Diazepam effects and kinetics in Caucasians and Orientals.

Mental and psychomotor effects and diazepam kinetics were studied in Caucasian and Orientals. 12 Caucasian and 13 Oriental young adults received on one of two occasions, separated by 2 weeks, either 0.2-mg/kg diazepam or saline intravenously. Serum diazepam and desmethyldiazepam concentrations were measured by electron-capture gas-liquid chromatography in samples drawn up to 72 hr after injection. Serum protein binding was measured by equilibrium dialysis. Subjects were tested on a battery of psychological tests before and 0.5, 2, and 4 hr after treatment. While the free fraction of diazepam was identical in both races (0.02), volume of distribution at steady state (Vdss) was different when calculated as absolute volume (Vdss = 76.55 +/- 9.63 l in Caucasians and 54.96 +/- 4.55 l in Orientals, p = 0.04) and marginally significant when corrected for body weight (Vdssl/kg = 1.10 +/- 0.11 in Caucasian and 0.88 +/- 0.05 in Orientals, p = 0.07). total body clearance (Cl), but not elimination half-life (t 1/2), was higher in Caucasians than Orientals, p less than 0.01; t 1/2 = 37.70 +2- 5.53 hr in Caucasians and 41.77 +/- 3.80 in Orientals). Desmethyldiazepam levels were higher in Orientals than Caucasians. Mental and psychomotor effects were maximal at the first session (0.5 hr), followed by complete recovery by the 4-hr session. Effects were similar in both groups. If repeated dosing causes a higher rate of cumulated diazepam serum levels in Orientals, as expected, there might be deeper brain depression in that group.

Topical carbonic anhydrase inhibitors.

Ethoxzolamide and several derivatives (1-6) were synthesized and evaluated for carbonic anhydrase inhibition (CAI), solubility, pKa, distribution, and corneal permeability. The 6-hydroxy (5) and, particularly, the 6-chloro (6) analogues have the best combination of properties for penetrating the site of action and reducing intraocular pressure. Both 5 and 6 exhibited topical effectiveness in the normal rabbit, with 6 showing greater potency.

N-Substituted sulfonamide carbonic anhydrase inhibitors with topical effects on intraocular pressure.

N-Methylacetazolamide was shown to be active topically in reducing intraocular pressure (IOP) to a small but statistically significant level in the normotensive rabbit eye. In vivo experiments with N-methylacetazolamide suggest that ocular metabolism to acetazolamide was responsible for the observed topical activity. Examination of initial rate kinetics of carbonic anhydrase catalyzed p-nitrophenyl acetate hydrolysis showed that N-methylacetazolamide was a competitive inhibitor, in contrast to noncompetitive inhibition seen with acetazolamide and other primary sulfonamide inhibitors. N-Substituted and unsubstituted 4-chlorobenzene- and 4-nitrobenzenesulfonamides were also synthesized, and their biochemical characteristics and in vivo ability to lower IOP when applied topically were determined. The primary sulfonamides were reversible noncompetitive inhibitors of carbonic anhydrase, with no effect on IOP after topical administration. 4-Nitrobenzene- and 4-chlorobenzenesulfonamides containing both N-hydroxy and N-methyl substituents were model irreversible inhibitors of carbonic anhydrase and exhibited a trend toward topical activity in reducing IOP in normotensive rabbit eyes. Therefore, this paper describes the synthesis and characterization of two types of carbonic anhydrase inhibitors; the N-methyl-substituted sulfonamides are reversible competitive inhibitors of carbonic anhydrase, while the N-hydroxy-N-methyl-substituted sulfonamides are irreversible inhibitors.

A bioavailability comparison in rabbits of two steroids formulated as high-viscosity gels and reference aqueous preparations.

