Investigation of human pharmacoscintigraphic behavior of two tablets and a capsule formulation of a high dose, poorly water soluble/highly permeable drug (efavirenz).

Human pharmacoscintigraphic behavior of two tablets and a capsule formulation of a high dose, poorly water soluble, highly permeable, micronized drug (efavirenz) was investigated. The tablets and capsule, prepared with samarium oxide and neutron activated to produce radioactive samarium-153, were evaluated for their in vivo disintegration and gastrointestinal (GI) transit in healthy subjects under fasted condition. Scintigraphic images were acquired to coincide with blood sampling times to assess the plasma concentration-time profile in relation to in vivo disintegration and GI transit. The mean gastric emptying times were approximately the same for all three formulations. Although in vivo dosage form disintegration was faster for Tablet A as compared to Tablet B and was similar between Tablet A and the capsule, Tablet A showed a slower rate and extent of drug absorption than Tablet B and the capsule. The results of this study eliminated the initial hypothesis that the difference in in vivo performance between the two tablet formulations is due to a different rate of in vivo disintegration and suggest that for this drug the in vivo dissolution rate of the drug from its disintegrated dosage form was a more important factor affecting the rate and extent of drug absorption.

Synthetic macromolecular inhibitors of human leukocyte elastase. 1. Synthesis of peptidyl carbamates bound to water-soluble polymers: poly-alpha,beta-[N-(2-hydroxyethyl)-D,L-aspartamide] and poly-alpha-[N5-(2-hydroxyethyl)-L-glutamine].

The design and synthesis of macromolecular peptidyl carbamate inhibitors of human leukocyte elastase (HLE), based on coupling of a low-molecular-weight peptidyl carbamate, succinylalanylalanylprolylmethyl isopropyl carbamate, with a linear hydrophilic polymer, poly-alpha,beta-[N-(2-hydroxyethyl)-D,L-aspartamide] or poly-alpha-[N5-(2-hydroxyethyl)-L-glutamine], is described. The covalent linkage between a flexible linear polymer and a peptidyl carbamate inhibitor of HLE did not compromise in vitro inhibitory capacity. The macromolecular peptidyl carbamates reported here represent a novel class of elastase inhibitors with a Ki ranging from 35.5 to 2.0 nM.

Preparation of [18-F]haloperidol.

A procedure is described that permits the preparation of [18F]haloperidol in 140 min at specific activities ranging from 4-5 muCi mg-1. A key step in the synthetic route involves the incorporation of 18F into the molecule through a Schiemann-type reaction, which involves the pyrolysis of the diazonium tetrafluoroborate salt of 4-[4-(p-chlorophenyl)-4-hydroxypiperidino]-4'-aminobutyrophenone.

Synthesis of [82Br]bromperidol and preliminary tissue distribution studies in the rat.

A procedure is described for the preparation of [82Br]bromperidol with specific activity 440 muCi/mg. The incorporation of bromine-82 into the molecule was accomplished through Brackman and Smit's modification of the Sandmeyer reaction, during the last step of the synthetic route. This involved the formation of a complex between Cu82Br2 and nitric oxide gas in acetonitrile, which was then allowed to react with 4-[4-(aminophenyl)-4-hydroxy-piperidinyl]-1-(4-fluorophenyl)-1-butanone (aminoperidol, 10) to give [82Br]bromperidol in about 1.5 h. Cupric 82Br]bromide was prepared in situ from K82Br and CuSO4.5H2O. The radiochemical and chemical yields for the preparation of [82Br]bromperidol from K82Br were 10.4 and 12%, respectively. Preliminary tissue distribution studies with the labeled bromperidol in the rat showed that the uptake of radioactivity by the liver, brain, kidneys, and the lungs was very fast and was in the declining phase in the latter organs 15 min after iv administration.

Peptidyl carbamates incorporating amino acid isosteres as novel elastase inhibitors.

The design and synthesis of 13 novel peptidyl carbamates are described. When tested for inhibitory activity toward porcine pancreatic elastase, trypsin, and chymotrypsin, six compounds were found to specifically inhibit elastase without affecting the other two serine proteases. All the active inhibitors had an amino acid isostere at the P1 position. Kinetic studies indicated that the inhibition was competitive with Ki values ranging from 4.23 X 10(-5) to 2.4 X 10(-6) M. The degree of inhibition was found to be dependent on the specificity of the peptide chain for the extended subsites on the enzyme as well as on the nature of P1'. Preliminary work on one inhibitor indicates that the inhibition is reversible and proceeds via the rapid formation of a strong enzyme-inhibitor complex, followed by slow acylation of the serine residue on the active site of the enzyme. Peptidyl carbamates represent a novel class of elastase inhibitors.

