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This paper presents a consistent quantum mechanical model of Child-Langmuir (CL) law, including electron exchange-correlation interaction, electrode's surface curvature, and finite emitter area. The classical value of the CL law is increased by a larger factor due to the electron tunneling through the space-charge potential, and the electron exchange-correlation interaction becomes important when the applied gap voltage Vg and the gap spacing D are, respectively, on the order of Hartree energy level, and nanometer scale. It is found that the classical scaling of Vg(3/2) and D(-2) is no longer valid in the quantum regime, and a new scaling of Vg(1/2) and D(-4) is established. The smooth transition from the classical regime to the quantum regime is also demonstrated.

A semi-automatic, high-throughput method has been developed to rapidly assess plasma protein binding of new chemical entities in drug discovery phase. New chemical entities are mixed with plasma and the unbound fractions are separated from the bound fraction by ultrafiltration in a 96-well filtrate assembly. The unbound fractions are then analyzed by fast liquid chromatography-tandem mass spectrometry (LC-MS/MS). Sample handling is automated by a robotic system. Employing a cocktail approach where multiple new chemical entities are allowed to bind to plasma proteins in the same well has further increased the throughput. We have validated the method with 12 commercially available compounds. The plasma protein binding data obtained by this method are comparable with the literature values. This method enables the determination of protein binding for 32 compounds in one single experiment instead of 1-2 compounds using the conventional methods.

The first observations and measurements are reported on suppression of the third-order intermodulation (IM3) product arising from nonlinear mixing of two drive frequencies in a klystron, by externally injecting a wave at the IM3 product frequency. Optimum amplitude and phase of the injected wave for maximum suppression are examined. Results indicate that suppression of the IM3 product by as much as 30 dB can be achieved. Experimental results compare favorably with predictions of a 1D simulation code that takes into account all kinematical and dynamical effects including charge overtaking and space charge forces.

We present a novel, simple asymptotic expansion for the spectrum of radiation that is backscattered from a laser by a counterpropagating (or copropagating) electron. The solutions are presented in such a way that they explicitly show the relative merit of using an intense laser and of an energetic electron beam in x-ray production in the single particle regime. Simple scaling laws are given.

Sixty-four compounds with diverse structures were used in evaluation of intrinsic clearance by various hepatocyte preparations from rats, dogs, monkeys, and humans. Intrinsic clearance (CL(int)) was calculated from the ratio of the initial amount of the test compound minus the amount remaining after 2 h of incubation and the corresponding area under the concentration versus time curve. The predictive potential of this in vitro model was tested by comparing the intrinsic clearance with in vivo clearance using linear regression analysis. In addition, the intrinsic clearance values obtained from three different types of hepatocytes (cryopreserved, fresh, and sandwich-cultured) from the same species were compared to determine the influence of preservation and culture conditions. It seems that intrinsic clearance determined from human cryopreserved hepatocyte (R(2) = 0.87) was the best predictor for the corresponding human in vivo clearance. Dog and rat hepatocyte clearances were also demonstrated to be reasonable predictors (R(2) ranged 0.68-0.85 in dogs and 0.65-0.72 in rats) for their corresponding in vivo clearances. Monkey hepatocyte clearance seems to be the worst predictor for the corresponding in vivo clearance (R(2) = 0.35-0.67). Comparison of intrinsic clearance generated from cryopreserved and fresh hepatocytes demonstrated a very good correlation in dog (R(2) = 0.82) followed by rat (R(2) = 0.77), and then by monkey (R(2) = 0.70). A similar correlation profile was shown between cryopreserved hepatocytes and sandwich-cultured hepatocytes. These results demonstrated the predictive potential of intrinsic clearance for rat, dog, and human in vivo clearance, but also showed some limitation of the approach for monkey.

