J Pharm Pharmaceut Sci (www.cspscanada.org) 9(2): 2006
Canadian Society for
Pharmaceutical Sciences
Proceedings
of the 9th Annual meeting of the Canadian Society for Pharmaceutical Sciences.
May
24-27, 2006
Ottawa, Ontario, Canada
Scientific Planning Committe Chairs
Gordon McKay and Brian Foster
INVITED PODIUM PRESENTATIONS
Prediction of Drug Absorption
and Disposition Based on the BCS
Leslie Z. Benet, Department of Biopharmaceutical Sciences,
The Biopharmaceutics Classification System (BCS) was developed to allow
prediction of in vivo pharmacokinetic performance of drug products from
measurements of permeability and solubility Although the BCS is useful for
characterizing drugs in Class 1 (high permeability; high solubility) for which
drug dosage form dissolution alone may be amenable for waiver of in vivo bioequivalence
studies, there is little predictability concerning drugs in Classes 2 (high
permeability; low solubility), 3 (low permeability; high solubility) and 4 (low
permeability; low solubility). Last year (Pharm Res 22:13-22, 2005), we
suggested that a modification of such a classification system, designated the
Biopharmaceutics Drug Disposition Classification System (BDDCS), may be useful
in predicting overall drug disposition including: routes of drug elimination;
the effects of efflux and absorptive transporters on oral drug absorption; when
transporter-enzyme interplay will yield clinically significant effects (e.g.,
low bioavailability and drug-drug interactions); the direction, mechanism and
importance of food effects; and transporter effects on post-absorptive systemic
drug concentrations following oral and i.v. dosing. In BDDCS, Classes 1 and 2
drugs are predominantly eliminated by metabolism, while Classes 3 and 4 drugs
are predominantly eliminated unchanged via urinary or biliary excretion. Transporter
effects will be negligible for Class 1 compounds. Efflux transporter effects
will predominate in predicting the oral exposure of Class 2 compounds, while
absorptive transporters will have a major influence on the oral exposure of
Class 3 compounds. We suggest that the BDDCS, using elimination and solubility
criteria, may provide predictability of disposition profiles for all classes of
drugs.
Edward (Ted) M. Hawes, Professor Emeritus,
The vast array of metabolic reactions of xenobiotics can be conveniently
classified as either oxidations, reductions,
conjugations, or nucleophilic trapping processes. Most conjugations involve SN2
reactions of electrophilic adenosine-containing cofactors with nucleophilic
sites in xenobiotics, while formation of amino acid conjugates requires prior
activation of the carboxylic acid substrate. Nucleophilic trapping processes
involve reactions of water, glutathione, or other cellular nucleophiles
(including protein and nucleic acid) with electrophilic xenobiotics.
Categorization of each general route of drug metabolism as resulting in either
increase or decrease in toxicity and pharmacological activity is problematic.
For example, although glucuronidation is traditionally regarded as a route of
detoxification, acyl glucuronides and narcotic analgesic ether glucuronides
have been associated with toxicity and pharmacological activity, respectively.
The gene families of many non-P450 enzymes are well categorized (e.g., UGTs and
GSTs), and in some cases genetic variations are a potential major factor in
affecting interindividual variations (FMO3, NAT2, TPMT, UGT1A1). Virtually any
type of metabolic reaction may play a role in drug activation; widely quoted
examples include O-acetylation of aromatic amines, glutathione S-conjugation of
haloalkanes, and O-sulphation of various N-hydroxy functional groups. Although
drug toxicity cannot be accurately predicted, approaches can be used in drug
discovery and development to minimize potential problems. These approaches
include use of structure-metabolism relationships in drug design, small
molecule trapping agents, and radiolabelled substrate. Selected routes of
non-P450 drug metabolism will be discussed, including involvement in
contributing to interindividual variation and bioactivation.
