adjustment for the different subsequent death rates of men who had served in different Army corps. All seven deaths from diseases of the digestive system were of veterans. There was no excess of deaths among veterans from cancer or from atypical causes of death in this group of young men. Three-quarters of deaths of both veterans and non-veterans were from external causes, often involving motor vehicle accidents.

Organic anions with similar properties, but different molecular weights, were bound to anion-exchange resins with different cross-linking. It was found that the capacity of the ion-exchange resin for the anions, the percentage of organic anion released, and the rate of release depends on the crosslinking of the ion-exchange resin, the molecular weight of the anion, and the moisture content of the resin. Self-diffusion coefficients for the release rates were determined.

The triplet state EPR spectra of magnetically aligned whole cells of Rhodopseudomonas viridis and Rhodopseudomonas palustris display a marked dependence on the orientation of the static EPR field with respect to the alignment field direction. This observation implies that the primary donor species on which the triplets are localized are ordered within the membranes. We have developed a theoretical model for the system to enable calculation of the orientation of the magnetic axes of the primary donor species with respect to the membranes in which they reside. The triplet state spectra are generated by an ensemble of partially ordered magnetic systems and a computer simulation of the experimental results. The triplet orientation is very similar for the two organisms studied, where one axis lies predominantly in the plane of the membrane and the other two axes have approximately equal projections onto the normal to the membrane.

Accuracy and consistency of death certification and coding practices in Australia were examined in relation to a defined population of 552 young men who died aged 20 to 40 years. For each death a mock death certificate was prepared by a team of physicians using all available clinical, pathological, and coronial data. These certificates, and their International Classification of Diseases (ICD 8) codes derived by an Australian Bureau of Statistics (ABS) nosologist, were compared with the actual death registration papers and their ICD 8 codes. Data for deaths from malignant neoplasms were also compared with results of histopathological review. The mock death certificates were superior to the true death certificates in both completeness of content and accuracy of the recorded cause of death. The major source of discrepancies was unsatisfactory narrative on the original death certificates, the standard of which varied with Australian state and with cause of death. The cause of death coding by ABS was of high standard, but its coding practices generally identify only death certificates where content is inadequate for specific coding. Where the narrative is sufficiently specific to be coded there is no mechanism for checking the accuracy of the cause of death recorded. Suggestions are made for improving the narrative.

The nonexponential phosphorescence decay of a highly folded form of poly-riboadenylic acid (poly rA) with noncovalently bound dye is explained by a novel application of a well-known theory of electronic excitation transfer based on the Förster mechanism. This theory, originally used to describe singlet-singlet energy transfer from donor molecules to an acceptor in a solution, is here applied to the transfer of triplet excitation from the adenine (in poly rA) to the singlet manifold of either of the bound dyes, ethidium bromide or proflavine. New experimental data are presented that allow straight-forward theoretical interpretation. These data fit the form predicted by the theory, U(t) exp(-Bt1/2), where U(t) is the decay of the poly rA phosphorescence in the absence of dye, for a range of relative concentrations of either dye. The self-consistency of these theoretical fits is demonstrated by the proportionality of B to the square root of the Förster triplet-singlet overlap integrals for transfer from poly rA to each of the dyes, as demanded by the theory. From these self-consistent values of B, the theory enables one to deduce the mean packing density of nucleotides in this folded poly rA, which we estimate to be approximately 1 nm-3. We conclude that some variations of the method described here may be useful for deducing packing densities of nucleotides in other compact nucleic acid structures.

Anion-exchange resin beads in the theophylline form were prepared and coated with paraffin or encapsulated with ethylcellulose by various methods; one of these products was subsequently coated with paraffin in order to achieve a wide range of release rates and patterns of release. The various products were subjected to rate studies, and it was found that the patterns of release and the rates could be controlled by the cross-linking of the resin and the coating procedures used. A medium and almost constant rate of release was obtained by using ethylcellulose microencapsulation, a coating of hard paraffin, or a combination of both. The amount of coating and the total amount of drug released was also determined.

A method is presented for encapsulating high molecular weight biological materials such as viral antigen, concanavalin A, and other proteins with cellulose acetate phthalate. The method is simple, inexpensive, and rapid; the process takes approximately 15 min. Capsules generated by this method are produced as microspheres 1-3 mm in diameter. They are stable for at least 6 h in simulated gastric conditions, but disintegrate rapidly under simulated intestinal conditions. Encapsulation had no effect on the activity of the biological materials. The method has potentially wide application for encapsulation of drugs and other substances.

