3,4-anhydro-1,2-O-isopropylidene-beta-D-tagatopyranose and 4,5-Anhydro-1,2-O-isopropylidene-beta-D-fructopyranose.

3,4-Anhydro-1,2-O-isopropylidene-beta-D-tagatopyranose (8) and 4,5-anhydro-1,2-O-isopropylidene-beta-D-fructopyranose (10) have been prepared by treatment of 3,5-di-O-acetyl-1,2-O- isopropylidene-4-O-toluene-p-sulfonyl-beta-D-fructopyranose with methanolic sodium methoxide. The structures of 8 and 10 were assigned by 1H and 13C NMR spectroscopy and that of 10 by X-ray crystallography; both exist in half-chair conformations. Compounds 8 and 10 interconvert in aqueous sodium hydroxide, giving a ratio of 1:2 at equilibrium. The monoacetates of 8 and 10 (5-O-acetyl-3,4-anhydro-1,2-O-isopropylidene-beta-D-tagatopyranose and 3-O-acetyl-4,5-anhydro-1,2-O-isopropylidene-beta-D-fructopyranose) undergo stereospecific epoxide ring opening in 80% acetic acid to give mainly the axial monoacetates 5-O-acetyl-1,2-O-isopropylidene-beta-D-fructopyranose and 4-O-acetyl-1,2-O-isopropylidene-beta-D-tagatopyranose, respectively.

Methane output and lactation response in Holstein cattle with monensin or unsaturated fat added to the diet.

We measured effects of continuous vs twice-daily feeding, the addition of unsaturated fat to the diet, and monensin on milk production, milk composition, feed intake, and CO2-methane production in four experiments in a herd of 88 to 109 milking Holsteins. Methane and CO2 production increased with twice-daily feeding, but the CO2:CH4 ratio remained unchanged. Soybean oil did not affect the milkfat percentages, but fatty acid composition was changed. All saturated fatty acids up to and including 16:0 decreased (P < .01), whereas 18:0 and trans 18:1 increased (P < .001). The 18:2 conjugated dienes also increased (P < .01) when the cows were fed soybean oil. Monensin addition to the diet at 24 ppm decreased methane production (P < .01); the CO2:CH4 ratios reached 15, milk production increased (P < .01), and milkfat percentage and total milkfat output decreased (P < .01), as did feed consumption, compared with cows fed diets without monensin (P < .05). Milk fatty acid composition showed evidence of depressed ruminal biohydrogenation: saturated fatty acids (P < .05) decreased and 18:1 increased (P < .001); most of the increase was seen in the trans 18:1 isomer. As with soybean oil feeding, addition of monensin also increased (P < .05) the concentration of conjugated dienes. The monensin feeding trial was repeated 161 d later with 88 cows, of which 67 received monensin in the diet in the first trial and 21 cows were newly freshened and had never received monensin. Methane production again decreased (P < .05), but this time the CO2:CH4 ratio did not change and all other monensin-related effects were absent. The ruminal microflora in the cows that had previously received monensin seemed to have undergone some adaptive changes and no longer responded as before.

Binding and biological activity of C-terminally modified melanocortin peptides: a comparison between their actions at rodent MC1 and MC3 receptors.

Five subtypes of melanocortin receptors have to date been identified, but to date little is known about the different structural requirements for binding and biological activity at these receptors. In this study, the role of C-terminal melanocortin peptide residues in imparting selectivity for the receptor subtypes was examined. C-terminally modified analogues of alpha-MSH and gamma-MSH were synthesized and their interaction with MC1 and MC3 melanocortin receptors was investigated. This study provides further evidence for an important role of proline 12 (numbering with respect to alpha-MSH) for binding and activity at the MC1 receptor. Although the influence of C-terminal amino acids on binding and activity at MC3-R was less marked, some of them were nevertheless observed to be beneficial for the interaction with this receptor subtype.

Genes encoding proteins homologous to halobacterial Gvps N, J, K, F & L are located downstream of gvpC in the cyanobacterium Anabaena flos-aquae.

