Recommended Best Practices in Freeze Dryer Equipment Performance Qualification: 2022.

Best practices for performing freeze dryer equipment qualification are recommended, focusing on identifying methods to quantify shelf thermal uniformity (also known as "shelf surface uniformity"), equipment capability, and performance metrics of the freeze dryer essential to the pharmaceutical Quality by Design paradigm. Specific guidelines for performing shelf temperature mapping, freeze dryer equipment limit testing (the capability curve), and condenser performance metrics have been provided. Concerning shelf temperature mapping and equipment capability measurements, the importance of paying attention to the test setup and the use of appropriate testing tools are stressed. In all the guidelines provided, much attention has been paid to identifying the balance between obtaining useful process knowledge, logistical challenges associated with testing in the production environment vs that at laboratory scale, and the frequency of the testing necessary to obtain such useful information. Furthermore, merits and demerits of thermal conditions maintained on the cooled surfaces of the freeze dryer condenser have been discussed identifying the specific influence of the condenser surface temperature on the process conditions using experimental data to support the guidelines. Finally, guidelines for systematic leak rate testing criteria for a freeze dryer are presented. These specific procedural recommendations are based on calculations, measurements, and experience to provide useful process and equipment knowledge.

Recommended Best Practices for Process Monitoring Instrumentation in Pharmaceutical Freeze Drying-2017.

Recommended best practices in monitoring of product status during pharmaceutical freeze drying are presented, focusing on methods that apply to both laboratory and production scale. With respect to product temperature measurement, sources of uncertainty associated with any type of measurement probe are discussed, as well as important differences between the two most common types of temperature-measuring instruments-thermocouples and resistance temperature detectors (RTD). Two types of pressure transducers are discussed-thermal conductivity-type gauges and capacitance manometers, with the Pirani gauge being the thermal conductivity-type gauge of choice. It is recommended that both types of pressure gauge be used on both the product chamber and the condenser for freeze dryers with an external condenser, and the reasoning for this recommendation is discussed. Developing technology for process monitoring worthy of further investigation is also briefly reviewed, including wireless product temperature monitoring, tunable diode laser absorption spectroscopy at manufacturing scale, heat flux measurement, and mass spectrometry as process monitoring tools.

A comparative study of freeze-drying heat transfer in polymeric vials and glass vials.

Implementation of polymeric vials for freeze-dried drug products has been practically non-existent because of unique moisture barrier and thermodynamic technical challenges. Hybrid vials, which combine the benefits of polymer and glass, have been shown to address the challenges of ordinary polymeric vials. Tackling thermodynamic challenges starts with a clear understanding of the heat transfer mechanism. To this end, multi-physics simulations and experimentation were used to compare the heat transfer between hybrid cyclic olefin polymer (COP) vials and borosilicate glass vials during freeze-drying. Parametric models were developed for hybrid COP and glass vials to systematically study the effect of five design parameters based on the arrangement of the vials on a tray inside a lyophilization chamber. Heat transfer in glass vials were dominated by heat conduction with the surrounding vapor, while hybrid COP vials were governed by conduction with the bottom shelf. Furthermore, hybrid COP vials exhibited more consistent heat flow rate and total heat transfer coefficient compared to glass vials, suggesting higher product quality as a result. The distance between adjacent vials and the drug product height were the most important parameters affecting heat transfer irrespective of vial type. Results indicated that hybrid COP vials can be filled to higher fill volumes with higher heat transfer and without the risk of breakage. Results of this study can help design innovative primary packaging systems for freeze drying or optimizing heat transfer for existing glass or hybrid COP vial systems regarding product consistency and drying time.

Pharmacodynamic effects of cotinine in abstinent cigarette smokers.

The nicotine metabolite cotinine was administered to abstinent cigarette smokers to determine whether it has pharmacologic activity as assessed by various physiologic and subjective measurements. By means of a randomized, double-blind, placebo-controlled counterbalanced-order design, subjects received cotinine base (30 mg) intravenously after 48 hours of abstinence from cigarette smoking. Serum cotinine concentrations increased to levels commonly achieved during daily cigarette smoking, whereas no change in serum nicotine concentration was observed. Cotinine compared with placebo produced subjective differences in self-reported ratings of restlessness, anxiety and tension, insomnia, sedation, and pleasantness. Cotinine had minimal effects on cardiovascular measurements. These findings indicate that cotinine is behaviorally active in the setting of cigarette abstinence at blood concentrations similar to those commonly achieved through daily cigarette smoking.

Early ADME in support of drug discovery: the role of metabolic stability studies.