Carbon-13 nuclear magnetic resonance investigations into the interactions of bisulfite with pyrimidine nucleosides and nucleotides.

Carbon-13 NMR is utilized to demonstrate the attack of bisulfite anion on uridine, 5-fluorouridine, and uridine 5'-monophosphate. The attack produces a pair of diastereomeric adducts similar in structure to those seen in the uracil series. Intensity data from the equilibrium system give an estimate for the individual equilibrium constants. Thymidine and thymidine 5'-monophosphate show no evidence of nucleophilic attack by bisulfite. This evidence indicates that bisulfite addition to nucleosides and nucleotides models the enzymatic methylation of uridine by the enzyme thymidylate synthetase better than the uracil bisulfite system.

Preparation of nitroso-13N-labeled nitrosoureas.

A method is described for the preparation of 13N-labeled N-nitrosoureas, specifically 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea. The 13N is generated as ammonia by the 12C(d,n)13N reaction on methane gas. The product is selectively trapped and subsequently oxidized to nitrous acid which reacts with the parent urea in solution to form the 13N-labeled nitrosourea.

Radionuclide gastric emptying studies in patients with anorexia nervosa.

To evaluate gastric emptying in anorexia nervosa patients, 26 patients (17 females, two males, ranging in age from 13 to 40 yr) with upper GI symptoms ingested 150-200 microCi [99mTc]triethelenetetraamine polysterene resin in cereal and were imaged in the supine position. Data were accumulated at 5 min intervals to obtain the gastric emptying time (GET). The results of the studies were divided into three categories: prolonged, 13 patients; rapid, 11; and normal 3. Twelve of 13 patients with prolonged GET were given 10 mg metoclopramide i.v. injections; nine of the 12 patients had a good response and three had no response. Five of the nine patients underwent metoclopramide therapy and four of the patients showed benefit from the therapy. One patient discontinued metoclopramide therapy because of somnolence. Although all patients had subjective symptoms of gastric dysfunction, our results indicated only 50% had objectively prolonged GET, and another 50% showed normal or even rapid GET. Therefore, this radionuclide study enables quantitatively objective documentation of gastric emptying, separation of those patients with rapid or normal GET from those with prolonged GET, thereby avoiding the possible side effects from metoclopramide medication, and prediction of effectiveness of metoclopramide therapy in patients with prolonged GET.

Gastrointestinal behavior of orally administered radiolabeled erythromycin pellets in man as determined by gamma scintigraphy.

The behavior of single 250-mg doses of a multiparticulate form of erythromycin base (ERYC(R)), each including five pellets radiolabeled with neutron-activated samarium-153, was observed by gamma scintigraphy in seven male subjects under fasting and nonfasting conditions. The residence time and locus of radiolabeled pellets within regions of the gastrointestinal tract were determined and were correlated with plasma concentrations of erythromycin at coincident time points. Administration of food 30 minutes postdosing reduced fasting plasma erythromycin Cmax and area under the plasma erythromycin versus time curve (AUC) values by 43% and 54%, respectively. Mean peak plasma concentration of erythromycin (Cmax) in the fasting state was 1.64 micrograms/mL versus 0.94 micrograms/mL in the nonfasting state. Total oral bioavailability, as determined by mean AUC (0-infinity) of the plasma erythromycin concentration versus time curve, was 7.6 hr/micrograms/mL in the fasted state, versus 3.5 hr/micrograms/mL in the nonfasting state. Mean time to peak plasma erythromycin concentration (tmax) in the fasting state was 3.3 hours, versus 2.3 hours in the nonfasting state. Plasma concentrations of erythromycin in both fasting and nonfasting states were within acceptable therapeutic ranges. Evidence provided by this study: 1) indicates that pellet erosion and absorption of active erythromycin base begins when the enteric-coated pellets reach the highly vascular mucosa of the jejunum and proximal ileum, and is essentially completed within the ileum, with a significant portion absorbed in the medial-to-distal ileum; 2) confirms that acceptable therapeutic plasma levels of erythromycin are attained in nonfasting subjects (Cmax = 0.94 microgram/mL) and that superior plasma erythromycin concentrations (Cmax = 1.64 micrograms/mL) are achieved by administration of the dose on an empty stomach 1 to 2 hours before or after meals; 3) corroborates other comparative studies reporting greater fasting bioavailability with this multiparticulate dosage form of erythromycin base than with reference single tablet or particle-in-tablet formulations; and 4) indicates that neutron activation of stable isotopes incorporated as a normal excipient in industrially-produced formulations provides an effective means for in vivo evaluation of dosage forms through gamma scintigraphy.