Only two gas vesicle genes have been previously identified in the cyanobacteria, gvpA and gvpC, both of which encode structural gas vesicle proteins. Analysis of the nucleotide sequence immediately downstream of gvpC in the cyanobacterium Anabaena flos-aquae has revealed the presence of 4 ORFs (open reading frames) the products of which share significant homology with a number of the gene products derived from halobacterial gvp gene clusters. In halobacteria the gas vesicle gene clusters consist of 14 genes involved in gas vesicle synthesis and assembly. The product of Anabaena ORF 1, located immediately downstream of gvpC is homologous to halobacterial GvpNs. For the remaining ORFs the predicted gene products show homology to both GvpJ and GvpA for ORF 2, to GvpK and GvpA for ORF 3, and to both GvpF and GvpL for ORF 4.

A scale for predicting nonphysiological contributions to pain.

BACKGROUND: Waddell and colleagues have identified a series of nonorganic signs which may appear in low back pain patients. The occurrence of these signs helps predict nonphysiological factors in the etiology of the pain. This study describes the development of the Waddell Equivalency Scale (WES) which elicits comparable information from patients with pain at other sites. METHODS: One hundred fifty patients presenting to a comprehensive pain clinic with low back pain were evaluated by physicians for Waddell signs and by psychologists using the Pain Presentation Inventory (PPI) developed by Psychometric Designs Inc. From three scales of the PPI, correlation, analysis of variance and multiple regression identified a formula which predicted Waddell signs from the PPI. The study was replicated on a new sample of 150 patients with almost identical results. The two samples were combined, again with highly similar results. An interpretive scheme was devised for use of the WES scale. RESULTS: The WES score derived from three scales of the PPI predicts the number of Waddell signs elicited from low back pain patients. Since it comes from a paper-and-pencil test rather than a physical examination, it can be used with patients whose pain locus is other than the back. CONCLUSIONS: Behavioral expectations and treatment plans, à la Waddell, can now be applied to patients with other than low back pain. Clinical application of the WES information over a variety of pain populations will be required to determine the parameters of its usefulness.

Methane and carbon dioxide emissions from dairy cows in full lactation monitored over a six-month period.

Methane and CO2 emissions from a herd of 118 lactating cows were measured directly by continuous monitoring with an infrared gas analyzer from 24 gas sampling locations. A total of 112 d of gas output was recorded between June 1993 and November 1993. Recordings were integrated at .5-h intervals, so that there were 48 data points for each 24-h period. The mean 24-h CH4 emission per cow was 587 +/- 61.3 L; the range was 436 to 721 L. The mean 24-h CO2 emission per cow was 6137 +/- 505 L, and the range was 5032 to 7427 L. These values were not corrected for gas emissions from stored manure, which contributed 5.8 and 6.1%, respectively, to CH4 and CO2 output under conditions of this experiment.

GvpCs with reduced numbers of repeating sequence elements bind to and strengthen cyanobacterial gas vesicles.

We have previously shown that the gas-vesicle protein GvpC is present on the outer surface of the gas vesicle, can be reversibly removed and rebound to the surface, and increases the critical collapse pressure of the gas vesicle. The GvpC molecule, which contains five partially conserved repeats of 33 amino acids (33-RR) sandwiched between 18 N-terminal and 10 C-terminal amino acids, is present in a ratio of 1:25 with the GvpA molecule, which forms the ribs of the gas vesicle. By using recombinant techniques we have now made modified versions of GvpC that contain only the first two, three or four of the 33-amino-acid repeats. All of these proteins bind to and strengthen gas vesicles that have been stripped of their native GvpC. Recombinant proteins containing three or four repeats bind in amounts that give the same ratio of 33-RR:GvpA (i.e. 1:5) as the native protein, and they restore much of the strength of the gas vesicle; the protein containing only two repeats binds at a lower ratio (1:7.7), however, and restores less of the strength. Ancestral proteins with only two, three or four of the 33-amino-acid repeats would have been functional in strengthening the gas vesicle but the progressive increase in number of repeats would have provided strength with increased efficiency.

A Pain Rating Scale and a Pain Behavior Checklist for clinical use: development, norms, and the consistency score.

