Extracellular volume estimation from ratios of bromide to chloride in urine or saliva.

Use of either urine or saliva samples to estimate extracellular water volume was investigated in 10 men using nonradioactive bromide (Br) and in seven newborn piglets using radioactive Br (82Br) and chloride (36Cl). The relation to Br to Cl concentrations in urine enabled an estimation of Br dilution volume from human urine (267 +/- 42 ml/kg, mean +/- SD) that was not significantly different (P = 1.0) from the Br dilution volume calculated from plasma Br concentration (268 +/- 20 ml/kg). Although the Br dilution volume estimated from saliva was not different from that of plasma, the error in the estimates of Br dilution volume from saliva was too large (mean difference, -36 +/- 64 ml/kg) to make its use practical. The data from piglets showed good agreement between 82Br and 36Cl dilution volumes calculated from 4-hr plasma samples (356 +/- 14 ml/kg and 347 +2- 12 ml/kg; P greater than 0.1) and between 82Br dilution volumes calculated from urine 82Br:36Cl and plasma 82Br (360 +/- 31 ml/kg and 356 +/- 14 ml/kg; P greater than 0.1). Extracellular water volume can be estimated in both adult and young animals using the Br dilution volume calculated from urine samples. It requires (i) two urine collections: one before and one 4 to 8 hr after administration of Br; (ii) a measurement or estimate of plasma Cl concentration; and (iii) a correction factor that describes the relationship of the ratio of Br to Cl in urine to that ratio in plasma.     

Atrial natriuretic peptide during water deprivation or hemorrhage in rats. Relationship with arginine vasopressin and osmolarity.

The concentrations of atrial natriuretic peptide (ANP) in atria, hypothalami and plasma were investigated in relation to the variations of the plasma endogenous immunoreactive arginine vasopressin (Ir-AVP) during water deprivation or hemorrhage in normal conscious Wistar rats. Furthermore, the in vitro and in vivo effect of extracellular hyperosmolarity on ANP release from right atrium and hypothalamus was examined. Water deprivation elevated circulating immunoreactive ANP (Ir-ANP: pg/ml) to 153 +/- 7 (24 h); 174 +/- 1 (48 h) from the control level (109.6 +/- 7.8). This increase in Ir-ANP concentration which correlated with atrial (r = -0.93) or hypothalamic (r = -0.87) Ir-ANP content decrease, was associated with significantly enhanced levels of plasma Ir-AVP, plasma sodium, osmolarity and hematocrit. An acute volume depletion by hemorrhage significantly reduced plasma Ir-ANP (67 +/- 8.4 pg/ml) from the sham operated level (140 +/- 18 pg/ml). Plasma Ir-AVP was elevated dramatically (207.4 +/- 53.4 pg/ml) compared with the sham operated level (8.8 +/- 2.6 pg/ml). These results, indicating the lack of correlation between plasma Ir-ANP and Ir-AVP in vivo, suggest that the ANP secretion, which is regulated mainly by plasma volume, may be modulated by a change in plasma osmolarity. Extracellular hyperosmolarity stimulated the ANP release from superfused sliced normal rat atria and hypothalami.     

Comparative plasma pharmacokinetics of meloxicam in sheep and goats following intravenous administration

Meloxicam, a novel cyclooxygenase-2 selective nonsteroidal anti-inflammatory drug (NSAID), has been used extensively in humans and recently in some domestic animal species. Although it is an attractive NSAID for use in small ruminants, meloxicam pharmacokinetics have not been investigated in sheep and goats and this information is essential for rational therapeutic use of the drug in these species. In this investigation, comparative pharmacokinetic properties of meloxicam were studied in sheep and goats after a single intravenous dose of 0.5 mg kg(-1) body mass. Blood samples were collected via jugular venepuncture into heparinised tubes at predetermined times after drug administration. Plasma concentrations of meloxicam were determined by reversed-phase high performance liquid chromatography. The plasma concentrations of meloxicam were detectable in sheep and goats up to 72 and 48 h, respectively. The plasma concentration versus time data of meloxicam in both sheep and goats were adequately described by a two-compartment open model. The values obtained for sheep and goats for distribution half-life, volume of distribution at steady state and volume of the central compartment were almost similar in sheep and goats. The elimination half-life (t(1/2beta)), area under the plasma concentration-time curve (AUC), mean residence time (MRT) and total systemic clearance (Cl(B)) in sheep were significantly different from those of goats. The mean+/-S.E. values of t(1/2beta), MRT, AUC and Cl(B) in sheep were 10.85+/-1.21 h, 15.13+/-1.67 h, 31.88+/-2.97 microg h mL(-1) and 0.016+/-0.002 L h(-1) kg(-1), respectively whereas the respective values in goats were 6.73+/-0.58 h, 9.37+/-0.83 h, 19.23+/-2.23 microg h mL(-1) and 0.03+/-0.01 L h(-1) kg(-1). The results indicate that elimination kinetics of meloxicam differ significantly between sheep and goats and the elimination of the drug tends to be faster in goats compared to sheep.     

