Anhydrolide macrolides. 2. Synthesis and antibacterial activity of 2,3-anhydro-6-O-methyl 11,12-carbazate erythromycin A analogues.

A series of 3-descladinosyl-2,3-anhydro-6-O-methylerythromycin A 11, 12-cyclic carbazate analogues was prepared and evaluated for antibacterial activity. These 2,3-anhydro macrolides were found to be potent antibacterial agents in vitro against macrolide-susceptible organisms including Staphylococcus aureus 6538P, Streptococcus pyogenes EES61, and Streptococcuspneumoniae ATCC6303. These compounds were also very active against some organisms that show macrolide resistance (S. aureus A5177, S. pyogenes PIU2584, and S. pneumoniae 5649). The compounds generally showed poor activity against organisms with constitutive MLS resistance. Selected compounds were evaluated in vivo in mouse protection studies. Although most of the compounds tested in vivo showed poor efficacy, two compounds, 38 and 57, were more active than clarithromycin against S. pneumoniae ATCC6303.

Anhydrolide macrolides. 1. Synthesis and antibacterial activity of 2,3-anhydro-6-O-methyl 11,12-carbamate erythromycin A analogues.

A series of 3-descladinosyl-2,3-anhydro-6-O-methylerythromycin A 11, 12-carbamate analogues have been synthesized and evaluated for antibacterial activity. These compounds were found to be potent antibacterial agents against Gram-positive organisms in vitro, many having MIC values below 1 microg/mL for the macrolide-susceptible Staphylococcus aureus, Streptococcus pyogenes, and Streptococcus pneumoniae, as well as improved activity compared to erythromycin A against the inducibly MLS (macrolide, lincosamide, and streptogramin B)-resistant organisms. Structure-activity studies revealed that arylalkyl carbamates with two and four carbon atoms between the aromatic moiety and carbamate nitrogen have the best in vitro activity. All of the C-10 epi analogues evaluated were found to have substantially less activity than the corresponding natural C-10 isomer. Several analogues demonstrated moderate antibacterial activity against the constitutively resistant S.aureus A-5278, S. pneumoniae5979, and S.pyogenes 930. However, despite potent in vitro activity, these analogues showed only moderate in vivo activity in mouse protection studies.

Calyceal diverticula. Ureteroscopic management.

In selected cases, RIRS management of calyceal diverticula and its related problems has been shown to be more efficacious than ESWL monotherapy and avoids the potential complications and discomfort of percutaneous and laparoscopic procedures. The advent of the Holmium laser energy source, with innovations such as the flexible ureteroscope and the tipless stone basket, have expanded the role of RIRS in the management of calyceal diverticula and associated problems. Presently, RIRS is the initial treatment choice for the management of low to moderate stone burdens that are contained in calyceal diverticula or trapped behind any narrowed intrarenal segment (e.g., infundibular stenosis). If repair of the stenotic segment is not successful, thereby excluding stone treatment, then under the same anesthesia, the patient undergoes a percutaneous antegrade renal surgery. The authors feel that the percutaneous approach as a first choice is more suitable for posteriorly located diverticula with a large stone burden. In selected cases of anteriorly located diverticula with large stone burdens, the laparoscopic approach is more expedient.

Prognostic factors and molecular markers for renal cell carcinoma.

Renal cell carcinoma is the most common cancer in the kidney, affecting nearly 30,000 Americans every year and is associated with over 12,000 deaths annually. If detected early, renal cell carcinomas can be cured surgically. However, once metastatic disease develops the prognosis for long-term survival is poor. Unfortunately, one-third of patients have metastatic disease at the time of diagnosis and approximately 50% of the patients undergoing surgical resection for less advanced disease eventually relapse. This review examines the clinical and molecular prognostic tools currently available or under investigation for kidney cancer.

Decreased glutathione in patients with anorexia nervosa. Risk factor for toxic liver injury?