A Pain Rating Scale [PRS] and a Pain Behavior Checklist [PBC] were developed for use during a clinical pain interview. Norms were based on 395 chronic pain patients referred for pain management. The PRS obtained ratings of present pain, and worst and least pain during the previous 30 days. The PBC recorded 16 pain behaviors during the 45- to 60-min interview. Normative data were given for seven measures: (1) worst pain rating, past month; (2) least pain rating, past month; (3) present pain rating; (4) difference between worst and least pain ratings; (5) pain behavior score; (6) total pain score (based on Nos. 1, 2, 3, and 5), and (7) consistency score. The consistency score reflects the agreement (or discrepancy) between the pain behavior and present pain ratings. A moderate overall relationship was found between pain behavior and present pain ratings (r = 0.46, p < 0.001). The frequency of good agreement between pain behavior and present pain ratings was unaffected by sex, race, age, pain site, type of injury, duration of pain, legal representation, and evaluating psychologist; but it varied markedly with conscious symptom magnification. Patients seen as consciously exaggerating pain (n = 127) gave higher pain ratings (all p values < 0.001); and had frequent (64.6%) discrepancies between their pain report and pain behavior compared to others (14.2%). The report of present pain was unrelated to pain behavior for conscious exaggerators (r = 0.04, NS); in contrast there was a moderately high relationship for other patients (r = 0.68, p < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

As-needed medication use in asthma usage patterns and patient characteristics.

Daily inhaled bronchodilator medication usage was recorded using an electronic device and airway obstruction by daily peak flow measurement. The demographic, clinical, and psychological characteristics of the subjects were noted. Subjects were allocated to as-needed (prn) medication usage groups according to the mean number of inhaler activations on days with zero, moderate, and severe airway obstruction. Segregation into arbitrary and nonarbitrary use, and into overuse, appropriate use, and underuse resulted in six usage groups. Appropriate use was observed in only 10 of 39 subjects. The major psychological variable to differ among groups was the MMPI variable Pt, representing general anxiety. Arbitrary users had a significantly higher mean score than nonarbitrary users. The variable Specific Internal Awareness, representing a perceived difficulty in recognizing the premonitory symptoms of an asthma attack, also differed among the usage groups, with arbitrary users having the lowest scores. These findings raise the possibility that reliance on an objective measurement of airway obstruction rather than on subjective symptomatology might enhance compliance with prn medication in some patients.

Isolation and partial purification of a melanocyte-stimulating hormone receptor from B16 murine melanoma cells. A novel approach using a cleavable biotinylated photoactivated ligand and streptavidin-coated magnetic beads.

The alpha-melanocyte-stimulating hormone (alpha-MSH) receptor of B16 mouse melanoma cells was characterized by photoaffinity labelling using radiolabelled photoactive derivatives of alpha-MSH. A doublet band of 43-46 kDa representing a ligand-receptor complex was identified. A novel adaptation of the streptovadin/biotin-based affinity system was used to isolate the alpha-MSH receptor. A probe was synthesized which contained biotin connected to a photolabelled alpha-MSH analogue via a cleavable disulphide linker and which displayed high affinity for the alpha-MSH receptor. Streptavidin-coated magnetic beads were used as a solid support instead of an affinity column. Covalently linked probe-receptor complexes solubilized in Triton X-100 were equilibrated with the beads, and after magnetic separation and washing, specifically bound complexes were treated with dithiothreitol to cleave the disulphide bridge in the biotin-peptide spacer arm and so release the receptor-ligand complex. The identity of the isolated protein was established by SDS/PAGE analysis. Methods to achieve purification to homogeneity and to allow quantitative isolation of the receptor are discussed.

Compliance in clinical trials of two nonbronchodilator, antiasthma medications.

An electronic monitoring device was used to assess patient compliance during clinical trials of two new aerosolized nonbronchodilator, antiasthma drugs. Compliance was poor, but similar, in both trials. Patients took the study drug as instructed on a mean of 37.3% days (range 10% to 77%) in one trial and 37.4% days (range 0% to 63%) in the other. Both underuse and overuse were observed. Underuse was seen on a mean of 38.9% days (range 9% to 81%) and 46.0% (range 15% to 80%) respectively; overuse on a mean of 23.5% days (range 6% to 54%) and 16.6% days (range 0% to 41%). In some patients ten or more activations of the device were recorded at the same time. In six patients such multiple simultaneous activations were recorded on two or more separate days. These patients were among the most noncompliant in the study. Multiple simultaneous activations frequently followed underuse days or preceded followup visits, suggesting the possibility of duplicity. Overall compliance was such that valid conclusions about efficacy of the drugs could only have been drawn in six of 34 patients.