Oxygen binding and acid base status of the blood from the freshwater turtle, Phrynops hilarii

Oxygenation studies with the whole blood of Phrynops hilarii show a P50 of 38 torr at extracellular pH (pHe) of 7.4 which corresponds to an intracellular pH (pHi) of 7.05 at 25 degrees C. The blood CO2 Bohr effect was -0.56 when related to pHi. pHi is related to pHe by the following equation: pHi = 0.75.pHe + 1.54 (r = 0.99); pHi = 0.72. pHe + 1.72 (r = 0.96) at 10 and 25 degrees C respectively. Blood pHe, for 25 degrees C, was 7.519 +/- 0.254 (n = 6). Blood gas partial pressures were: pCO2 = 25.8 +/- 3.8 torr (n = 6); pO2 = 61.7 +/- 21.2 torr (n = 6). The major red cell phosphates, in mmole/l erythrocytes, n = 6, were: ATP (3.66 +/- 0.86); GTP (0.53 +/- 0.28); 2.3-DPG (0.32 +/- 0.12) and inorganic phosphates (2.00 +/- 0.35). The plasma inorganic ion composition, n = 6, was, in mEq/l: K+ (3.04 +/- 0.40); Na+ (148.4 +/- 12.6); Ca2+ (4.75 +/- 1.32); Cl- (106.6 +/- 5.0). Additional blood parameters of interest (n = 6) were: lactate (2.07 +/- 1.72 mM in plasma); erythrocytes/mm3 (416 X 10(3) +/- 4.6 X 10(3)); leucocytes/mm3 (44636 +/- 2618); haematocrit (%) (14.5 +/- 3.6); haemoglobin, g/dl (3.2 +/- 0.5); plasma protein g/dl (4.4 +/- 0.4); osmolarity (293 +/- 10 mOsm/l). The non-bicarbonate buffer value was -22.6 mmol/kg H2O/pH. For a constant CO2 content, delta pHe/delta t = 0.0141 +/- 0.002 (n = 18) and delta pHi/delta t = 0.0157 +/- 0.003 (n = 18).     

Development and validation of a gas chromatography-mass spectrometry method for the determination of isoimperatorin in rat plasma and tissue: application to the pharmacokinetic and tissue distribution study

Isoimperatorin is one of the major furanocoumarins isolated from the dried root of Angelica dahuricae Benth.et Hook. The aim of the present study is to develop a procedure based on gas chromatography-mass spectrometry (GC-MS) to describe the analysis of isoimperatorin in rat plasma and tissue. The method was set up and adapted for the analysis of small biological samples taken from rats. Biological samples were extracted by liquid-liquid extraction. Extracted compounds were acetic ether/light petroleum (1:2). They were separated by GC on a DB-5MS analytical column and determined by a quadrupole mass spectrometer detector operated under selected ion monitoring mode. Excellent linearity was found between 0.027-5.32 microg/mL (r >0.99) for plasma samples and 0.108-21.28 microg/g (r >0.99) for the tissue samples. The limit of detection (LOD) was 1.0 ng/mL or 1.0 ng/g (three times signal/noise ratio). Within- and between-day precisions expressed as the relative standard deviation (RSD) for the method were 2.81-5.22% and 4.72-6.52%, respectively. The method recoveries for all samples were >80%. The main pharmacokinetic parameters obtained were T(max)=(1.06+/-0.12)h, C(max)=(0.72+/-0.14) microg/mL, AUC=(2.11+/-0.29)h microg/mL and K(a)=(1.76+/-0.13)/h. The concentrations of isoimperatorin in rat liver, heart, cerebellum and cerebrum were higher than those in other organs. The results presented here clearly indicate that this proposed method could be applicable to investigate the pharmacokinetic and tissue distribution of isoimperatorin in rats after administration.     