The area of Drug Discovery has undergone an amazing evolution in the past decade. This evolution is typified by the development of automated combinatorial synthesis and high throughput pharmacological testing. This, in turn, has lead to the ability to create and mine extensive databases and then model this new information. The overall result is a substantial increase in the rate of target identification, generation of new leads, and finally, optimization of those leads into clinical candidates. ADME studies have always played a critical role in helping to optimize the pharmacokinetic (PK) properties of new drugs thereby increasing their success rate. As a consequence of the increased throughput of drug discovery, ADME studies have evolved to keep pace. These so-called "early ADME" studies, are characterized by parallel processing and higher throughput than before. A primary concern of medicinal chemists is to design molecules that will have not only the desired activity, but also suitable potency and duration of action, which is influenced by pharmacokinetic properties such as bioavailability and half-life. This article focuses on a particular subset of eADME studies known as "metabolic stability", which can be an important contributor for a good pharmacokinetic profile. Metabolic stability studies represent the adaptation of more complex metabolism rate studies to a minimized system suitable for parallel processing of large numbers of compounds. The theoretical basis for metabolic stability lies in its relationship to the concept of metabolic intrinsic clearance. Typical metabolic stability protocols are discussed with respect to their relation to drug design. How metabolic stability studies have evolved to keep pace with advances in drug discovery is also discussed. Several case studies of the role of metabolic stability in drug design over the past few years are summarized to exemplify the utility of this kind of study. Finally, future trends in drug metabolism and analytical chemistry and how they may influence metabolic stability studies are reviewed.

Pentopril-cimetidine interaction caused by a reduction in hepatic blood flow.

The interactive effects of the coadministration of steady-state cimetidine and single-dose pentopril, an angiotensin converting enzyme inhibitor, on the pharmacokinetic disposition of each other were studied in humans. Cimetidine reduced the clearance of pentopril by 11 to 14%. This reduction in clearance was shown to be caused by a reduction in liver blood flow probably mediated through H2 receptor blockade. Meanwhile pentopril induced the oral clearance of cimetidine by 21%, presumably by a reduction in the bioavailable fraction of cimetidine. The mechanism of this interaction is unknown.

Evaluation of the BBMEC model for screening the CNS permeability of drugs.

Combinatorial synthesis and high-throughput pharmacology screening have greatly increased compound throughput in modern drug-discovery programs. For CNS drugs, it is also important to determine permeability to the blood--brain barrier. Yet, given the increased pace of discovery, it difficult to conduct this screen in a timely fashion. In this presentation, we describe several improvements to an existing CNS permeability screen, the bovine brain microvessel endothelial cell (BBMEC) model. By implementation of these incremental process improvements, we have achieved a robust, facile screen for determination of CNS permeability of multiple compounds.

Determination of ML-1035 enantiomers in plasma by chiral high-performance liquid chromatography.

ML-1035, is a gastroprokinetic agent structurally related to metoclopramide. Because ML-1035 contains an asymmetric chiral sulphoxide moiety, a chiral HPLC method has been developed to separate and quantitate its R- and S-enantiomers in plasma. The ML-1035 enantiomers present in plasma are extracted with dichloroethane under alkaline conditions, the extract evaporated to dryness and reconstituted in the mobile phase. Samples are chromatographed on a Chiralcel OD HPLC column with hexane-absolute ethanol (1% TEA) (1:1, v/v) as the mobile phase. The enantiomers of the unchanged drug are determined by fluorescence measurement (ex: 310 nm, em: 350 nm). The method provides a linear response for both enantiomers over a concentration range of 25 (limit of determination) to 2500 ng ml-1 with correlation coefficients of 0.9987 or greater. The inter-assay precision is 9.5% or less and the accuracy ranges from 93.9 to 103.4% of the theoretical value. The method is used to determine the plasma concentrations of the R- and S-enantiomers following oral and intravenous administration of R- or S-enantiomers to dogs. The method is also adapted to measure enantiomer levels from in vitro reaction mixtures so that the possibility of metabolic inversion may be assessed. The data suggest that no significant level of inversion between the enantiomers occurred either in vivo or in vitro.

The effects of hepatic microsomal enzyme inducers on the pharmacokinetics of ouabain after portal and systemic administration to rats.