In order to assess the bioavailability of steroids from a high-viscosity gel, rabbit cornea and aqueous humor levels were measured over 12 hr following topical instillation of a gel and reference preparation. Concentrations associated with a 1% tritiated prednisolone acetate gel were found to be approximately four times larger than those of the reference preparation. A graphical comparison of the 1% tritiated prednisolone sodium phosphate data showed the area under corneal and aqueous time curves to be five and 10 times greater, respectively, with the gel than data associated with the reference preparation. The gel vehicle is well retained in the rabbit eye and is responsible for the large increases in bioavailability.

The effect of flurbiprofen on herpes simplex virus type 1 stromal keratitis in mice.

The use of steroidal compounds to reduce the inflammation and scarring associated with herpes simplex virus type 1 (HSV-1) stromal keratitis can result in severe exacerbation of the corneal disease. We compared the nonsteroidal anti-inflammatory drug (NSAID) flurbiprofen sodium with dexamethasone for the treatment of HSV-1 induced corneal stromal disease in an inbred mouse model. Stromal disease was induced by the direct intrastromal injection of HSV-1. A stromal opacity and corneal neovascularization (CNV) developed in 100% of the injected eyes, with no epithelial involvement until late in the course, when the stromal disease was quite severe, such that it was possible to test the effectiveness of drugs in animals with an intact epithelium. Dexamethasone treatment had a variable effect on the severity of disease, ranging from a significant reduction in severity to significant exacerbation of disease, compared with placebo-treated controls. The most frequent effect of dexamethasone treatment was a worsening of corneal stromal opacities and CNV. In contrast, treatment with the NSAID did not exacerbate HSV-1 stromal disease. Flurbiprofen treatment resulted in a significant reduction of the maximum intensity of stromal opacity in some experiments, whereas in other experiments the effect was not statistically significant. In vitro studies of the effect of the anti-inflammatory drugs on HSV-1 replication in Vero cells revealed that both dexamethasone and flurbiprofen inhibited HSV-1 replication in a dose-dependent manner. Flurbiprofen had a greater inhibitory effect, which appears to be due, at least in part, to a direct virucidal effect. Dexamethasone did not exhibit virucidal activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Characterization of arylamine acetyltransferase in the rabbit eye.

The activity of arylamine acetyltransferase with p-aminobenzoic acid (PABA), sulfamethazine (SMZ), and aminozolamide as substrates was studied in rabbit tissue homogenates of the corneal epithelium, stroma-endothelium, iris-ciliary process, and liver. Rabbits were classified as rapid or slow acetylators with respect to their rate of hepatic acetylation of SMZ. The ocular disposition of aminozolamide in the two phenotypes was compared using a topical ocular infusion method that permitted a constant concentration to remain in contact with the intact cornea. The effect of hepatic-acetylator phenotype on the intraocular pressure (IOP) recovery rate and drug concentrations in tissues after single-dose administration of aminozolamide also was studied. In general, the rank order of arylamine acetyltransferase activity regardless of substrate was liver greater than iris-ciliary process greater than corneal epithelium greater than stroma-endothelium. The specific activity with aminozolamide as substrate was greater than that with SMZ in each tissue homogenate and greater than with PABA as substrate in all tissues except the stroma-endothelium of slow hepatic-acetylator rabbits. Very low enzyme activity ratios for ocular acetylation between rapid and slow hepatic-acetylating rabbits indicated that acetylation in the ocular tissues did not correspond with the acetylation phenotype. At various times during and after topical infusion to the anesthetized rabbit, assay determinations of drug and metabolite in ocular tissues indicated that there were no significant differences between phenotypes in the disposition of either drug or metabolite. These results correlate with the IOP measurements after topical infusion; they also showed no difference in the effect of aminozolamide between hepatic-acetylator phenotypes. These results indicate that the ocular disposition and the decrease in IOP from topical application of aminozolamide is independent of the hepatic-acetylation phenotype in the rabbit. There are significant amounts of acetyltransferase activity in the ocular tissues of the rabbit with these three substrates, indicating that acetylation may be occurring for other arylamine drugs used in the eye.