HPLC-atmospheric pressure chemical ionization mass spectrometric method for enantioselective determination of R,S-propranolol and R,S-hyoscyamine in human plasma

A method for the simultaneous determination of R- and S-propranolol and R- and S-hyoscyamine in human plasma was developed, validated and applied to the analysis of samples from a clinical study. Sample preparation was performed by solid-phase extraction of 2 ml of human plasma using Oasis MCX cartridges and the enantioselective separations were achieved using a Chirobiotic V chiral stationary phase. The chromatography was carried out using gradient elution with a mobile phase composed of methanol:acetic acid:triethylamine which was varied from 100:0.05:0.04 to 100:0.05:0.1 (v/v/v) over 30 min and delivered at a flow rate 1 ml/min. The internal standard was R,S-propranolol-d7 and the analytes were quantified using a single quadrupole mass spectrometer employing APCI interface operated in the positive ion mode with single ion monitoring. The enantioselective separation factors, alpha, were 1.15 and 1.07 for S- and R-propranolol and R- and S-hyoscyamine, respectively. The standard curves were linear for all target compounds with coefficients of determination (r2) ranging from 0.9977 to 0.9999. The intra- and inter-day precision and accuracy were 相似文献   

Transport of uterine PGF2 alpha to the ovaries by systemic circulation and local lymphovenous-arterial diffusion during luteolysis in sheep.

The theory of countercurrent vascular transfer of PGF2 alpha during luteolysis was examined. In the first experiment, pulmonary clearance of PGF2 alpha was determined to re-examine whether the total amount of PGF2 alpha was degraded in the lungs after one passage. Cardiac output was measured by the Fick method and PGF2 alpha by radio-immunoassay before and after vascular lung supply, using pulmonary catheterization and the interventional radiology method in ten anaesthetized ewes on day 16 of the oestrous cycle. Cardiac output remained stable (7156 +/- 439 ml min-1). Infusion of 5 iu oxytocin resulted in an increase in plasma PGF2 alpha concentrations at 30 min in the uterine vein and the pulmonary and femoral arteries (3811 +/- 806, 224 +/- 55 and 18 +/- 4 pg ml-1, respectively). The PGF2 alpha concentrations decreased exponentially and the half-time decreases were 27 (r = 0.99), 16 (r = 0.99) and 18 (r = 0.98) min, respectively. Pulmonary clearance of PGF2 alpha was estimated at 6338 +/- 451 ml min-1. In a second experiment, an arterio-arterial gradient of plasma PGF2 alpha concentrations was analysed between the proximal and distal segments of the ovarian artery to verify whether the total amount of PGF2 alpha flowing to the ovary was from the local venous-arterial countercurrent pathway. Surgical catheterization techniques were performed on 11 ewes on day 16 of the oestrous cycle. The ovarian arterial blood flow was measured by the implantable Doppler method (8 +/- 1 ml min-1). The maximum plasma PGF2 alpha concentrations in the femoral and distal ovarian arteries were 23 +/- 6 and 42 +/- 11 pg ml-1 (P < 0.05), respectively. Plasma PGF2 alpha decreased exponentially in the femoral artery and the half-time decrease was 26 min (r = 0.98), and in the distal ovarian artery close to the ovary PGF2 alpha decreased linearly and the half-time decrease was 108 min (r = 0.96). Consequently, the arterio-arterial diffusion gradient of PGF2 alpha concentrations was extended to 3 h. These experiments showed that the PGF2 alpha flow rate in the pulmonary artery was 42.275 +/- 10.793 micrograms per 150 min (n = 10) and the systemic arterial PGF2 alpha flow rate was 5.359 +/- 1.658 micrograms per 150 min (n = 10). Therefore, 12% of the PGF2 alpha was not oxidized by the lungs. The proximal ovarian PGF2 alpha flow rate was 6.909 +/- 2.341 ng per 150 min, while the distal flow rate was 21.003 +/- 5.703 ng per 150 min (n = 11). Thus, 33% of the PGF2 alpha was transported rapidly to the ovary via the systemic route, while 67% was transported by slow local countercurrent diffusion, which extended the duration of luteolytic activity to four times that of the PGF2 alpha surge. These results indicate both rapid systemic transport of PGF2 alpha to the ovaries and a slower buffer mechanism involving a local diffusion pathway, rather than a direct countercurrent system.     