We have developed a new approach to the theory of linear dichroism in partially ordered systems. The description of the partially ordered ensemble uses a density of states function, D(theta, phi, approximately delta), which gives the probability that the direction of polarization for incident polarized light has spherical angles theta and phi in an axis system fixed with respect to the molecule; approximately delta = (delta 1, delta 2...delta n) is a set of parameters that describes the partial ordering. We derive new formulas for linear dichroism using the density of states function and then apply these formulas to the analysis of linear dichroism in reaction centers and whole cells of photosynthetic bacteria. One advantage of our approach is that the order parameter, approximately delta, provides a more complete description of the distribution function than the traditional order parameters used by other authors. Knowledge of approximately delta gives a good physical description of the partial ordering and allows one to calculate accurate limits for the range of possible orientations of the transition moments.

Absorption spectra of the bacteriochlorophyll a-protein from Prosthecochloris aesturaii were measured at temperatures from 2.9 to 300 K. Fourth and eight derivatives of the spectra were calculated from the digital data. From an analysis of 34 scans taken from 750 to 850 nm at 5 K, and 130 scans taken from 822 to 838 nm, we find evidence for nine peaks, six of which are probably 0--0 excitonic and three probably higher vibronic features. The major peaks are resolved in the derivative spectra to 300 K, and all shift with temperature by less than 1 nm compared to their 5 K positions, except for the 825 nm peak which shifts about 2 nm. The most prominent fourth derivative peak at 300 K shifts from 812.9 nm in the standard buffer solution to 814.1 nm in the cryogenic solution in which our low temperature measurements were made. We conclude that the conformation of the protein at 5 K is essentially the same as at 300 K.

Ion-exchange resin beads were coated with various waxes to improve and control their release. The in vitro release rates of benzoate ions from the coated-resin beads were then investigated using a rotating sieve basket technique. The dramatic differences in release rates observed with the different waxes can be discussed in terms of the wax to resin ration and the solubility characteristics of the waxes. The initial release rates can be expressed in terms of a mathematical expression previously reported for the diffusion of ions in ion-exchange resins, thereby aiding in the elucidation of the effect of the waxes on release.

Seventy-nine patients with chronic bronchitis were randomly allotted to four treatment regimens-placebo throughout the winter months for five years; tetracycline for the first two winters and placebo for the next three; placebo for the first two winters and tetracycline for the next three; and tetracycline for five winters. In addition all groups recevied a five-day course of tetracycline for any acute exacerbation. There was a significant reduction in the number of exacerbations among the more susceptible patients-that is, those who suffered more than one exacerbation each winter. Though the average decline in F.E.V.(1) over the five-year period was less in the treated groups this was not statistically significant. There was no significant difference between the groups in respect of lung volumes, diffusing capacity, and blood gases.

A method for appraising the distribution of diffusing capacity of the lungs (D(L)) in relationship to pulmonary capillary blood flow ([unk]Q(C)) in normal human subjects was derived from measurements of oxygen diffusing capacity (D(LO2)) and carbon monoxide diffusing capacity (D(LCO)) performed during breath holding. This method utilizes the fact that the observed D(LO2) is considerably reduced in value if uneven distribution of D(L) with respect to [unk]Q(C) (uneven D(L)/[unk]Q(C)) is present. In contrast, D(LCO) is barely affected by uneven D(L)/[unk]Q(C), and from its measured value one can calculate the value D(LO2) would have if no uneven D(L)/[unk]Q(C) were present (true D(LO2)). Once observed D(LO2) and true D(LO2) are known, the degree of uneven D(L)/[unk]Q(C) in the lung can be calculated. In five normal, resting, sitting subjects average values for true D(LO2) were 57 ml per (minute x mm Hg), and the directly measured D(LO2) was 33 ml per (minute x mm Hg). These values could be explained if one-half of total [unk]Q(C) were distributed to approximately 15% of total D(L). These measurements did not permit the determination of the alveolar to end capillary O(2) gradient, but calculations demonstrate that an important factor in determining its size may be the pattern of uneven D(L)/[unk]Q(C) present in the lungs. Estimations of the alveolar-end capillary O(2) gradient from measurements of D(LCO) or D(LO2) that do not take into account uneven D(L)/[unk]Q(C) may underestimate its size.