Reina Bendayan,
Several families of membrane transport proteins can regulate drug
distribution in the central nervous system. In particular, members of the
ATP-binding Cassette (ABC) superfamily of transporters, such as P-glycoprotein
(ABCCB1/MDR1), multidrug resistance-associated proteins (MRPs/ABCC) and breast
cancer resistance protein (BCRP/ABCG2), localized at the brain barriers and
brain parenchyma can play a major role in the overall disposition of drugs in
the brain. The goals of this presentation are: i) to provide an overview on the
molecular expression, cellular/subcellular location and functional activity of
ABC membrane-associated transporters, primarily P-glycoprotein and MRPs in
several brain cellular compartments (i.e., brain microvessel endothelial cells,
astrocytes and microglia) and ii) to discuss their clinical implications in the
pharmacotherapy of neurological disorders (i.e., HIV infection).
Medicinal Cannabis and a New
Oro-mucosal Aerosol
Ethan Russo, Senior Medical Advisor, GW Pharmaceuticals,
THC inhibits cAMP through G-protein receptor coupling. It is a partial
agonist on CB1 receptors, especially in pain pathways. THC actions include
analgesia, muscle relaxant and anti-inflammatory effects. Cannabidiol (CBD) has
anti-anxiety, anti-psychotic, anti-oxidant, anti-inflammatory, immunomodulatory
effects, and prevents glutamate excitotoxicity. Sativex® is a
well-characterised botanical drug product derived from two clonal cannabis
chemovars, one THC-predominant (Tetranabinex®) and another CBD-predominant
(Nabidiolex®), yielding a botanical drug substance (BDS) of defined composition
with controlled reproducibility. THC and CBD comprise some 70% (w/w) of the
BDS, with minor cannabinoids (5 – 6%), terpenoids (6 – 7%, most GRAS), sterols
(6%), triglycerides, alkanes, squalene, tocopherol, carotenoids and other minor
components (also GRAS). Sativex is administered oromucosally with each 100
Lpump-action spray providing 2.7 mg of THC and 2.5 mg of CBD, in an ethanol:
propylene glycol vehicle with 0.05% peppermint flavouring. To date, RCTs show
Sativex to have statistically significant benefits in several medically
intractable conditions: pain associated with peripheral neuropathy, rheumatoid
arthritis, cancer pain unresponsive to opiates and neuropathic pain in multiple
sclerosis (indication for the NOC/C of Sativex in Canada in 2005), as well as
spasticity, and lower urinary tract symptoms associated with MS, and sleep
quality in these various disorders. In all studies, Sativex has been used as
add-on therapy in patients who have not responded adequately to their existing
medication. These trials and their safety-extension studies (up to four years)
have demonstrated no abuse or diversion of Sativex, no tolerance to symptomatic
benefits, or significant withdrawal effects upon sudden discontinuation.
SESSION 2: Safety Issues
Issues in Hepatotoxicity
Jack Uetrecht, Leslie Dan Faculty of Pharmacy,
Adverse drug reactions have become a major cause of drug candidate
failure. Idiosyncratic reactions are especially difficult to deal with, and
hepatotoxicity is the most common type of idiosyncratic reaction leading to
drug withdrawal. It would be a major advantage if risk could be determined
early. Current animal testing does not predict the risk of idiosyncratic
reactions. It appears that most idiosyncratic reactions are mediated by
reactive metabolites; therefore, screens for reactive metabolites may make drug
candidates safer. Several companies have invested heavily in this concept, but
it is not clear whether it really decreases risk. In clinical trials, the best
predictor of idiosyncratic hepatotoxicity is an elevation of ALT; however,
several drugs are associated with a significant incidence of elevated ALTs
without posing a significant risk for liver failure. After the drug reaches the
market, databases of spontaneous reports can provide an important early
warning; however, causality assessment is very important. This is especially
important because there are many other causes of liver failure. In order to
improve on our ability to predict the risk of hepatotoxicity, we need a much
better understanding of the basic mechanisms involved. Idiosyncratic
hepatotoxicity has been divided into immune-mediated reactions and metabolic
idiosyncrasy; however, metabolic idiosyncrasy in particular is undefined. An
important tool for mechanistic studies is valid animal models and two animal
models of idiosyncratic reactions will be presented; however, developing animal
models is difficult because such reactions are also idiosyncratic in animals. Funded by CIHR.