Heart design: free ADP scales with absolute mitochondrial and myofibrillar volumes from mouse to human

Our aim was to estimate a number of bioenergetic parameters in the beating mouse, rat and guinea pig heart in situ and compare the values to those in hearts of mammals over a 2000-fold range in body mass. For the mouse, rat and guinea pig heart, we report a phosphorylation ratio of 1005+/-50 (n=16), 460+/-32 (n=10) and 330+/-22 (n=5) mM(-1) and a free cytosolic [ADP] concentration of 13, 18 and 22 microM, respectively. When each parameter was plotted against body mass, they scaled closely to the quarter power (-0.28, r=0.99 and -0.23, r=0.97). A similar regression slope was found when the inverse of free [ADP] was plotted against absolute mitochondrial (slope=-0.26, r=0.99) and myofibrillar volumes (slope=-0.24, r=0.99). The similar slopes indicate that the ratio of absolute mitochondria and myofibrillar volumes in the healthy mammalian heart is a constant, and independent of body size. In conclusion, our study supports the hypothesis that the mammalian heart has a number of highly conserved thermodynamic and kinetic parameters that obey quarter-power laws linking the phosphorylation ratio, ATP turnover rates, free [ADP] and absolute mitochondrial volumes to body size. The results are discussed in terms of possible mechanisms and potential deviations from these laws in some disease states.     

Pharmacokinetic study of orphenadrine using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS)

We developed and validated a simple, rapid, and accurate HPLC-MS/MS method with simple protein precipitation for the determination of orphenadrine. Injection-to-injection running time was 3 min with a retention time of orphenadrine of 1.1 min. The linear assay range was 1-200 ng/mL (r2 > 0.99). The intra- and inter-assay imprecisions were CV 0.6-4.2% and CV 1.6-6.1%, respectively. The accuracy, extraction recovery, specificity and stability were satisfactory. Using the measured plasma concentrations of orphenadrine in 24 healthy subjects, pharmacokinetic profiles of orphenadrine were evaluated (AUC(0-72,) 1565+/-731 ng h/mL, Cmax 82.8+/-26.2 ng/mL, Tmax 3.0+/-0.9 h, elimination half-life 25.8+/-10.3 h).     

A comparison of chloride, bromide, and sucrose dilution volumes in neonatal pigs

Use of 36Cl, 82Br, and [3H]sucrose to estimate extracellular water volume was evaluated in 14 piglets (7-14 days old). 36Cl and 82Br were distributed in approximately the same volume, but a period of 5-6 hr after injection was required to reach equilibrium in the neonatal pig. Dilution volumes calculated before equilibration (2-5 hr) for 36Cl (326 +/- 11 ml/kg) and 82Br (328 +/- 13 ml/kg) were different from equilibration (6-8 hr) phase volumes (356 +/- 13 ml/kg and 355 +/- 13 ml/kg, respectively; P less than 0.001). A 3-hr sample estimated the same volume distribution calculated by extrapolation of the 6- to 8-hr period because of the relationship between the two slopes of the plasma clearance curves. After the 82Br and 36Cl had achieved equilibration, each was distributed in a volume equivalent to total body chloride space (362 +/- 29 ml/kg) measured by neutron activation; no statistical differences were found (P = 0.6). The early equilibration phase measured a 10% smaller, faster exchangeable fraction of total body Cl. Sucrose dilution volume (332 +/- 19 ml/kg) required multiple plasma samples for extrapolation and measured a dilution volume 7% smaller (P less than 0.05) than total body chloride space.     

The relation between selenium, zinc and copper concentration and the trace element dependent antioxidative status.