OBJECTIVE: To investigate glutathione and amino acids related to glutathione metabolism in patients with anorexia nervosa in order to test the hypothesis that these patients exhibit a deficiency of glutathione and therefore might be at an increased risk of developing toxic liver injury. DESIGN: Controlled observatory study and case report. SETTING: University Hospital. SUBJECTS: Subjects included 11 female patients with anorexia nervosa and 12 healthy female controls. INTERVENTIONS: Determination of fasting free and total glutathione, homocysteine, vitamins B(6) and B(12) and folic acid in plasma. RESULTS: A 14-y-old patient with a body mass index of 12.6 kg/m(2) presented with markedly elevated transaminases (ALAT >50 x upper limit of normal), and paracetamol was detected in her blood. Patients with anorexia nervosa exhibited lower circulating concentrations of free cysteine (8.9+/-1.5 vs 12.0+/-1.4 micromol/l) and free and total glutathione (5.0+/-1.3 vs 7.1+/-1.2 and 11.2+/-3.8 vs 16.2+/-5.0 micromol/l, respectively). The plasma concentrations of homocysteine (17.5+/-4.9 vs 12.0+/-3.8 micromol/l) and also of glycine (194+/-37 vs 143+/-41 micromol/l) and glutamine (422+/-51 vs 353+/-51 micromol/l) were significantly higher in patients with anorexia nervosa who were not deficient in folic acid, vitamin B(6) and B(12). CONCLUSIONS: Lower plasma concentrations of glutathione suggest lower intracellular concentrations of the tripeptide. Higher homocysteine, glycine and glutamine concentrations point to a decreased utilization of these amino acids for glutathione synthesis and an impairment of trans-sulfuration. Consequently, the capacity of patients with anorexia nervosa to detoxify electrophilic metabolites and reactive oxygen species via glutathione may be impaired.

A modelized distribution of actomyosin interactions in the vertebrate cardiac muscle.

Preliminary assumption of this model is that interactions between actin and myosin presupposes an exact three-dimensional geometrical correspondence between sites, due to the very short time constants present under physiological conditions. Only small and controlled torsions of the actin filaments are accepted. The model uses geometrical information concerning orientations and dimensions of myosin crossbridges and actin monomeres to modelize the distribution of their inter-actions. An orientation map of actin sites in the cross-section perpendicular to the filament axis is proposed, adapted to the specific filament array of vertebrate muscle. Orientation of myosin crossbridges follows Luther's rules. According to the model, any interaction between actin and myosin implies the superimposition of their respective cross-sectional planes. The axial length of actin monomere is 55 A; the distance between two crossbridges along the myosin filament axis is 143 A. The following properties are derived: 1) The shortening step of the sliding actin filament must be a multiple of 11 A (highest common factor). Taking into account the staggered disposition of the two actin strands and the presence of two heads for each cross-bridge, the most probable value for this shortening step is equal to 99 A. A specific scheme is proposed to describe the shortening process. The behavior of the modelized crossbridge does not need any elastic structure--2) Planes situated at 715 A (lowest common multiple) of actin and myosin coinciding planes are also in coincidence. In a hemi-sarcomere the maximal number of these planes, referred to as simultaneously activable planes, is 10 (20 if both myosin heads are considered). The proportion of interactions authorized by the site orientations is 1/12. In the model, the concept of randomly recruited crossbridges is replaced by a discretized recruitment, based on geometrical properties at an ultrastructural level. The proposed distribution is homogeneous: it can be extended radially in the sarcomere and authorizes the actin filament sliding in the whole physiological range under the control of a dual activation function, reproducing Ca++ temporal and spatial distribution.

A microbiological assay on microtitre plates of thiamine in biological fluids and foods.