Purpose: The in vivo hepatic extraction ratio of cynomolgus monkeys was correlated with the corresponding in vitro extraction ratios that were determined in monkey microsomal incubations., Method: For compounds that are eliminated mainly through liver phase I metabolism, the extraction ratio calculated from liver microsomal stability studies should correlate with their in vivo hepatic extraction ratios and also with their oral bioavailability in monkey. We used both well-stirred and parallel tube models of intrinsic clearance for the correlation. We also calculated extraction ratios for compounds within a given therapeutic area from fraction absorbed values that were estimated from the Caco-2 absorption model., Result: The present data show that in vitro extraction ratios in monkey microsomes are predictive of the in vivo hepatic extraction ratios in monkeys. All compounds with high extraction ratio (>70%) in vivo were successfully classified as high-extraction-ratio compounds based on the in vitro monkey microsomal stability data. From the results of this study, it appears that the parallel tube model provided a slightly better classification than the well-stirred model. CONCULUSIONS: The present method appears to be a valuable tool to rapidly screen and prioritize compounds with respect to liver first-pass metabolism in monkeys at an early phase of drug discovery.

Introduction: Enzyme induction is undesirable in new drug discovery process, with consequences spanning from auto-induction to toxicity. Cytochrome P450 (CYP) 1A1 has long been known to be one of the metabolic enzymes involved in activating many procarcinogens, the first step toward tumor formation during chemical carcinogenesis. Induction of CYP1A1 during drug treatment may predispose the patients to some risk of chemical carcinogenesis., Methods: Based on the signal-transduction mechanism of CYP1A1 induction, a high-throughput reporter-gene system was established by stable transformation of H4IIE cells to incorporate the luciferase gene under control of CYP1A1 promoter. This stable cell line was validated with known CYP1A1 inducers, such as 3-methylcholanthrene (3-MC), beta-naphthoflavone (beta-NF), alpha-naphthoflavone (alpha-NF) and 3-indocarbinol. Thirty in-house new chemical entities (NCEs) were then screened with this reporter-gene system, and also administered to rats to evaluate in vivo CYP1A1 induction., Results: CYP1A1 reporter gene system can be used to identify strong inducers, such as 3-MC, beta-NF and alpha-NF, and weak inducers, such as 3-indocarbinol. In vitro induction of 30 in-house compounds in reporter gene system did not correlate with in vivo induction in rat liver microsome measured by ethoxyresorufin-O-dealkylation (EROD) activity, but had a reasonable correlation with Western blot signals., Discussion: This reporter-gene system may be useful in eliminating compounds that can cause CYP1A1 induction at an early stage of drug discovery.

This paper presents, for the first time, a simple analytic theory for the two-dimensional (2D) Child-Langmuir law. For electron emission over a finite patch on a planar cathode, the limiting current density is derived approximately from first principles. The scaling laws are in excellent agreement with simulation results. They predict the onset of virtual cathode formation in a 2D geometry; they also indicate that electrons emitted from a cathode over only a restricted area may have a current density much exceeding the classical (1D) Child-Langmuir value.

Using a mean-field theory, we have studied the quantum extension on the limiting current density in a crossed-field nanogap. When the gap spacing is less than the electron wavelength, our results show that the limiting current density is increased by a large factor from the classical values due to the effects of electron tunneling. The effects of the external magnetic field diminish with a decrease of gap spacing. Smooth transition from the classical regime to the quantum regime is demonstrated.

Objective: To compare the cellular yield and sensitivity of cancer detection by urine cytology using alcoholic carbowax as a prefixative and formal alcohol as a final fixative with the conventional centrifugation-and-smearing method., Study Design: Four hundred seventeen voided urine specimens were examined. Each specimen was divided into two equal portions. One portion was centrifuged and wet fixed (CS). The other was treated with alcoholic carbowax as a prefixative and then fixed in formal alcohol (CW). The smears were studied for cellular yield, morphology, cancer detection rate and typing., Results: The CW method yielded significantly more cells with excellent morphology. It detected 25 more abnormal cases out of a total of 86, including 5 more transitional cell carcinomas, as compared to the CS method. This finding was statistically significant (P < .0000003)., Conclusion: The CW method is simple and enhances cellular yield and the cancer detection rate as compared to the conventional centrifuge-and-smear technique.