Explaining variable absorption of a hypolipidemic agent (CGP 43371) in healthy subjects by gamma scintigraphy and pharmacokinetics.

The gastrointestinal absorption of a hypolipidemic agent (CGP 43371) was investigated using an external scintigraphy technique in six healthy men. After an overnight fast, subjects received a single 800-mg oral dose of CGP 43371 (4 capsules of 200 mg each) and one capsule of radioactive samarium-153 oxide (100-130 microCi) as a nonabsorbable marker of gastrointestinal transit and fecal recovery for CGP 43371. In vivo gastrointestinal transit of samarium-153 was monitored via gamma scintigraphy for 48 hours after administration to coincide with blood sampling. Samarium-153 content in whole fecal samples was determined by external gamma scintigraphy, and CGP 43371 content in both fecal and plasma samples was determined using high-performance liquid chromatography (HPLC). The results of fecal analysis indicated that transit of the two compounds in the gastrointestinal tract were similar, and bioavailability of CGP 43371 was calculated to be 9% based on the difference between the cumulative amounts of the nonabsorbable radioactive marker and CGP 43371 found in the feces. The onset of drug absorption occurred 4 hours after administration when radioactive samarium-153 was in the distal small bowel, and peak plasma drug level occurred 6 hours after administration, which corresponded with the arrival of samarium-153 in the terminal ileum and ileal/cecal junction. This observation supported the concept that primary absorption of this compound was in the distal to terminal portion of the ileum. Although the onset of drug absorption was delayed, it was curious that the rate of gastric emptying also affected the extent of absorption. A positive correlation (r = 0.91) between area under the drug curve (AUC) and area under the transit curve (AUTC) of the gastric emptying showed that longer gastric residence improved oral absorption of CGP 43371.

Distribution of 13N in rat tissues following intravenous administration of nitroso-labeled BCNU.

The concentrations of label in 16 major organs and tissues of pentobarbital-sedated normal male rats were measured at six time points ranging from 0.2 to 50 min after IV injection of the antitumor drug BCNU labeled in the nitroso position with cyclotron-produced nitrogen-13. Initial (12-s) concentrations in the lungs, kidneys, and heart were 41, 13, and 11 times the whole-body average, respectively. Time for clearance of the first 50% of the injected dose from the circulation was of the order of several seconds. Estimated first-pass extractions of 70% or more were noted in the heart, kidneys, brain, stomach, small intestine, muscle, fat, and bone. Washout of label from the heart and lungs was quite rapid, removing most of the initially extracted 13N from these organs by 2 min after injection. Label concentrations in the kidneys exceeded those in all other tissues studied between 2 and 50 min. Secondary accumulations of 13N were observed in muscle, skin, liver, small intestine, and fat. Label concentrations in a number of tissues closely paralleled the steadily decreasing concentration in blood for various intervals between 5 and 50 min. The results suggest that the toxic insult to lung tissue from IV administered BCNU is effected in a period of several minutes. They also suggest that intra-arterial administration of the drug would significantly raise the target/non-target dose ratio and lower the incidence of pulmonary toxicity.

Assessment of new formulations of nonoxynol-9 coprecipitated with polyvinylpyrrolidone and iodine as possible vaginal contraceptives.