Drug, Food, Natural Health
Product Interactions
Brian C. Foster, Therapeutic Products Directorate, Health
Natural health products (NHPs) and functional foods are widely used as
complementary alternative medicines (CAMs). Traditional uses of CAMs have
generally proven safe, but their current pattern of consumption and uses in the
global context has changed. Many NHPs are being used concurrently by every
patient population and serious adverse events have been identified. Health
Safety Assessment in Clinical
Trials
Jim Gallivan, Clinical Trials Division, Centre for Evaluation of
Radiopharmaceuticals and Biotherapeutics, Biologics and Genetic Therapies
Directorate, Health Products and Food Branch, Health Canada, Ottawa, Canada
Safety assessment is a continuous process that begins prior to the first
exposure of humans and continues throughout drug development. Clinical trials
are essential to the drug development process because they provide quantitative
information needed for the assessment of efficacy and safety for the market
authorization of new drugs and for continued marketing. This presentation will
review the regulatory framework and procedures to evaluate safety and to
minimize the risks to subjects prior to, and during, clinical trials. Future
directions in safety assessment in clinical trials will also be considered.
Pharmacogenomics: The state
and art of the Regulatory Science
Agnes V. Klein, Centre
for the Evaluation of Radiopharmaceuticals and Biotherapeutic Products,
Biologics and Genetic Therapies Directorate, Health
Pharmacogenomics as a part of drug development and the search for
rational targets for therapeutics is based on scientific activities that apply
the knowledge acquired during the Genome Project. The fact that the era of
blockbuster drugs appears to have drawn to a close, has lent additional impetus
to this and other related “omics” areas. Genetics as a science has provided
insights into the predisposition of individuals to diseases and has opened up
areas where Biomarkers have come to the forefront. Thus, opportunities have
been opened to the conversion of these Biomarkers as targets for therapy
creating a promise for better tailored and more individualized therapy. The era
of Pharmacogenomics has spawned such products as Herceptin and Erbitux that are
targeted at specific mutations of tumour cells which then can be suppressed by
the appropriately targeted product. It has also become clear that many of the
metabolic variants for pharmaceuticals result in different levels of efficacy
for different subpopulations and that the safety of therapeutics may be linked
to genetic factors. However, PGx has not been incorporated as yet into daily drug
development. In order to facilitate this reality both regulatory and policy
bodies have been preoccupied with the development of enabling policies and
nomenclature. Hence a documents produced by CIOMS, another by the EMEA, and yet
another guidance document generated by the FDA.
SESSION 3: Pharmacokinetics: focus on dose
proportionality and other applications of population pharmacokinetics (
Jake J. Thiessen, Leslie Dan Faculty of
During the 1990-92 period, the Canadian Expert Advisory Committee (EAC)
on bioavailability, chaired by Dr. John Ruedy, produced Reports A, B, C
offering bioavailability and bioequivalence guidelines encompassing a spectrum
of drugs and products. Report A was made official by Health Canada in 1992, and
provided a guidance for conventional formulations of oral drugs that
haduncomplicated characteristics.Various exceptions were identified in this
guidance including modified release formulations (a 1996 official guidance
dealt with this (Part B: Oral Modified Release Formulations)) and drugs with
so-called complicated or variable pharmacokinetics, including non-linear
kinetics. More recently, Health
Introduction and Background
on the Use of Population Pharmacokinetics: Differences with Conventional
Methods, Strengths and Limitations of This Approach.
Thomas M. Ludden, GloboMax, a Division of ICON plc,
Since the introduction of the concept of using mixed effect modeling
(MEM) to study pharmacokinetics (population pharmacokinetics) in 1972, the
application of this methodology has grown until today it is used throughout the
drug development process and in post-approval monitoring of drug safety and
efficacy. MEM involves the development of a mathematical description of the
influences of fixed effects (time, dose, demographics, concurrent drug therapy,
etc.) and random effects (variability between individuals, between different
dosing occasions, residual variability) on an outcome variable (drug
concentration or effect). Such an analysis differs from conventional two-stage
analysis in that it estimates both fixed and random effects simultaneously. The
two-stage approach fails to correctly partition variability among its various
sources. This is particularly true if the data per individual are sparse but
less so with dense and informative data. The primary advantage of MEM is
primarily its ability to extract information from data sets with moderate
information per individual. The primary disadvantage of MEM is that it may
require substantial time devoted to model building efforts, a process that
makes it difficult to have a rigorous statistical interpretation of the
results. A dual-phased approach to a MEM analysis can sometimes be used to
balance the advantages and disadvantages of MEM. The initial phase can address
previously stated hypotheses, while the second phase can be devoted to
exploratory analyses.