A microbiological method for the determination of thiamine in biological fluids and food using 96-well microtitre plates and automatic plate reader, suitable for using in routine clinical diagnosis is described. Thiamine was extracted from samples by acid digestion (acetate buffer, pH 4.5) at 110 degrees C for 20 minutes. Assay recovery and reproducibility were optimally evaluated. Results obtained from this method were compared with those obtained using traditional microbiological and HPLC methods. The values resulting from this method were more or less the same as those of the traditional microbiological method, but much higher than those of HPLC assay. However, the new assay presents many advantages: it reduces the use of serum volume, which is a main benefit for clinical analysis. It lowers also the reagent costs and increases the number of analyses as well as it is easy to perform for routine clinical laboratory.

Simple determination of retinol, alpha-tocopherol and carotenoids (lutein, all-trans-lycopene, alpha- and beta-carotenes) in human plasma by isocratic liquid chromatography.

Retinol, alpha-tocopherol, lutein, all-trans-lycopene, and alpha- and beta-carotenes can be simultaneously determined in human plasma by reversed-phase liquid chromatography with a programmable and variable UV wavelength detector. Plasma (200 microliters) is deproteinized with 200 microliters of ethanol containing retinyl acetate, as internal standard, then extracted with 1.0 ml of n-hexane-2,6-di-tert.-butyl-p-cresol (BHT). The organic layer is removed and evaporated and the residue is dissolved in 100 microliters of a mixture of acetonitrile-tetrahydrofuran-methanol (68:22:7, v/v/v). Aliquots (15 microliters) are injected onto a 250 x 4.6 mm I.D. column of Nucleosil 100-5 C18 with a pre-column of Lichrosorb RP-18, 7 microns, 15 x 3.2 mm I.D. A mobile phase of acetonitrile-tetrahydrofuran-methanol (68:22:7, v/v/v), adjusted to 100 (v/v) with 1% ammonium acetate, at a flow-rate of 1.5 ml/min is used. Usual run time is 15 min. Retinol and retinyl acetate are monitored at 325 nm, tocopherol at 290 nm, lycopene at 470 nm, lutein and alpha- and beta-carotenes at 450 nm. The intra-batch coefficients of variation (C.V.) were 2.5, 2.2, 2.4, 5.8, 5.1 and 4.7% for retinol, alpha-tocopherol, lutein, trans-lycopene, alpha- and beta-carotenes, respectively. The inter-batch C.V.s of experiments performed on 30 different days over 12 weeks were 5.7, 3.9, 4.5, 10.9, 11.3 and 10.5%, respectively.

Sample preparation and liquid chromatographic determination of vitamin D in food products.

Vitamin D in different fortified foods is determined by using liquid chromatography (LC). Sample preparation is described for fortified skim milk, infant formulas, chocolate drink powder, and diet food. The procedure involves 2 main steps: saponification of the sample followed by extraction, and quantitation by LC analysis. Depending on the sample matrix, additional steps are necessary, i.e., enzymatic digestion for hydrolyzing the starch in the sample and cartridge purification before LC injection. An isocratic system consisting of 0.5% water in methanol (v/v) on two 5 microns ODS Hypersil, 12 X 0.4 cm id columns is used. Recovery of vitamin D added to unfortified skim milk is 98%. The results of vitamin D determination in homogenized skim milk, fortified milk powder, fortified milk powder with soybean, chocolate drink powder, and sports diet food are given.

Dietary vitamin C intake and concentrations in the body fluids and cells of male smokers and nonsmokers.