OBJECTIVE: To assess the in vitro spermicidal activity of new formulations of nonoxynol-9, coprecipitated with polyvinylpyrrolidone (PVP) or iodinated PVP, against human spermatozoa via the use of the Sander-Cramer test and the cervical mucus penetration test. DESIGN: Solutions of PVP-nonoxynol-9 and iodinated PVP-nonoxynol-9 containing nonoxynol-9 whole molecule (oligomers 1 to 18) and its isolated fractions (oligomers 8 to 10, 4 to 6, and 1 to 3) at various concentrations (microgram/mL) were prepared via serial dilutions. Spermicidal solutions were mixed with human semen to determine the minimal lethal dose (microgram/mL). In the Sander-Cramer test, the lethal dose was reported as the minimal dose capable of killing spermatozoa within 20 seconds. In the cervical mucus penetration test, the lethal dose was reported as the minimal dose capable of preventing penetration of spermatozoa into cervical mucus beyond the second millimeter length of the capillary. SETTING: Andrology laboratory, University of Kentucky, Lexington, Kentucky. PATIENT(S): Normospermic male donors. MAIN OUTCOME MEASURE(S): Spermicidal lethal dose determination of various nonoxynol-9 preparations containing the whole nonoxynol-9 molecule and its isolated fractions coprecipitated with PVP or iodinated PVP. RESULT(S): The use of PVP increased the aqueous solubility of the nonoxynol-9 formulations containing oligomers 1 to 18 and 8 to 10 slightly. The coprecipitation of the nonoxynol-9 formulations containing nonoxynol-9 oligomers 4 to 6 and 1 to 3 with PVP significantly increased their solubilization and spermicidal action in vitro. Moreover, the incorporation of iodine significantly decreased the minimal nonoxynol-9 dose required for complete killing of spermatozoa in preparations containing nonoxynol-9 oligomers 4 to 6 and 1 to 3. CONCLUSION(S): Incorporation of all three components tested in this study (PVP, nonoxynol-9, and iodine) enhanced the efficiency of the spermicidal preparations, especially for nonoxynol-9 preparations containing nonoxynol-9 oligomers 4 to 6 and 1 to 3.

Studies with [11C]alprazolam: an agonist for the benzodiazepine receptor.

We have built a system for the synthesis of high specific activity carbon-11 alprazolam (Xanax), a high affinity agonist for the benzodiazepine receptor. The system produces 30-40 mCi of the compound with a specific activity of > 12,000 Ci per millimole. Using this compound we have performed PET studies on 6 normal subjects and studied the cerebral influx and efflux of the compound. The uptake in the brain was low, approx. 1% of the administered dose. However, the levels of the compound in the circulation at early time points are heavily affected by the specific activity of the tracer, i.e. when pharmacologically active doses are used as blocking doses the concentration of radioactive material is higher in the circulation and more material enters the brain. We attribute this to a depot effect where the compound is trapped in saturatable sites in an organ, probably the lungs, and is slowly released over time. In the presence of blocking doses of agonist, the compound washes out of the brain more quickly suggesting that some blockade of the receptors is occurring. However, the pharmacological activity of the compound does not permit the administration of enough material to ensure complete receptor blockade. The compound shows definite signs of acting as a receptor binding ligand but the unusual pharmacokinetics complicate the interpretation of the data.

Detection of tetrafluoroputrescine in RBCs by fluorine-19 nuclear magnetic resonance spectroscopy.

Fluorine-19 nuclear magnetic resonance spectroscopy is used to detect the in vitro uptake of tetrafluoroputrescine, a novel putrescine analogue, in red blood cells.

Tumor selective enhancement of radioactivity uptake in mice treated with alpha-difluoromethylornithine prior to administration of 14C-putrescine.

A group of EMT6 tumor bearing male BALB/c mice which had been treated with alpha-difluoromethylornithine (DFMO, a specific, irreversible inhibitor of ornithine decarboxylase, the enzyme which catalyzes the biosynthesis of putrescine), 8 mg/mouse, ip, 20 and 5 hrs before the 14C-putrescine dose, and a group of control animals were administered 14C-putrescine (0.5 muCi, 0.1 mCi/mmol, iv) 60 min prior to sacrifice. Radioactivity uptake data was obtained for the tumor and 13 major normal organs. In the control animals the tumor exhibited one of the highest uptakes of radioactivity. For DFMO-pretreated mice the radioactivity distribution among most of the normal tissues was not very different from that obtained for the control animals. However, the uptake into the tumor was enhanced by a factor of approximately 4. So, high tumor-to-tissue ratios (3.8, lung to 38, brain) were attained as a result of DFMO treatment.