Ocular disposition of aminozolamide in the rabbit eye.

We have previously determined that 6-amino-2-benzothiazolesulfonamide (aminozolamide) significantly lowers IOP in rabbits and, more importantly, in ocular hypertensive human subjects. Results from in vitro experiments established that both the inhibitory activity of aminozolamide against carbonic anhydrase B as well as the penetration rate across excised rabbit corneas were equivalent to ethoxzolamide. Consequently, we have investigated the ocular disposition of aminozolamide to explain its activity when instilled topically to the eye. A constant concentration of 67.4 micrograms/ml of drug was applied for 90 min to the left eye of anesthetized rabbits. Drug and metabolite were measured in both aqueous humor and iris/ciliary body over time. The metabolite was collected and purified. Identification using mass spectroscopy, high pressure liquid chromatography (HPLC) and fluorescence scans indicated that the metabolite was 6-acetamido-2-benzothiazolesulfonamide. Relatively high levels of metabolite were identified in the cornea and iris/ciliary body but were much lower in aqueous humor. Tissue concentrations over time for the metabolite in iris/ciliary body were approximately 2-fold higher than levels of metabolite measured in aqueous humor. When compared to drug levels measured in either aqueous humor or iris/ciliary body, metabolite levels in these respective tissues were much higher. It is hypothesized that topical activity is a consequence of both metabolite retention in the iris/ciliary body as well as inhibition of 99+% of carbonic anhydrase. Both of these events must occur over a sufficient time period to effect a significant lowering of IOP.

Furosemide (frusemide). A pharmacokinetic/pharmacodynamic review (Part II).

Part I of this article, which appeared in the previous issue of the Journal, covered the physical properties, pharmacology, toxicology and pharmacokinetics of furosemide (frusemide). In part II the authors examine the pharmacodynamics of the drug, and suggest various areas for future study.

Furosemide (frusemide). A pharmacokinetic/pharmacodynamic review (Part I).

Furosemide (frusemide) is a potent loop diuretic used in the treatment of oedematous states associated with cardiac, renal and hepatic failure, and for the treatment of hypertension. Therapy is frequently complicated by apparently erratic systemic availability from the oral route and from unpredictable responses to a given dosage. The exact mechanism of action is not fully understood, but furosemide is believed to act at the luminal surface of the ascending limb of the loop of Henle by inhibiting the active reabsorption of chloride. The response to a given dosage is modulated by the fluid and electrolyte balance of the individual. Acute and delayed tolerance has been demonstrated both in animals and in man, and is postulated to be due to the intervention of homeostatic mechanisms influencing fluid and electrolyte balances. Furosemide is delivered to its site of action by active secretion via the nonspecific organic acid pump. Comparisons between the observed diuresis/saluresis and plasma furosemide concentrations, urinary excretion rates and renal clearance found either negative or no correlations with plasma drug concentration but significant correlations with urine measurements. Response is related to the concentration of the drug in urine rather than in plasma. The most common adverse reactions attributable to furosemide therapy are essentially extensions of the therapeutic effects (i.e. fluid and electrolyte disturbances). The pharmacokinetic behaviour of furosemide is marked by a large degree of variability, derived from differences within and between both subjects and study protocols. Part of this variability can be attributed to differences in organ function, which is important in view of the types of patients treated with furosemide. On the other hand, a large proportion remains as inter- and intrasubject variation. The bioavailability of furosemide from oral dosage forms is highly variable. The poor bioavailability has been hypothesized to be due to the poor solubility of the compound, site-specific absorption, presystemic metabolism and/or other unknown mechanisms. Furosemide is highly bound to plasma proteins, almost exclusively to albumin. Although the drug is insoluble in water and favours partitioning into fatty tissue, the high degree of plasma protein binding restricts the apparent volume of distribution at steady-state to values within a multiple of 2 to 5 times the plasma volume. Furosemide has two documented metabolites--furosemide glucuronide and saluamine (CSA). The first is an accepted metabolic product, whereas the status of CSA as a metabolite is highly controversial.(ABSTRACT TRUNCATED AT 400 WORDS)

Aminozolamide gel. A trial of a topical carbonic anhydrase inhibitor in ocular hypertension.