A highly sensitive and selective HPLC-MS-MS method was developed for the determination of pramipexole in human plasma. The analytes, pramipexole and BHT-920 (internal standard), were extracted from plasma at basic pH with methyl tert.-butyl ether (MTBE). MTBE was evaporated to dryness and reconstituted in 100 microliters of (95:5) methanol-water. Chromatographic separation was achieved on a Zorbax SB-CN column with a mobile phase of (15:5:80) water-0.1 M ammonium acetate-methanol. The analytes were detected utilizing HPLC in conjunction with atmospheric pressure chemical ionization (APCI) tandem mass spectrometry (MS-MS). The assay was linear in the concentration ranges of 50 to 5000 pg/ml. The analysis of pooled quality controls (150, 750, and 3000 pg/ml) demonstrated excellent precision with relative standard deviations (R.S.D.) (n = 18) of 7.2%, 5.3% and 5.2%, respectively. The method is accurate with all intra-day (n = 6) and overall (n = 18) mean values being less than 11.7% from theoretical.

A sensitive and selective high-performance liquid chromatographic (HPLC) method was developed for the determination of pramipexole in human plasma and urine. Plasma/urine is made alkaline before pramipexole and BHT-920 (internal standard) are extracted by ethyl ether and back-extracted with a solution that contains heptanesulfonic acid. Separation is achieved by ion-pair chromatography on a Zorbax Rx C8 column with electrochemical detection at 0.6 V for plasma and ultraviolet detection at 286 nm for urine. The retention times of pramipexole and internal standard are approximately 14.4 and 10.7 min, respectively. The assay is linear in concentration ranges of 50 to 15,000 pg/ml (plasma) and 10 to 10,000 ng/ml (urine). The correlation coefficients are greater than 0.9992 for all curves. For the plasma method, the analysis of pooled quality controls (300, 3000, and 10,000 pg/ml) demonstrates excellent precision with relative standard deviations (R.S.D.) (n = 18) of 1.1%, 2.3%, and 6.8%, respectively. For the urine method, quality control pools prepared at 30, 300, and 3000 ng/ml had R.S.D. values (n = 18) of 2.9%, 1.7%, and 3.0%, respectively. The plasma and urine controls were stable for more than nine and three months, respectively. The mean recoveries for pramipexole and internal standard from plasma were 97.7% and 98.2%, respectively. The mean recoveries for pramipexole and internal standard from urine were 89.8% and 95.1%, respectively. The method is accurate with all intra-day (n = 6) and overall (n = 18) mean values for the quality control samples being less than 6.4 and 5.8% from theoretical for plasma and urine, respectively.

A simple, specific and sensitive high-performance liquid chromatographic (HPLC) method was developed for the determination of rifabutin in human plasma. Rifabutin and sulindac (internal standard) are extracted from human plasma using a C8 Bond Elut extraction column. Methanol (1 ml) is used to elute the compounds. The methanol is dried down under nitrogen and reconstituted in 250 microliters of mobile phase. Separation is achieved by HPLC on a Zorbax Rx C8 column with a mobile phase composed of 0.05 M potassium dihydrogen phosphate and 0.05 M sodium acetate at pH 4.0-acetonitrile (53:47, v/v). Detection is by ultraviolet absorbance at 275 nm. The retention times of rifabutin and internal standard were approximately 10.8 and 6.9 min, respectively. The assay is linear over the concentration range of 5-600 ng/ml. The quantitation limit was 5 ng/ml. Both intra-day and inter-day accuracy and precision data showed good reproducibility.

Objective: To compare the sensitivity of sputum cytology using the dithiothreitol homogenization method (DTT) with that of the standard pick-and-smear (PS) method in lung cancer detection., Study Design: In a two-year period, 2,428 sputum specimens were examined. Standard PS smears were prepared as controls; the remainder of the specimens were homogenized with DTT before smears were prepared. The sensitivity of the two methods in detecting positive cases was compared., Results: Among 260 positive cases, the DTT method detected 256, while the PS method detected only 212, representing a difference of 17%. The DTT method also detected 28 more suspicious cases and permitted definitive typing of the tumor in a further 11 cases., Conclusion: Homogenization of sputum with DTT is not only a simple and versatile method but also a more sensitive method as compared to the PS method in lung cancer detection.