Enhanced entrapment of a quaternary ammonium compound in liposomes by ion-pairing.

The encapsulation of a quaternary ammonium compound by multilamellar liposomes was enhanced by formation of ion-pairs with a counterion. Thus, [14C]methantheline bromide was synthesized and paired with a 25 M excess of trichloroacetate. Under these conditions, the amount of radioactivity entrapped by phosphatidylcholine liposomes was three times greater than when no trichloroacetate was present. The increased liposomal loading was probably due to the solubilization of the ion-pair in the lipid membrane of the liposome.

Noninvasive dissolution measurement using perturbed angular correlation.

A novel noninvasive technique was developed to measure dissolution of the water-soluble component of a solid dosage form using indium 111 and perturbed angular correlation. The method involves time-delayed coincidence counting of two cascading gamma-rays that exhibit angular correlation. This angular correlation can be perturbed if the intermediate excited state of the nucleus is reoriented due to an interaction with its environment. When such an interaction occurs, as in a phase change (solid to liquid), the perturbation changes can be shown by anisotropy. A highly perturbed condition in the solid state results in low values (0.02-0.04), while increasing values of anisotropy indicate dissolution. Anisotropy values reach 0.14-0.15 when the total unperturbed physical state (liquid) exists. The worth of this technique was demonstrated by both in vitro and in vivo determinations of dissolution rates.

Metabolism and distribution of 1-[14C]alprazolam in rats.

Using 14C-labeled alprazolam (1-[14C]APZ) as a model for 1-[11C]APZ, we evaluated the tissue distribution of total radioactivity and assessed the contribution of the polar metabolites of APZ (alpha-hydroxymethyl- and 4-hydroxy-APZ) to radioactivity levels in the plasma and brains of rats over the course of 1 h. The biodistribution data showed that the uptake of radioactivity by rat brain was 0.31% of the injected dose per gram of tissue weight at 10 min postinjection. Pretreating rats reduced the levels in brain to 0.21% of the injected dose at 10 min but had little effect on the distribution of radioactivity in plasma and other tissues studied. Analysis of the metabolites in plasma and brain homogenates by an extraction-thin-layer chromatography-liquid scintillation method revealed that greater than 94% of the radioactivity in the rat brain was due to 1-[14C]APZ over the course of 1 h. APZ, therefore, is stable to metabolic transformations in the rat brain, and the polar metabolites are readily conjugated and excreted so that their cerebral uptake is minimal.

Assessment of antiprotease activity of some carbamate derivatives.

Six carbamate derivatives were tested for inhibitory activity towards porcine pancreatic elastase, trypsin, and chymotrypsin. Only three esters that are isatoic anhydride derivatives were found to inhibit elastase competitively but nonspecifically. KI values for the best two inhibitors were determined from Dixon plots.

Cross-linking of gelatin capsules and its relevance to their in vitro-in vivo performance.

The present review deals with the chemistry of gelatin cross-linking under conditions that are relevant to pharmaceutical situations. Mechanistic rationalizations are offered to explain gelatin cross-linking under "stress" conditions. These include elevated temperature and high humidity conditions. In addition, the chemical interactions between gelatin and aldehydes, such as formaldehyde and other formulation excipients, are discussed. The literature on the in vitro and in vivo dissolution and bioavailability of a drug from stressed gelatin capsules and gelatin-coated tablets is reviewed. Cross-linking phenomena, occurring in stressed hard gelatin capsules and gelatin-coated tablets, could cause considerable changes in the in vitro dissolution profiles of drugs. However, in a few cases, the bioavailability of the drug from the stressed capsules is not significantly altered when compared to that obtained from freshly packed capsules. It is concluded that, as with other drug-delivery systems, careful attention should be paid to the purity and chemical reactivity of all excipients that are to be encapsulated in a gelatin shell. It is suggested that in vitro dissolution tests of hard gelatin-containing dosage forms be conducted in two stages, one in a dissolution medium without enzymes and secondly in dissolution media containing enzymes (pepsin at pH 1.2 or pancreatin at pH 7.2, representing gastric and intestinal media, respectively) prior to in vivo evaluation. Such in vitro tests may constitute a better indication of the in vivo behavior of gelatin-encapsulated formulations. Furthermore, testing for contamination with formaldehyde as well as low molecular weight aldehydes should be a standard part of excipient evaluation procedures.