A topical carbonic anhydrase inhibitor, 6-amino-2-benzothiazolesulfonamide (aminozolamide), which is an analogue of ethoxzolamide, was studied in 18 patients with ocular hypertension. Significant lowering of intraocular pressure was achieved with a single 50-microL gel application compared with the untreated control eye. The onset of action occurred within two hours and lasted at least eight hours. No systemic side effects were elicited. Topical carbonic anhydrase inhibitors may be an important alternative in the medical management of glaucoma.

Reduction of phenylephrine drop size in infants achieves equal dilation with decreased systemic absorption.

We studied the effect of reducing eye drop size on the efficacy and systemic absorption of topical 2.5% phenylephrine hydrochloride in neonates and infants. Eleven neonates received an 8-microL drop volume in one eye and a 30-microL drop volume (commercial size) in the fellow eye. Mean pupillary dilation at 60 minutes was equivalent (4.86 mm vs 4.57 mm) for both eyes, respectively. The plasma phenylephrine level was determined for the two drop sizes in a second group of infants. Eight infants received an 8-microL drop volume in both eyes, while nine infants received a 30-microL drop volume in both eyes. The mean phenylephrine level at ten minutes was 0.9 ng/mL for the 8-microL drop group and 1.9 ng/mL for the 30 microL drop group. In neonates and infants, reducing the drop volume of topical phenylephrine may improve the risk-benefit ratio.

Ethoxzolamide analogue gel. A topical carbonic anhydrase inhibitor.

An analogue of ethoxzolamide, 6-hydroxyethoxzolamide, was synthesized to enhance corneal permeability yet retain carbonic anhydrase inhibitory activity for use in lowering intraocular pressure. In a 1% suspension, the analogue caused a small but statistically significant unilateral reduction of IOP when applied to one eye of normal rabbits. When formulated in a gel vehicle, 6-hydroxyethoxzolamide caused a more prolonged and larger reduction in IOP in normal and ocular hypertensive rabbits compared with its effect in suspension or with the parent compound.

Aqueous vs viscous phenylephrine. I. Systemic absorption and cardiovascular effects.

We studied 30 patients undergoing vitreoretinal surgery to compare the systemic absorption and cardiovascular effects of 2.5% aqueous and 2.5% viscous (21 cp) ophthalmic solutions of phenylephrine hydrochloride. No significant differences were noted in the plasma levels or changes in blood pressure between the two groups, although there was a tendency toward higher mean plasma levels and blood pressures in groups receiving 2.5% aqueous phenylephrine hydrochloride. Maximum plasma levels were achieved within the first 20 minutes following topical application of phenylephrine eye drops, irrespective of the nature of the vehicle. This finding emphasizes the importance of monitoring these patients, especially those at high risk for any adverse cardiovascular effects during the first 20 to 30 minutes following instillation of phenylephrine eye drops. The patients in our study were supine and under general anesthesia. Therefore, there was no effect by lid blinking, the lacrimal pump, or gravity, which would ordinarily increase absorption by the nasal mucosa through the nasolacrimal system. By eliminating these variable factors, such as lid blinking, the study was performed in a stable and controlled manner, but the results may not be directly applicable to an upright awake patient.

Aqueous vs viscous phenylephrine. II. Mydriatic effects.