An imbalance in the antioxidative system was connected with the development of a number of pathological processes. In order to receive values of a healthy group and to evaluate pathological changes of the trace element dependent antioxidative status in future, we investigated 99 healthy volunteers (45 male and 54 female, mean age 37.4 +/- 11.7 years). We determined the concentrations of Se, Cu and Zn, the concentrations of malondialdehyde (MDA) and the activities of the Se dependent glutathione peroxidase (GSH-Px) and the Zn/Cu dependent superoxide dismutase (SOD). The plasma concentrations (mean +/- SD) for Se, Cu and Zn were 0.84 +/- 0.10 micromol/l, 15.6 +/- 2.78 micromol/l and 12.6 +/- 1.80 micromol/l, resp., and for non protein-bound and protein bound MDA 0.27 +/- 0.07 micromol/l and 1.11 +/- 0.25 micromol/l, resp. The activity of GSH-Px in plasma and erythrocytes was 130 +/- 20.8 U/l and 19.8 +/- 4.18 U/mg Hb, resp. and of SOD in erythrocytes 3,159 +/- 847.2 U/g Hb. In plasma positive correlations have been found between Se concentrations and GSH-Px activities (p < 0.002, r = 0.31) and between GSH-Px activities and concentrations of non protein-bound MDA (p = 0.004, r = 0.28). A negative correlation has been observed between GSH-Px activities in plasma and in erythrocytes. The higher the concentrations of Cu in erythrocytes, the higher were the activities of SOD (p = 0.03, r = 0.22) and GSH-Px in erythrocytes (r = 0.26, p = 0.01), while an increasing activity of GSH-Px in these cells correlated with a decreasing concentration of non protein-bound MDA (r = -0,31, p = 0.002). An increase in BMI was connected with an increase in protein-bound MDA and a decrease in GSH-Px activities in pLasma (p = 0.002 and r = 0.23). As the results demonstrate, Se and Cu concentrations in erythrocytes can improve the trace element dependent antioxidative status.     

Circulating leptin concentrations are positively related to leptin messenger RNA expression in the adipose tissue of fetal sheep in the pregnant ewe fed at or below maintenance energy requirements during late gestation

We have investigated the effects of maternal undernutrition during late gestation on maternal and fetal plasma concentrations of leptin and on leptin gene expression in fetal perirenal adipose tissue. Pregnant ewes were randomly assigned at 115 days of gestation (term = 147 +/- 3 days [mean +/- SEM]) to either a control group (n = 13) or an undernourished group (n = 16) that received approximately 50% of the control diet until 144-147 days of gestation. Maternal plasma glucose, but not leptin, concentrations were lower in the undernourished ewes. A significant correlation was found, however, between mean maternal plasma leptin (y) and glucose (x) concentrations (y = 2.9x - 2.4; r = 0.51, P < 0.02) when the control and undernourished groups were combined. Fetal plasma glucose and insulin, but not fetal leptin, concentrations were lower in the undernourished ewes, and no correlation was found between mean fetal leptin concentrations and either mean fetal glucose or insulin concentrations. A positive relationship, however, was found between mean fetal (y) and maternal (x) plasma leptin concentrations (y = 0.18x + 0.45; r = 0.66, P < 0.003). No significant difference was found in the relative abundance of leptin mRNA in fetal perirenal fat between the undernourished (0.60 +/- 0.09, n = 10) and control (0.70 +/- 0.08, n = 10) groups. Fetal plasma concentrations of leptin (y) and leptin mRNA levels (x) in perirenal adipose tissue were significantly correlated (y = 1.5x +/- 0.3; r = 0.69, P < 0.05). In summary, the capacity of leptin to act as a signal of moderate maternal undernutrition may be limited before birth in the sheep.     

VCO2 and VE kinetics during moderate- and heavy-intensity exercise after acetazolamide administration.

The effect of carbonic anhydrase inhibition with acetazolamide (Acz) on CO2 output (VCO2) and ventilation (VE) kinetics was examined during moderate- and heavy-intensity exercise. Seven men [24 +/- 1 (SE) yr] performed cycling exercise during control (Con) and Acz (10 mg/kg body wt iv) sessions. Each subject performed step transitions (6 min) in work rate from 0 to 100 W [below ventilatory threshold (VET)]. VE and gas exchange were measured breath by breath. The time constant (tau) was determined for exercise VET by using a three-component model (fit from the start of exercise). VCO2 kinetics were slower in Acz (VET, MRT = 75 +/- 10 s) than Con (VET, MRT = 54 +/- 7 s). During VET kinetics were faster in Acz (MRT = 85 +/- 17 s) than Con (MRT = 106 +/- 16 s). Carbonic anhydrase inhibition slowed VCO2 kinetics during both moderate- and heavy-intensity exercise, demonstrating impaired CO2 elimination in the nonsteady state of exercise. The slowed VE kinetics in Acz during exercise 相似文献   