Inhaled cigarette smoke releases a variety of oxidizing agents. Ascorbic acid is recognized as an important biological antioxidant. To better characterize the antioxidant protective role of ascorbic acid, a comparison of ascorbic acid concentrations in plasma, leukocytes, bronchoalveolar lavage fluid and alveolar macrophages from a homogeneous group of healthy male smokers (n = 10) and nonsmokers (n = 14) was investigated. The resulting ascorbic acid contents were (means +/- SD) 91 +/- 25 (n = 10) and 87 +/- 25 (n = 14) mumol/L in plasma, 2.09 +/- 0.62 (n = 7) and 2.12 +/- 0.77 (n = 11) mumol/10(9) cells in mononuclear leukocytes, 3.2 +/- 2.2 (n = 10) and 1.7 +/- 1.5 (n = 13) mumol/L in bronchoalveolar lavage fluid and 3.4 +/- 2.3 (n = 8) and 1.6 +/- 1.3 (n = 6) mumol/10(9) cells in alveolar macrophages from smokers and nonsmokers, respectively. Mean daily dietary vitamin C intake was 116 +/- 68 and 107 +/- 59 mg/d for smokers and nonsmokers, respectively. The ascorbic acid contents of bronchoalveolar lavage [3.9 +/- 1.9 mumol/L (n = 8)] and alveolar macrophages [4.1 +/- 2.1 mumol/10(9) cells (n = 6)] of smokers consuming 15 to 20 cigarettes/d were significantly higher (P less than 0.05) than those of nonsmokers. The increased content of ascorbic acid in bronchoalveolar lavage and in alveolar macrophages of smokers compared with nonsmokers may reflect a defensive mechanism against free radical species derived from cigarette smoke.

In situ mitochondrial function in volume overload- and pressure overload-induced cardiac hypertrophy in rats.

OBJECTIVES: Little comparative information is available on mitochondrial function changes during experimentally-induced hypertrophy. Respiratory control mechanisms are not exactly the same in situ and in isolated mitochondria. This study assessed in situ mitochondrial function in two myocardial hypertrophy models. METHODS: Cytochrome aa3 (Cytaa3) and myoglobin (Mb) absorption changes were monitored in isolated rat hearts using dual wavelength spectrophotometry (Cytaa3: 605-630 nm, Mb: 581-592 nm). Hypertrophy was induced by creation of an aortic stenosis or of an aorto-caval fistula. Optical monitoring was performed on diastole-arrested perfused hearts using the sequence O2 perfusion, N2 perfusion during 4 min, and reoxygenation. The plateaus of the Cytaa3 and Mb curves were used to quantify oxidation-reduction and oxygenation levels. Respiratory kinetics were characterized by the slopes of transition phase curves. RESULTS: Myoglobin oxygenation was comparable in the hypertrophied and control hearts. However, Cytaa3 oxidation-reduction levels in the hypertrophied hearts showed a shift towards greater reduction in comparison with the controls (controls: 0.580 +/- 0.008 DO605/DO630 nm, n = 34; fistula: 0.530 +/- 0.023, n = 23; stenosis: 0.522 +/- 0.016, n = 20, p < 0.001). The rate of Cytaa3 reduction and the rate of myoglobin deoxygenation were significantly accelerated (p < 0.005) in the volume overload group (0.507 +/- 0.043, n = 23), whereas the respiratory rate in the pressure overload group (0.389 +/- 0.034, n = 20) was comparable to that in the control hearts (0.358 +/- 0.026 delta DO 605 nm/DO630 nm.min-1, n = 34). CONCLUSION: We found mitochondrial function alterations in both volume overload- and pressure overload-induced cardiac hypertrophy, despite adequate cytosol oxygenation. The patterns of these alterations differed: the redox state showed a shift of similar magnitude toward greater reduction in both models, but the respiratory rate was increased in the volume-overloaded hearts and unchanged in the pressure-overloaded hearts. The modification in the oxidation-reduction state suggested that overload hypertrophy may induce changes in the metabolism of the myocardium, which may, in turn, load to persistent modifications in mitochondrial function. The differences between the two models suggest that adaptation to hypertrophy-inducing events exists at the level of the mitochondrion.

Comparative in vitro activity of ABT-773, a novel antibacterial ketolide.

The in vitro activities of ABT-773, erythromycin, clarithromycin, and azithromycin were compared. ABT-773 was the most active compound against macrolide-susceptible Streptococcus pneumoniae, Streptococcus pyogenes, Staphylococcus aureus, Staphylococcus epidermidis, Listeria monocytogenes, and Enterococcus spp. and multidrug-resistant Streptococcus pneumoniae. It also had good activity against gram-negative and atypical respiratory tract pathogens and Helicobacter pylori.