The microsomal enzyme inducers 3-methylcholanthrene, phenobarbitone and pregnenolone-16alpha-carbonitrile (PCN) are known to affect other aspects of hepato-biliary disposition in addition to metabolism. This study was designed to determine if presystemic elimination of the non-metabolized xenobiotic ouabain could be altered by these inducers. Male Sprague-Dawley rats were pretreated with inducers or saline for four days. A day later, ouabain (0.5 mg kg-1) was administered into either the ileocolic vein (portal administration) or the femoral vein (systemic administration). Blood and bile samples were collected for up to 90 min after ouabain administration. Biliary excretion rate and cumulative biliary excretion of ouabain were increased by pretreatment with PCN (75 mg kg-1 day-1) relative to controls. Phenobarbitone pretreatment (75 mg kg-1 day-1) also increased these parameters, but to a lesser extent than PCN. In contrast, 3-methylcholanthrene pretreatment (20 mg kg-1 day-1) had no effect on biliary excretion. Plasma concentrations of ouabain were much lower after PCN pretreatment relative to controls, whereas neither phenobarbitone nor 3-methylcholanthrene had any effect. Similarly, clearance (both biliary and total) and volume of distribution were increased by PCN, but not by phenobarbitone or 3-methylcholanthrene pretreatment. Interestingly, the magnitude of biliary and plasma effects induced by PCN appeared to be comparable whether ouabain was administered portally or systemically. Pretreatment of rats with PCN, but not phenobarbitone or 3-methylcholanthrene was shown to increase total clearance of ouabain, mainly via an increase in biliary clearance. Furthermore, because the enhanced clearance occurs after systemic as well as after portal administration of ouabain, a significant change in hepatic presystemic elimination was not detected.

Oxidative and conjugative metabolism of xenobiotics by livers of cattle, sheep, swine and rats.

Homogenate preparations from fresh livers of cattle, sheep, swine and rats were assayed for microsomal cytochrome P-450 content, for mixed-function oxidase activities and for a wide array of conjugative activities using numerous xenobiotic substrates. Results show that hepatic enzymatic capabilities toward xenobiotics do not parallel phylogenetic classifications, thus strengthening the view that most of the comparative data available at present is more descriptive than predictive of relationships among species. Livestock species differed widely from rats in having lower activities of benzo(alpha)pyrene hydroxylase, glutathione S-transferase and acetyltransferase toward isoniazid and sulfamethazine and UDP-glucuronosyl-transferase toward bilirubin. Acetyltransferase activities toward beta-naphthylamine and 2-aminofluorene were not detected in livers of livestock species studied. Cattle livers were remarkably high in activities of styrene oxide hydrolase, ethoxyresorufin O-deethylase, 2-naphthol sulfotransferase and p-aminobenzoic acid acetyltransferase; but notably low in activity of glutathione-S-transferase toward sulfobromophthalein and 1,2-dichloro-4-nitrobenzene. Swine livers had low activity of glutathione-S-transferase toward four of six substrates and low acetyltransferase activity toward four of five substrates. Sheep livers generally were higher than cattle livers in sulfo- and UDP-glucuronsyltransferase activities and lower in acetyl- and glutathionyl-S-transferase. Findings emphasize the risk of error in extra-polations among species and in extrapolations among substrates.

Experimental models for evaluating enzyme induction potential of new drug candidates in animals and humans and a strategy for their use.

Experimental models that have application for evaluating enzyme induction potential have been described in order of increasing complexity. The main focus was on models that have had wide application thus far. However, many new models are currently being developed that may have future applications in evaluating enzyme induction potential. A strategy to evaluate the enzyme induction potential of drug candidates was outlined. This scheme uses a combination of new and established techniques to evaluate data in a stepwise manner that is appropriate to the drug's current stage of development.

Magnifying lens for 800 MeV proton radiography.

This article describes the design and performance of a magnifying magnetic-lens system designed, built, and commissioned at the Los Alamos National Laboratory (LANL) for 800 MeV flash proton radiography. The technique of flash proton radiography has been developed at LANL to study material properties under dynamic loading conditions through the analysis of time sequences of proton radiographs. The requirements of this growing experimental program have resulted in the need for improvements in spatial radiographic resolution. To meet these needs, a new magnetic lens system, consisting of four permanent magnet quadrupoles, has been developed. This new lens system was designed to reduce the second order chromatic aberrations, the dominant source of image blur in 800 MeV proton radiography, as well as magnifying the image to reduce the blur contribution from the detector and camera systems. The recently commissioned lens system performed as designed, providing nearly a factor of three improvement in radiographic resolution.

Measurement of angular distributions of Drell-Yan dimuons in p+p interactions at 800 GeV/c.

We report a measurement of the angular distributions of Drell-Yan dimuons produced using an 800 GeV/c proton beam on a hydrogen target. The polar and azimuthal angular distribution parameters have been extracted over the kinematic range 4.5

Direct observation of the phenomenology of a solid thermal explosion using time-resolved proton radiography.

We present a new phenomenology for burn propagation inside a thermal explosion based on dynamic radiography. Radiographic images were obtained of an aluminum cased solid cylindrical sample of a plastic bonded formulation of octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine. The phenomenology observed is ignition followed by cracking in the solid accompanied by the propagation of a radially symmetric front of increasing proton transmission. This is followed by a further increase in transmission through the sample, ending after approximately 100 micros. We show that these processes are consistent with the propagation of a convective burn front followed by consumption of the remaining solid by conductive particle burning.

Measurement of Upsilon production for p + p and p + d interactions at 800 GeV/c.