Inverse relationships between steroid concentration and volume in preovulatory follicles of the golden hamster

In order to investigate variations in the microenvironment of oocytes within a cohort of maturing follicles the follicular volumes as well as the intrafollicular concentrations of oestradiol (E2) and progesterone (P) were measured in the golden hamster. At 10 h before ovulation the follicular volumes varied from 0.009 to 0.037 mm3 (mean +/- SD: 0.0187 +/- 0.0071 mm3; n = 36). Large follicles (greater than 0.025 mm3; n = 8) contained statistically significantly lower E2 and P levels (30.1 +/- 10.4 and 517 +/- 113 mumol/l, respectively) than the medium sized group (less than 0.025 and greater than 0.015 mm3; n =20): 46.9 +/- 16.0 (P less than 0.02) and 919 +/- 264 (P less than 0.0001) mumol/l, respectively. Small follicles (less than 0.015 mm3) showed the highest steroid levels: 97.0 +/- 33.3 and 1590 +/- 517 mumol/l for E2 and P (P less than 0.001 versus the medium sized group values). Correlation coefficients for the steroid concentrations and the follicular volumes appeared to be -0.674 for E2 and -0.612 for P (P less than 0.001). At the time studied a positive correlation between E2 and P concentrations in the follicles was found: r = 0.655 (P less than 0.001). The mean ratios of intrafollicular over serum steroid concentrations appeared to be approx 36 x 10(3) in the case of E2 and about 17 x 10(3) in the case of P. These results clearly show that there is an inverse relationship between follicular volume and intrafollicular steroid concentrations. The presence of a fine regulatory mechanism for a collective maturation of follicles is hypothesized.     

Quantitative ultrastructural study of nucleolus-organizing regions at some stages of the cell cycle (G0 period, G2 period, mitosis)

The quantitative characteristics of chromosomal nucleolus-organizing regions (NORs) and some other nucleolar components were studied on ultra-thin sections of pig embryo kidney cells (PK cells). It was shown that: 1) nucleoli-per-cell volumes were 3 times smaller in the G0 period than in the G2 period; 2) the number of fibrillar centers (FCs) per cell in the G0 period, the G2 period, and at metaphase was equal to 7, 33.7, and 8, respectively; 3) mean volumes of individual FCs in the G0 period (0.033 +/- 0.005 micron3), G2 period (0.014 +/- 0.001 micron3), and at metaphase (0.025 +/- 0.002 micron3) were significantly different; 4) the total volumes of FCs calculated per haploid set of chromosomes were practically the same in the G0 (0.105 micron3) and G2 (0.107 micron3) periods, but were twice as large as those at metaphase (0.04-0.05 micron3). These data show that partial activation and inactivation of ribosomal genes in interphase PK cells are not accompanied by a considerable change in the total volume of FCs and may be due to the fragmentation and fusion of individual FCs. Complete inactivation of ribosomal genes in mitosis results in a decrease of total volumes of FCs per cell; 5) in G0 and G2 periods the total volume of the dense fibrillar component per nucleolus is practically proportional to the nucleolus volume (r = 0.99); 6) in the G2 period, the nucleolus volume is also proportional to the number of FCs (r = 0.99; 7) the volume of the dense fibrillar component within individual fibrillar complexes is not a constant one.(ABSTRACT TRUNCATED AT 250 WORDS)     

Development and application of a chiral high performance liquid chromatography assay for pharmacokinetic studies of methadone

Methadone enantiomers and EDDP, the main metabolite of methadone, were separated (R(s) = 2.0 for methadone enantiomers) following liquid-liquid extraction from human serum and urine followed by reverse-phase high-performance liquid chromatography on a derivatized beta-cyclodextrin column and quantified at therapeutic concentrations with ultraviolet detection. Detector response was linear (r(2) > 0.98) to 1,000 and 2,500 ng x mL(-1) for methadone enantiomers and EDDP, respectively. The limit of quantification from a 1-mL biological sample was 2.5 and 5 ng x mL(-1) for methadone enantiomers and EDDP, respectively. Interday variation was <13% and intraday variation was <8% for the analytes of interest. The assay was applied to plasma protein and erythrocyte binding studies and a 96-h pharmacokinetic study in two healthy female volunteers following oral dosing with rac-methadone. The binding of methadone to plasma proteins was enantioselective with the active (-)-(R) enantiomer having the highest free fraction (mean +/- SD: 21.2+/-7.6% vs. 13.3+/-6.2% for (+)-(S)-methadone, n = 8). Binding of methadone to erythrocytes was not apparently enantioselective (38.6+/-1.3% and 38.1+/-1.4% bound for (-)-(R)- and (+)-(S)-methadone, respectively). The pharmacokinetic study revealed enantioselective disposition of methadone in one volunteer but not in the other. EDDP was observed in urine but was only in small or undetectable concentrations in serum. The method is applicable to in vitro and pharmacokinetic studies of rac-methadone disposition in humans.     