Capillary electrophoresis has been used to directly identify and measure the neurotransmitter, dopamine, in two vesicular compartments in a single nerve cell of Planorbis corneus. Dopamine in the cytoplasm and in easily released transmitter vesicles was separated from dopamine in what are apparently non-functional storage vesicles. In this method, the two peaks in the electropherogram attributed to dopamine were differentiated based on cell lyse time in a non-physiological buffer. The measurements presented here suggest that of the total dopamine present in the cell: 24% is in the cytoplasmic and easily released compartments and 76% is more centrally located, perhaps in a reserve compartment, in the cell. This methodology provides the means to determine molecular species in subcellular compartments and should allow kinetic parameters associated with membrane lysing to be evaluated at single nerve cells.

Ultrasmall glucose sensors have been constructed by using platinum-deposited carbon ring microelectrodes with glucose oxidase. Response times as low as 270 ms have been obtained with these sensors. Moreover, there is a linear relationship between sensor tip diameter and response times. The use of these sensors has been demonstrated in the detection of glucose in single-cell cytoplasm of the large dopamine cell of the pond snail Planorbis corneus. Current responses obtained at these sensors implanted into a cell increase following injection of 2 pL of glucose solution (3 M) into the cell. Results obtained from these experiments show that these sensors are suitable for glucose monitoring in ultrasmall environments. In addition, characterizations of these sensors have been investigated under different O2 concentrations. At atmospheric oxygen concentrations, glucose levels in the submillimolar range can be measured without oxygen interference; however, oxygen interference can be substantial at low oxygen concentrations.

Naflon-coated ultrasmall platinum ring electrodes have been implanted in the giant dopamine neuron of the pond snail Planorbis corneus, and the oxygen concentration inside these single neurons has been estimated. Experimental data suggest that the intracellular oxygen level in the identified dopamine neuron of P. corneus is approximately 0.032 mM. The oxygen concentration immediately outside the cell (ca. 10 microns away from the cell) is 0.041 mM. Furthermore, staircase voltammetry can be used to monitor dynamic changes in oxygen concentration inside the cell after bathing with Ringer's solution saturated with air/oxygen. Data obtained for intracellular oxygen concentrations suggest that intracellular oxygen consumption is increased following potassium chloride-induced stimulation of these cells.

The purpose of this study was to determine whether dietary n-3 polyunsaturates (PUFA) can affect the frequency of N-nitrosomethylurea (NMU)-induced Ha-ras mutations in virgin female F344 rat mammary glands. Two groups of 15 rats each were fed isocaloric diets containing either 23% w/w corn oil or corn oil plus menhaden oil (1:1) at starting 14 d before NMU administration (day 50 of age) and continuing for 13 wk. Mammary gland samples were taken serially at 3, 5, 9, and 13 wk post-NMU treatment. Total cellular DNA was isolated and analyzed by a newly devised enriched amplification procedure that involves predigestion of normal Ha-ras alleles at codon 12, amplification of the mutant alleles, and restriction fragment length polymorphism analysis. Selective amplification enabled the detection of Ha-ras mutations as early as 3 wk post-NMU treatment. Approximately 40-50% of all glands and 75% of all rats tested had the Ha-ras codon 12 mutation. No significant differences were found between the two dietary groups at any time point, indicating that the mammary tumor-inhibiting effect of n-3 PUFA is probably not exerted at the level of the Ha-ras activation step in NMU tumorigenesis.

An additional effect not considered in our earlier papers is found to enhance the rate of growth of spiral modes in disk-shaped galaxies. This enhanced growth appears to be attributable to the combined influence of differential rotation and tangential forces. For certain galaxy models, rapid temporal growths are possible (comparable to those found in numerical experiments), but in many real galaxies, these tendencies to extreme growth rates are probably reduced to lower levels by competing effects such as wave absorption at inner Lindblad resonance.

Discrete growing spiral modes are calculated, on the basis of principles of stellar dynamics, according to the recently developed asymptotic theory of density waves for spiral structure in galaxies. From the growth rates obtained, it appears likely that several spiral modes coexist in a given disk galaxy. General properties of these modes are described for two galaxy models, and some of their important observational implications are discussed.

A numerical scheme has been devised to calculate discrete spiral modes in a fluid-dynamical model of disk galaxies. The procedure is free from the usual restrictions imposed upon the asymptotic theory. A radiation condition has been formulated as one of the boundary conditions. For a test model galaxy with moderately low surface density, quantitative agreement is found between the asymptotic spiral modes and the corresponding ones obtained from the present scheme. The present scheme should find ready application to the study of open spiral modes.