We report a high statistics measurement of Upsilon production with an 800 GeV/c proton beam on hydrogen and deuterium targets. The dominance of the gluon-gluon fusion process for Upsilon production at this energy implies that the cross section ratio, sigma(p+d-->Upsilon)/2sigma(p+p-->Upsilon), is sensitive to the gluon content in the neutron relative to that in the proton. Over the kinematic region 0

Measurement of angular distributions of Drell-Yan dimuons in p+d interactions at 800 GeV/c.

We report a measurement of the angular distributions of Drell-Yan dimuons produced using an 800 GeV/c proton beam on a deuterium target. The muon angular distributions in the dilepton rest frame have been measured over the kinematic range 4.5

Optimization of metabolic stability as a goal of modern drug design.

Metabolism and other pharmacokinetic (PK) studies have always played a critical role in helping to optimize the bioavailability and duration of action of new drugs thereby increasing their success rate. With the advent of automated combinatorial synthesis, high-throughput pharmacological testing, and the ability to create extensive databases in the past decade, drug discovery has undergone an amazing evolution. With the increased throughput of drug discovery, metabolism and other PK studies have evolved to keep pace. Often called "early ADME" studies, these studies are characterized by parallel processing and higher throughput than before. This article focuses on a particular class of early ADME (absorption, distribution mechanism, and excretion) studies known as "metabolic stability" studies. The theoretical basis for metabolic stability and its relationship to the concept of metabolic intrinsic clearance is briefly presented. Some key relationships between structure and metabolism are summarized. Several case studies from recent medicinal chemistry literature are reviewed to exemplify how metabolic stability studies influenced drug design and led to improvements in bioavailability and half-life. Finally, future trends in drug metabolism and analytical chemistry and how they may influence metabolic stability studies are reviewed.

Hepatic presystemic elimination of diethylstilbestrol by rats and effect of pretreatment with inducers of UDP-glucuronosyltransferase.

In order to determine if the synthetic estrogen diethylstilbestrol (DES) undergoes hepatic presystemic elimination, male Sprague-Dawley rats were administered [3H]DES (0.005, 0.05, 0.5 mg/kg) into either the ileocolic (portal administration) or femoral (systemic administration) vein. Plasma and bile samples were collected and the concentrations of both DES and DES glucuronide fractions were determined. The concentration of the DES fraction was lower and the proportion of glucuronides was higher after portal than after systemic administration of all three doses. Comparison of the areas under the curve (AUC) of plasma concentration versus time for the DES fraction indicates that the liver diminishes this fraction by 50-70%. Biliary excretion of DES also showed route-dependent differences, in that it was higher for 10-20 min after portal than after systemic administration. To determine if these effects could be enhanced by induction of UDP-glucuronosyltransferase, rats were pretreated for 4 d with phenobarbital, 3-methylcholanthrene, or pregnenolone-16 alpha-carbonitrile and then administered DES (0.5 mg/kg) by the portal or systemic route. 3-Methylcholanthrene and pregnenolone-16 alpha-carbonitrile enhanced the plasma disappearance of the DES fraction to a greater extent after portal than systemic administration and thus further increased the apparent hepatic presystemic elimination. This effect does not appear to involve increased glucuronidation or biliary excretion. These data provide evidence that hepatic presystemic elimination of DES occurs in rats, and this effect can be enhanced by some microsomal enzyme inducers.

Actinomycin D-deoxynucleotide interactions: binding isotherms at the benzenoid and quinoid portions of the drug.

Titrations of actinomycin D (AMD) with dG and with dG-dC were monitored by circular dichroism at 380 nm and 470 nm. These wavelengths are sensitive predominantly to nucleotide binding processes at the benzenoid and quinoid portions, respectively, of the phenoxazone ring of the drug chromophore [Auer, H.E., Pawlowski-Konopnicki, B.E., Chiao, Y.C.C. & Krugh, T.R. (1978), Biopolymers, 17, 1891-1911.]. The temperature dependence of these isotherms was analyzed by the van't Hoff equation to obtain values for the enthalpy and entropy changes. For dG these are about -11 kcal mol-1 and -20 cal mol-1 deg-1, respectively, for complex formation at both the benzenoid and quinoid sites (1 cal = 4.184 J). The enthalpy and entropy changes for complex formation with dG-dC remain unchanged at the benzenoid site, but both values are more negative at the quinoid site. These results indicate that the additional process of binding C in the intercalated AMD-(dG-dC)2 complex, with respect to the simply stacked AMD-dG2 complex, has distinctive properties at the two sites, reflecting their structural differences. The ability to resolve binding processes at the two sites by circular dichroism has permitted us to suggest assignments for the two 31P magnetic resonance lines from the phosphodiester groups observed in the AMD-(pdG-dC)2 complex.