We performed four studies to determine whether there is a difference in the mydriatic effect of 2.5% aqueous vs 2.5% viscous phenylephrine hydrochloride solutions. The first study was performed under "room light" conditions, and the mean (+/- SD) dilation at one hour was 0.87 +/- 1.18 mm for the aqueous and 0.86 +/- 1.14 mm for the viscous solutions. The second study was performed in a darkened room, and the mean dilation at one hour was slightly greater than in room light but was still minimal (aqueous, 1.14 +/- 1.00 mm; viscous, 1.07 +/- 1.11 mm). In the third study, patients were pretreated with a topical anesthetic (0.5% proparacaine hydrochloride), and the mean one-hour dilation was approximately twice (aqueous, 2.30 +/- 0.81 mm; viscous, 2.41 +/- 0.88 mm) that found in patients who were not pretreated with proparacaine. In the fourth study, the two phenylephrine solutions were used in combination with 1% tropicamide, and the mean one-hour dilation was 3.8 +/- 0.82 mm for the aqueous and 3.8 +/- 0.98 mm for the viscous solutions. Our studies show that there is no difference in the mydriatic effect of 2.5% aqueous vs 2.5% viscous phenylephrine solutions when used alone or in combination with 0.5% proparacaine or 1% tropicamide.

Systemic absorption and cardiovascular effects of phenylephrine eyedrops.

We studied 24 patients undergoing vitreoretinal surgery to compare the systemic absorption and cardiovascular effects of 2.5% aqueous and 10% viscous ophthalmic solutions of phenylephrine hydrochloride. Plasma levels measured in patients receiving two drops of 10% viscous solution were consistently higher ten, 20, and 60 minutes after instillation (P less than .02). Although the mean systolic and diastolic blood pressure was generally higher with the 10% viscous solution, the difference was not statistically significant. Blood pressure was high in several isolated cases. Because maximum plasma levels are achieved within ten to 20 minutes after topical instillation, phenylephrine eyedrops should be administered under close observation so that if an adverse reaction occurs it can be readily treated.

The effect of reduced eyedrop size and eyelid closure on the therapeutic index of phenylephrine.

In this study we examined the relative effects of reducing eyedrop size (from 30 microliters to 10 microliters) and eyelid closure on the ocular efficacy and systemic absorption of 10% phenylephrine. Thirteen subjects participated in a quadruple crossover study that involved dilation with a 10-microliters and a 30-microliters drop of phenylephrine with and without eyelid closure. The 10-microliters drop was just as effective for pupillary dilation as the 30-microliters drop. Eyelid closure improved dilation for both drop sizes. Both eyelid closure and reducing the drug volume decreased systemic absorption of phenylephrine as measured by plasma concentration. When used together, eyelid closure and the smaller drop size reduced plasma concentration by 45%. The therapeutic index for 10% phenylephrine appears to be improved by using a 10-microliters drop followed by eyelid closure.

The effects of sigma ligands on protein release from lacrimal acinar cells: a potential agonist/antagonist assay.

Sigma receptor antagonists have been proposed as leading clinical candidates for use in various psychotic disorders. Prior to clinical testing, it is imperative that a new agent be correctly identified as an antagonist and not an agonist since the latter may worsen the psychosis. For sigma-ligands many behavioral and pharmacological assays have been developed in an attempt to classify agonist/antagonist activity. These assays evaluate a response or a behavior in an animal model that can be related to clinical efficacy. However, is the action by the presumed antagonist a consequence of sigma-receptor activity? Previously we have identified sigma-receptors in acinar cells of the main lacrimal gland of the New Zealand white rabbit and have measured protein release after the addition of various N,N-disubstituted phenylalkylamine derivatives known to be sigma-ligands by receptor binding studies. Although protein release from acinar cells has been attributed to either muscarinic or alpha-adrenergic stimulation, protein release from sigma-receptor stimulation was also confirmed. In the reported studies here, we isolated and incubated acinar cells with varying concentrations of known sigma-ligands and measured protein concentration. A knowledge of the receptor profile for the disubstituted phenylalkylamines permitted experiments to be designed in which various alpha, muscarinic, serotonergic, and dopaminergic antagonists could be added in equimolar concentrations. Under the conditions of these experiments, statistically significant increases in protein release for sigma-ligands could be attributed to stimulation of sigma-receptors. Haloperidol, an apparent sigma-antagonist, caused a statistically significant decrease in protein release and also inhibited protein release when tested with a known sigma-ligand, AF2975 [N,N-dimethyl-2-phenylethylamine]. In this system, stimulation and inhibition of protein release were defined as agonist and antagonist behavior, respectively. Of particular interest were the results for BMY14802 and +/- pentazocine, both of which were found to be agonists. Various antipsychotic and antidepressant drugs were measured for their agonist/antagonist behavior. Because of multireceptors present in acini, their agonist or antagonist behaviour could not be attributed solely to interaction with the sigma-receptor unless specific antagonists were added.