Effect of atorvastatin on plasma apoE metabolism in patients with combined hyperlipidemia

Atorvastatin, a synthetic HMG-CoA reductase inhibitor used for the treatment of hyperlipidemia and the prevention of coronary artery disease, significantly lowers plasma cholesterol and low-density lipoprotein cholesterol (LDL-C) levels. It also reduces total plasma triglyceride and apoE concentrations. In view of the direct involvement of apoE in the pathogenesis of atherosclerosis, we have investigated the effect of atorvastatin treatment (40 mg/day) on in vivo rates of plasma apoE production and catabolism in six patients with combined hyperlipidemia using a primed constant infusion of deuterated leucine. Atorvastatin treatment resulted in a significant decrease (i.e., 30-37%) in levels of total triglyceride, cholesterol, LDL-C, and apoB in all six patients. Total plasma apoE concentration was reduced from 7.4 +/- 0.9 to 4.3 +/- 0.2 mg/dl (-38 +/- 8%, P < 0.05), predominantly due to a decrease in VLDL apoE (3.4 +/- 0.8 vs. 1.7 +/- 0.2 mg/dl; -42 +/- 11%) and IDL/LDL apoE (1.9 +/- 0.3 vs. 0.8 +/- 0.1 mg/dl; -57 +/- 6%). Total plasma lipoprotein apoE transport (i.e., production) was significantly reduced from 4.67 +/- 0.39 to 3.04 +/- 0.51 mg/kg/day (-34 +/- 10%, P < 0.05) and VLDL apoE transport was reduced from 3.82 +/- 0.67 to 2.26 +/- 0.42 mg/kg/day (-36 +/- 10%, P = 0.057). Plasma and VLDL apoE residence times and HDL apoE kinetic parameters were not significantly affected by drug treatment. Percentage decreases in VLDL apoE concentration and VLDL apoE production were significantly correlated with drug-induced reductions in VLDL triglyceride concentration (r = 0.99, P < 0.001; r = 0.88, P < 0.05, respectively, n = 6). Our results demonstrate that atorvastatin causes a pronounced decrease in total plasma and VLDL apoE concentrations and a significant decrease in plasma and VLDL apoE rates of production in patients with combined hyperlipidemia.     

A non-invasive method for measuring inspiratory muscle fatigue during progressive isocapnic hyperventilation in man

Eleven normal adults each performed a ten minute progressive isocapnic hyperventilation (PIHV) test in which ventilatory levels were increased every two minutes. All subjects exhibited mechanical fatigue by failing to maintain the target of 80% of maximum voluntary ventilation (MVV). The mean ventilation at this level was 67.5 +/- 1.4% MVV. This fatigue was accompanied by a fall in transdiaphragmatic pressure. During the test the EMG of the sternomastoid (SM) was monitored by surface electrodes and was analyzed using fast-fourier transform. The centroid frequency (Fc) fell as ventilation increased, and correlated negatively with the inability to achieve target ventilation(r = -0.99, p less than 0.015). Five subjects performed the test while the diaphragmatic EMG was recorded from an oesophageal electrode (DIes) and from surface electrodes (DIs). The Fc of DIes fell with increasing ventilation levels (r = -0.95, p less than 0.05) and there was a correlation between the Fc changes of both DIes and the SM (r = -0.92, p less than 0.001). The Fc of DIs did not correlate with either mechanical performance or the Fc of DIes, because of contamination of surface signals by signals from expiratory muscles. It is concluded that the PIHV along with surface monitoring of EMG activity from the sternomastoid can serve as a non-invasive method for evaluating inspiratory muscle fatigue.     

Muscle capillary blood flow kinetics estimated from pulmonary O2 uptake and near-infrared spectroscopy.

