The Changes in Fluorescence of Acridine Orange-Stained Hela Cells During Ultraviolet Illumination

HeLa cells were stained with a 1/12,000 concentration of acridine orange at pH 7.2 for 3 min and the fluorescence emission was measured quantitatively for effects of ultraviolet illumination with durations including intervals between 5 and 210 min. The total photometric fluorescence intensity increased for the first 30 min, then decreased with illumination time. The initial maximum fluorescence intensity occurred at 525 nm and shifted progressively to shorter wavelengths. Fluorescence intensity above 580 nm decreased with increasing duration of illumination time while that below 580 nm showed an initial increase in intensity followed by a gradual fading.     

The preparation and biological activity of novel amino acid analogs of butirosin

The aminoglycoside antibiotic butirosin (1) has been chemically modified in its amino acid side-chain. The (S)-()-4-amino-2-hydroxybutyryl side-chain was removed by alkaline hydrolysis of the tetrakis(5,5-dimethyl-3-oxo-1-cyclohexen-1-yl) derivative. The resulting deacylated, trisubstituted derivative 3 was reacylated with a wide variety of mono and polyfunctional amino acids. The cyclohexenyl protecting groups were then removed by chlorine gas and the new analogs isolated chromatographically. Structure-activity relationships were determined with five microorganisms, including Pseudomonas aeruginosa. The specific side-chain structures that exhibited maximum potency against Pseudomonas are identified.Butirosin¹, a new aminoglycoside antibiotic complex produced by mucoid strains of Bacillus circulans NRRL B-3312 and B-3313, is the first example of the aminocyclitol class of compounds to contain an amino acid in its chemical structure. Complete structural assignments for butirosin A and B have been described² and are shown in formula 1. The amino acid, which is connected by an amide linkage to N¹ of the deoxystreptamine moiety, was found to be the unique (S)-()-4-amino-2-hydroxybutyric acid². The butirosins have marked activity against Gram-positive and Gram-negative bacteria, including Pseudomonas aeruginosa, both in vitro and in vivo, as well as low toxicity in mammalian species^3,4. They were therefore likely candidates for structural-modification studies.     

Liquid scintillation counting of radioactive monomeric and polymeric carbohydrates on paper chromatograms

A simple, improved scintillation counting procedure was developed for the assay of radioactive mono- and polysaccharides on paper chromatograms. Segments of chromatograms are placed in scintillation vials and soaked in water to completely elute the carbohydrate before addition of Aquasol, a xylene-based scintillation fluid. The resulting water-Aquasol solution is counted in a liquid scintillation counter. Evaluation of numerous experimental variables revealed optimal conditions for complete elution of mono- and polysaccharides with water before counting in Aquasol.The water elution-Aquasol procedure allows water-soluble substances (¹⁴C- and ³H-labeled) on paper to be assayed with increased accuracy and sensitivity (three- to fivefold improvement in counting efficiency of tritiated samples). The simplicity of the procedure allows entire radiochromatograms to be assayed readily.     

Rapid Method for Measuring Extracellular Water in Yeast Preparations

A rapid procedure for the quantitative determination of extracellular water in bulk bakers' yeast was developed on the basis of the solute dilution principle. A reagent is prepared by synthesizing the diazonium ion of p-aminobenzoic acid and coupling it to peptone. This "azopeptone reagent" permits direct colorimetric measurement, which accounts for the rapidity and simplicity of the test. Potential errors due to osmotic effects are avoided by supplementing the reagent with saline and, more importantly, minimizing the duration of contact between reagent and cells. The new method has acceptable accuracy and precision, and may also be suitable for use with other microorganisms.     

Seasonal movement and spatial distribution of the sheep ked (Diptera: Hippoboscidae) on Wyoming lambs.

When populations of adult sheep ked, Melophagus ovinus (L.), infesting unshorn lambs were monitored at the University of Wyoming Paradise Farm during 1986, we determined the body regions on which keds would be found at various times of the year and their seasonal population trends for optimal sampling. Results suggested that ked populations were consistently greater on the ribs than on any other area of the lamb. No significant differences were detected for ked populations between sides of a lamb. Distinct and similar ked population trends over time occurred only in the rib, thigh, shoulder, hind leg, belly, and hind flank areas of the lambs, suggesting that a significant seasonal migration did not occur. Analyses for seasonal population fluctuations indicated that ked populations increased in the winter and spring, decreased in summer and then increased again in the fall. Thus, sampling for keds in the rib area at shearing, which begins in March in Wyoming and runs through mid-April, would be an opportune time to detect keds. At other times of the year, the rib area should be inspected for presence of sheep ked.     

Activation of adenylate cyclase during swelling of S49 cells in hypotonic medium is not involved in subsequent volume regulation

Experiments in S49 mouse lymphoma cells indicate that adenylate cyclase activity is increased following swelling in hypotonic medium through a mechanism independent of the G-proteins which are involved in hormonal regulation of the enzyme. An intact actin cytoskeleton is apparently required for stimulation of adenylate cyclase by mechanical forces. It was hypothesized that this increase in cAMP may be involved in triggering subsequent volume regulatory events. Manipulation of intracellular CAMP content and protein kinase A activity in S49 cells prior to swelling or during the regulatory volume decrease following swelling provided no evidence of a significant role for CAMP in regulating the extent of initial volume increase or the subsequent regulatory volume decrease. Treatment of S49 cells with 10-200 µM miconazole, previously shown to inhibit adenylate cyclase activity, attenuated the initial volume increase with medium dilution and accelerated the rate of regulatory decrease in a dose-dependent and time-dependent manner. However, incubation with 100 µM miconazole for 20 min, which completely inhibited swelling-induced increases in cAMP content, had no significant effect on either the initial volume expansion or the extent of regulatory volume decrease.