Significance of melanin binding and metabolism in the activity of 5-acetoxyacetylimino-4-methyl-delta2-1,3,4,-thiadiazolin e-2-sulfonamide.

5-Acetoxyacetylimino-4-methyl-delta2-1,3,4,-thiadiazoline -2-sulfonamide (compound (1)) is an ester prodrug that lowered intraocular pressure (IOP) in albino New Zealand rabbits, but was found to be inactive in pigmented Dutch Belt rabbits. In order to explain the differences in pharmacological activity for the two rabbit species, metabolism and melanin binding were studied. Depending on the initial concentration, the binding of compound (1) to natural melanin (Sepia officinalis) was 20-60%. The binding constant, K, at 37 degrees C was 4.32 x 10(5) M(-1) and the maximum moles bound to melanin, r(max), was 4.5 x 10(-7) mol/mg of melanin. From a determination of binding at temperatures between 25 degrees C and 47 degrees C, a van't Hoff plot was constructed to determine enthalpy and entropy changes accompanying the binding process, deltaH and deltaS, respectively. Values calculated from the plot were -12.7 and -15.4 kcal/(mol deg), respectively. Negative values for these parameters are consistent with charge transfer interactions and therefore suggest that this may be an operative mechanism between compound (1) and melanin. The in vitro incubation of compound (1) was also studied with various ocular tissues from both albino and pigmented rabbits which were iris-ciliary body, intact cornea, stroma/endothelium and aqueous humor. A major metabolite, MET 1, was identified and also observed from in vivo analyses of the same tissues following topical application. The metabolite was isolated and subjected to mass spectroscopy and proton nuclear magnetic resonance spectroscopy analysis. From these analyses, it was hypothesized that the formation of MET 1 involved a GSH conjugation mechanism which displaced the sufonamide (-SO2NH2) group. The metabolism was found to be less extensive in the pigmented rabbit than in the albino rabbit and suggested that the binding affinity of compound (1) for melanin was a better explanation for the lack of IOP activity in the pigmented rabbit than differences in metabolism.

Relationship between steroid permeability across excised rabbit cornea and octanol-water partition coefficients.

Permeability rates were determined across excised rabbit corneas for 11 steroids. Permeability coefficients for each steroid were calculated, and their logarithms were plotted against their respective log octanol-water partition coefficients. A parabolic relationship resulted, with an optimum log permeability and coefficient observed at a log Po of 2.9. From these experimental results, an improvement in ophthalmic bioavailability of dexamethasone acetate as compared to dexamethasone is predicted and correlates with literature results.

Effect of particle size on ophthalmic bioavailability of dexamethasone suspensions in rabbits.

Three suspensions of 0.1% [3H]dexamethasone were prepared with mean particle sizes of 5.75, 11.5, and 22.0 micron. The suspensions were dosed topically to the right eyes of rabbits. Their bioavailability was compared by measuring aqueous humor and corneal levels over 5 hr. A statistically significant rank-order correlation was observed between increasing drug levels and decreasing particle size.