The near-infrared spectroscopy (NIRS) signal (deoxyhemoglobin concentration; [HHb]) reflects the dynamic balance between muscle capillary blood flow (Q(cap)) and muscle O(2) uptake (Vo(2)(m)) in the microcirculation. The purposes of the present study were to estimate the time course of Q(cap) from the kinetics of the primary component of pulmonary O(2) uptake (Vo(2)(p)) and [HHb] throughout exercise, and compare the Q(cap) kinetics with the Vo(2)(p) kinetics. Nine subjects performed moderate- (M; below lactate threshold) and heavy-intensity (H, above lactate threshold) constant-work-rate tests. Vo(2)(p) (l/min) was measured breath by breath, and [HHb] (muM) was measured by NIRS during the tests. The time course of Q(cap) was estimated from the rearrangement of the Fick equation [Q(cap) = Vo(2)(m)/(a-v)O(2), where (a-v)O(2) is arteriovenous O(2) difference] using Vo(2)(p) (primary component) and [HHb] as proxies of Vo(2)(m) and (a-v)O(2), respectively. The kinetics of [HHb] [time constant (tau) + time delay [HHb]; M = 17.8 +/- 2.3 s and H = 13.7 +/- 1.4 s] were significantly (P < 0.001) faster than the kinetics of Vo(2) [tau of primary component (tau(P)); M = 25.5 +/- 8.8 s and H = 25.6 +/- 7.2 s] and Q(cap) [mean response time (MRT); M = 25.4 +/- 9.1 s and H = 25.7 +/- 7.7 s]. However, there was no significant difference between MRT of Q(cap) and tau(P)-Vo(2) for both intensities (P = 0.99), and these parameters were significantly correlated (M and H; r = 0.99; P < 0.001). In conclusion, we have proposed a new method to noninvasively approximate Q(cap) kinetics in humans during exercise. The resulting overall Q(cap) kinetics appeared to be tightly coupled to the temporal profile of Vo(2)(m).     

Plasma obestatin is lower at fasting and not suppressed by insulin in insulin-resistant humans

Obestatin, a recently discovered 23-amino acid peptide, is involved in the regulation of appetite and body weight in antagonistic fashion to ghrelin, both deriving from a common precursor peptide. Ghrelin was shown to be associated with insulin resistance, which may also affect obestatin. We investigated the association between insulin resistance and plasma concentrations of obestatin and ghrelin in nondiabetic individuals with high (IS; n = 18, 13 females and 5 males, age 47 +/- 2 yr, BMI = 25.5 +/- 0.9 kg/m(2)) and low (IR; n = 18, 12 females and 6 males, age 45 +/- 2 yr, P = 0.49, BMI = 27.5 +/- 1.1 kg/m(2), P = 0.17) insulin-stimulated glucose disposal (M), measured by 2-h hyperinsulinemic (40 mU.min(-1).m(-2)) isoglycemic clamp tests. M(100-120 min) was higher in IS (10.7 +/- 0.7) than in IR (4.4 +/- 0.2 mg.min(-1).kg(-1), P < 10(-9)), whereas insulin-dependent suppression of free fatty acids (FFA) in plasma was reduced in IR (71 +/- 6% vs. IS: 82 +/- 5%, P < 0.02). In both groups, plasma ghrelin concentrations were comparable at fasting and similarly reduced by 24-28% during insulin infusion. IR had lower fasting plasma obestatin levels (383 +/- 26 pg/ml vs. IS: 469 +/- 23 pg/ml, P < 0.02). Clamp insulin infusion reduced plasma obestatin to approximately 81% of basal values in IS (P < 0.00002), but not in IR. Fasting plasma obestatin was correlated positively with M (r = 0.34, P = 0.04), HDL cholesterol (r = 0.45, P = 0.01), and plasma ghrelin concentrations (r = 0.80, P < 0.000001) and negatively with measures of adiposity, plasma FFA during clamp (r = -0.42, P < 0.01), and systolic blood pressure (r = -0.33, P < 0.05). In conclusion, fasting plasma concentrations of obestatin, but not of ghrelin, are reduced in insulin resistance and are positively associated with whole body insulin sensitivity in nondiabetic humans. Furthermore, plasma obestatin is reduced by insulin in insulin-sensitive but not in insulin-resistant persons.