Liquid chromatography-electrospray tandem mass spectrometry method for determination of indapamide in serum for single/multiple dose bioequivalence studies of sustained release formulations

Indapamide and internal standard (5-chloro-2-methoxy-N-[2-(4-sulphamoylphenyl)ethyl]benzamide) were isolated from plasma by a single step liquid-liquid extraction in t-butyl methyl ether. The chromatographic separation was achieved on a reversed-phase C(18) monolithic column with a mobile phase consisting in a methanol/aqueous 0.1% formic acid mixture and a flow rate of 0.8 ml/min, in isocratic conditions, within 11 min. Target compounds were transferred in an ion trap analyzer via an atmospheric pressure electrospray interface (AP-ESI). The mass analyzer was used in a selected reaction monitoring (SRM) mode, in order to enhance on detection selectivity. Whole method produces quantitation limit for indapamide of 1 ng/ml. Method was successfully applied to assess bioequivalence of two sustained release marketed pharmaceutical formulations of indapamide 1.5 mg coated tablets, carried-out in a single/multiple doses, randomized design.     

Total body skeletal muscle and adipose tissue volumes: estimation from a single abdominal cross-sectional image.

A single abdominal cross-sectional computerized axial tomography and magnetic resonance image is often obtained in studies examining adipose tissue (AT) distribution. An abdominal image might also provide additional useful information on total body skeletal muscle (SM) and AT volumes with related physiological insights. We therefore investigated the relationships between abdominal SM and AT areas from single images and total body component volumes in a large and diverse sample of healthy adult subjects. Total body SM and AT volumes were derived by whole body multislice magnetic resonance imaging in 123 men [age (mean +/- SD) of 41.6 +/- 15.8 yr; body mass index of 25.9 +/- 3.4 kg/m(2)] and 205 women (age of 47.8 +/- 18.7 yr; body mass index of 26.7 +/- 5.6 kg/m(2)). Single abdominal SM and AT slice areas were highly correlated with total body SM (r = 0.71-0.92; r = 0.90 at L(4)-L(5) intervertebral space) and AT (r = 0.84-0.96; r = 0.94 at L(4)-L(5) intervertebral space) volumes, respectively. R(2) increased by only 5.7-6.1% for SM and 2.7-4.4% for AT with the inclusion of subject sex, age, ethnicity, scanning position, body mass index, and waist circumference in the model. The developed SM and AT models were validated in an additional 49 subjects. To achieve equivalent power to a study measuring total body SM or AT volumes, a study using a single abdominal image would require 17-24% more subjects for SM and 6-12% more subjects for AT. Measurement of a single abdominal image can thus provide estimates of total body SM and AT for group studies of healthy adults.     

Estimating whole body intermuscular adipose tissue from single cross-sectional magnetic resonance images.

Intermuscular adipose tissue (IMAT), a novel fat depot linked with metabolic abnormalities, has been measured by whole body MRI. The cross-sectional slice location with the strongest relation to total body IMAT volume has not been established. The aim was to determine the predictive value of each slice location and which slice locations provide the best estimates of whole body IMAT. MRI quantified total adipose tissue of which IMAT, defined as adipose tissue visible within the boundary of the muscle fascia, is a subcomponent. Single-slice IMAT areas were calculated for the calf, thigh, buttock, waist, shoulders, upper arm, and forearm locations in a sample of healthy adult women, African-American [n = 39; body mass index (BMI) 28.5 +/- 5.4 kg/m2; 41.8 +/- 14.8 yr], Asian (n = 21; BMI 21.6 +/- 3.2 kg/m2; 40.9 +/- 16.3 yr), and Caucasian (n = 43; BMI 25.6 +/- 5.3 kg/m2; 43.2 +/- 15.3 yr), and Caucasian men (n = 39; BMI 27.1 +/- 3.8 kg/m2; 45.2 +/- 14.6 yr) and used to estimate total IMAT groups using multiple-regression equations. Midthigh was the best, or near best, single predictor in all groups with adjusted R2 ranging from 0.49 to 0.84. Adding a second and third slice further increased R2 and reduced the error of the estimate. Menopausal status and degree of obesity did not affect the location of the best single slice. The contributions of other slice locations varied by sex and race, but additional slices improved predictions. For group studies, it may be more cost-effective to estimate IMAT based on one or more slices than to acquire and segment for each subject the numerous images necessary to quantify whole body IMAT.     