Structure-activity studies with fragments and analogous of salmonid melanin-concentrating hormone.

A number of cyclic and linear fragments and analogues of MCH were synthesized and their biological potencies tested using the isolated carp scale melanophore assay. In this system the cyclic portion MCH(5-14) exhibited only 0.1% bioactivity, which was markedly enhanced by the addition of the exocyclic sequences MCH(15-17) and MCH(1-4). The exocyclic sequence itself, MCH(1-4,15-17), had minimal activity, however. Substitution of Tyr11 with phenylalanine reduced the potency of the ring structure MCH(5-14) by about 4-fold. Substitution of Gly8 with D-alanine reduced the potency of MCH(5-14) 16-fold, while both substitutions together caused a still more marked reduction (200-fold) in bioactivity. Linearized fragments of MCH, extending from MCH(15-17) to [Cys(Acm)5,14]MCH(1-17), showed a progressive increase in potency. The linearized forms of MCH, MCH(5-17) and MCH(5-14), were approximately 100-fold or less potent than their cyclic forms. The significant increases in bioactivity produced by the addition of the C- and N-terminal exocyclic sequence even to these linearized forms further emphasizes the importance of these regions for interaction at the receptor site.

Melanin-concentrating hormone: a structural and conformational study based on synthesis, biological activity, high-field NMR, and molecular modeling techniques.

A series of Melanin-concentrating hormone (MCH) fragments have been synthesized and their biological activities compared with the parent peptide. The substructural units, 5-14 linear and 5-14 cyclic, have been used as models for MCH-- H-Asp1-Thr-Met-Arg-Cys-Met-Val-Gly-Arg HO-Val17-Glu-Trp-Cys-Pro-Arg-Tyr-Val in 1H-nmr conformational studies. Conformational features predicted by molecular dynamics analyses find support in the nmr experiments.

Accessible conformations of melanin-concentrating hormone: a molecular dynamics approach.

Molecular dynamics simulations have been used to search for the accessible conformations of the melanin-concentrating hormone (MCH). The studies have been performed on native MCH and two of its peptide fragments, a cyclic MCH(5-14) fragment and a linear MCH(5-14) fragment. An analysis of the molecular dynamics trajectories of the three peptides indicates that two regions of the peptide have characteristic conformational properties that may be important for the biological activity. One is a region around Gly8, which is conformationally mobile, and the other is around Pro13, which shows unusual rigidity. The molecular dynamics simulation results are discussed in terms of backbone structural features like beta turns, side-chain interactions, and orientations of the disulfide bridge. The results of this analysis are used to suggest new analogues that will modify the conformational features of the peptide and further define the conformational requirements for activity. Finally, the results are related to nmr studies of the peptide and reveal agreements between the experimental nuclear Overhauser effect constraints and some of the accessible conformations obtained from the simulation.

Multidimensional analysis of peak pain symptoms and experiences.

Peak pain symptoms and experiences were explored within a group of 243 intractable pain patients seen consecutively at a pain clinic. Using a 5-point scale, patients rated the frequency with which 99 symptom adjectives occurred when their pain was at its worst. Key cluster analysis identified 11 reliable, conceptually clear symptom clusters: Four affective symptom categories, Angry Depression, Diminished Drive, Intropunitive Depression and Anxiety, describing emotional states concomitant with peak pain; two somatic symptom categories, Ecto-Pain and Endo-Pain, describing surface and deep bodily pain, respectively; and five additional symptom categories including Cognitive Dysfunction, Sleep Disturbance, Fatigue, Withdrawal and Disequilibrium. Among the affective symptom clusters, symptoms of Angry Depression were reported to occur frequently by 32% of the patients while only 11% reported the frequent occurrence of Intropunitive Depression. For the somatic symptom clusters, 25 and 52% reported the frequent occurrence of Ecto-Pain and Endo-Pain, respectively. Pain reports measured by Ecto-Pain and Endo-Pain were nearly independent of all other symptom categories. The results suggest that the experiential context of pain differs widely among intractable pain patients. The study derived a Pain Symptom Checklist to measure each symptom cluster as one way to identify coping styles among chronic pain patients.