Expression of lauroyl-acyl carrier protein thioesterase in brassica napus seeds induces pathways for both fatty acid oxidation and biosynthesis and implies a set point for triacylglycerol accumulation

Expression of a California bay lauroyl-acyl carrier protein thioesterase (MCTE) in developing seeds of transgenic oilseed rape alters the fatty acid composition of the mature seed, resulting in up to 60 mol% of laurate in triacylglycerols. In this study, we examined the metabolism of lauric acid and 14C-acetate in developing seeds of oilseed rape that express high levels of MCTE. Lauroyl-CoA oxidase activity but not palmitoyl-CoA oxidase activity was increased several-fold in developing seeds expressing MCTE. In addition, isocitrate lyase and malate synthase activities were six- and 30-fold higher, respectively, in high-laurate developing seeds. Control seeds incorporated 14C-acetate almost entirely into fatty acids, whereas in seeds expressing MCTE, only 50% of the label was recovered in lipids and the remainder was in a range of water-soluble components, including sucrose and malate. Together, these results indicate that the pathways for beta-oxidation and the glyoxylate cycle have been induced in seeds expressing high levels of MCTE. Although a substantial portion of the fatty acid produced in these seeds is recycled to acetyl-CoA and sucrose through the beta-oxidation and glyoxylate cycle pathways, total seed oil is not reduced. How is oil content maintained if lauric acid is inefficiently converted to triacylglycerol? The levels of acyl carrier protein and several enzymes of fatty acid synthesis were increased two- to threefold at midstage development in high-laurate seeds. These results indicate that a coordinate induction of the fatty acid synthesis pathway occurs, presumably to compensate for the lauric acid lost through beta-oxidation or for a shortage of long-chain fatty acids.     

Hybrid density functional theory with a specific reaction parameter: hydrogen abstraction reaction of difluoromethane by the hydroxyl radical

Accurate potential energy surfaces for the OH + CH2F2 --> H2O + CHF2 reaction are constructed using hybrid and hybrid meta density functional theory methods (mPW1PW91, B1B95, and mPW1B95) with specific reaction parameters in conjunction with the 6-31 + G(d,p) basis set. The accuracy of a surface is examined by comparing the calculated rate constants with the experimental ones. The rate constants are calculated over the temperature range 200-1,500 K using variational transition state theory with multidimensional tunneling contributions. The hybrid density functional theory methods with specific-reaction-parameter Hartree-Fock exchange contributions (39.2-41.0% for mPW1PW91, 41.0-42.2% for B1B95, and 44.9-46.3% for mPW1B95, respectively) provide accurate rate constants over an extended temperature range. The classical barrier height for the hydrogen abstraction reaction on these potential energy surfaces is determined to be 5.0-5.3 kcal mol(-1), and the best estimate value is 5.14 kcal mol(-1).     

Achiral-chiral LC/LC-FLD coupling for determination of carvedilol in plasma samples for bioequivalence purposes

Bioequivalence data for two pharmaceutical formulations (solid oral dosage forms) containing carvedilol is presented for both racemic and enantiomers of the active substance. This was achieved by on-line coupling of two liquid chromatographic separations followed by fluorescence detection. The first LC dimension was used for a fast separation of racemic carvedilol from propranolol (IS) and the endogenous matrix, by means of a reversed phase mechanism. The peak of racemic carvedilol was on-line transferred to the second enantioselective LC dimension, based on a reversed phase separation on cellulose tris(3,5-dimethyl-phenylcarbamate) stationary phase. Both stages were monitored over a single run by means of a fluorescence detector operated at an excitation wavelength of 285 nm and an emission wavelength of 355 nm. Automated shortcutting of the racemic carvedilol peak to the chiral column and simultaneous detection over the two LC dimensions have been obtained by using an experimental set-up based on two six-port rotative switching valves. Linearity was demonstrated on the interval 2-150 ng/mL for racemic carvedilol and on 1-75 ng/mL intervals for enantiomers. LLOQ fits between 0.7 and 1.4 ng/mL. Recoveries of the target compounds are 87+/-4 and 81+/-4% for the IS. Precision ranged from 0.6 to 2.5% and the mean accuracy obtained on quality control samples (measured as % bias) over the whole study falls between -0.8 and 6.3%.     

Persistence of native and exotic plants 10 years after prairie reconstruction

Prairie reconstructions are a critical component of preservation of the imperiled tallgrass prairie ecosystem in the Midwestern United States. Sustainability of this endeavor depends on establishment of persistent cover of planted native species and resistance to noxious weeds. The goal of this study was to understand the influence of early reconstruction practices on long‐term outcomes. Twelve replicates of three planting methods (dormant‐season broadcast, growing‐season broadcast, and growing‐season drill) and three seed mix richness levels (10, 20, or 34 species), fully crossed in a completely randomized design were planted in 2005 on nine former agricultural fields located in Iowa and Minnesota. Cover by species was estimated in 2005–2007, 2010, and 2015. In 2015, cover of planted species, native nonplanted species, and exotic species were similar to those recorded in 2010. Cover of the noxious weed Cirsium arvense had also declined by an average of 49% without herbicide from a peak in 2007 to low stable levels from 2010 to 2015. Richness of planted forbs, on the other hand, were still increasing in high‐richness broadcast treatments (e.g. 17–59% increase 2010–1015 in Minnesota). Two results in 2015 are reasons for concern: cover of planted species is only slightly over 50% in both Minnesota and Iowa, though with forbs still increasing, this may improve; and the cool‐season exotic grasses Poa pratensis and Bromus inermis are increasing at both Minnesota and Iowa sites. Control of these invasive grasses will be necessary, but care will be needed to avoid negative impacts of control methods on natives.     

Driving with Central Visual Field Loss II: How Scotomas above or below the Preferred Retinal Locus (PRL) Affect Hazard Detection in a Driving Simulator

We determined whether binocular central scotomas above or below the preferred retinal locus affect detection of hazards (pedestrians) approaching from the side. Seven participants with central field loss (CFL), and seven age-and sex-matched controls with normal vision (NV), each completed two sessions of 5 test drives (each approximately 10 minutes long) in a driving simulator. Participants pressed the horn when detecting pedestrians that appeared at one of four eccentricities (-14°, -4°, left, 4°, or 14°, right, relative to car heading). Pedestrians walked or ran towards the travel lane on a collision course with the participant’s vehicle, thus remaining in the same area of the visual field, assuming participant''s steady forward gaze down the travel lane. Detection rates were nearly 100% for all participants. CFL participant reaction times were longer (median 2.27s, 95% CI 2.13 to 2.47) than NVs (median 1.17s, 95%CI 1.10 to 2.13; difference p<0.01), and CFL participants would have been unable to stop for 21% of pedestrians, compared with 3% for NV, p<0.001. Although the scotomas were not expected to obscure pedestrian hazards, gaze tracking revealed that scotomas did sometimes interfere with detection; late reactions usually occurred when pedestrians were entirely or partially obscured by the scotoma (time obscured correlated with reaction times, r = 0.57, p<0.001). We previously showed that scotomas lateral to the preferred retinal locus delay reaction times to a greater extent; however, taken together, the results of our studies suggest that any binocular CFL might negatively impact timely hazard detection while driving and should be a consideration when evaluating vision for driving.