Using Mixed Effects Modeling
(MEM) to Evaluate Dose Proportionality (DP) and the Effect of Concomitantly
Administered Medications (CM) in Patients
Diane R Mould, Projections Research Inc.,
The development of new therapeutic agents has become more complex over
time. Because patients are seldom treated with only one agent, and may require
dose adjustments, evaluations of DP and CM are critical. Furthermore, the
effects of age and disease can alter the pharmacokinetics in patients.
Determining appropriate dose regimens requires understanding patient specific
changes in disposition and drug sensitivity, as well as evaluation of the
likely drug combinations that may be used. The development of therapeutic
agents should investigate these issues, however, evaluating dose
proportionality and drug interactions in patients may be difficult due to
ethical constraints or for other practical reasons. The use of population
pharmacokinetic (PPK) analysis has become an important tool in drug
development. Using MEM, sparse sampling schedules can provide information about
PPK in patients. Although model based evaluations inherently include
assumptions and the model can not always be completely specified prior to
analysis, several methodologies have been proposed for inferential evaluation.
For such work, the study design is important, the analysis plan should be
pre-specified to the extent possible, and the anticipated power and type I
error should be evaluated for the design and the model. The PPK model
performance must be evaluated rigorously and the assumptions used in the model
should be verified. Two cases are presented as examples of using MEM for the
evaluation of DP and CM for anti-viral agents. The results show that with an
adequate study design, DP and the effect of CM can be evaluated with good
precision.
Strengths and advantages of
doing
This presentation will be broken up into three parts. First, we will
review the differences between individual and population approaches in PK/PD and
explain the key elements supporting the increasing use of the population
approach. Secondly, we will present the current state of the art of using
population PK/PD studies within the drug development process and contrast this
with what has been done in the past. Finally, case examples will be presented
to highlight the advantages and strengths of this approach within the drug
development process.
SESSION 5: Regulatory issues
Emerging regulatory issues in
pharmaceuticals
Omer Boudreau, Therapeutic Products Directorate, Health
This talk will provide an update on the Therapeutic Products Directorate
(TPD) activities and accomplishments. TPD is currently outlining its plan for
the upcoming year. Planning is the key for the success of any organizations but
it can be derailed by unexpected situations. New developing scientific issues
are always a challenge that TPD must be ready to face. Performance
Sustainability and Modernizing the Regulatory Framework are two major
objectives in our strategic plan that could strongly impact the emerging
regulatory issues in pharmaceuticals. Modernizing of the regulatory framework
through a new framework (Progressive Licensing) will enable us to monitor and
assess the entire product cycle. As there are many regulatory gaps in the
current framework, the new framework would allow sound risk management and
access to new drug therapies while continuously monitoring and reassessing for
safety, quality and efficacy. The presentation will also cover some notable
activities in 2005, how the elimination of backlog and performance
sustainability have been achieved, and point out some emerging regulatory
issues/trends.
Emerging biotechnology
regulatory issues in biologics and genetic therapies directorate
Pierre Charest, Biologics and Genetic Therapies Directorate, Health
Not available at time of publication.
Nanotechnology and the
management of health risks: Regulatory oversight and challenges
Hans Yu, Chief, Departmental Biotechnology Office, Health
Nanotechnology is a complex and rapidly evolving field of science and
discovery that involves creating and/or manipulating materials at a very small
scale (in the billionths of a metre). It is considered a transformative
technology widely perceived to have significant socio-economic impacts in
Challenges in Product
Classification
Micheline Ho, Office of Risk Management, Therapeutic Products
Directorate, Health
Health Canada has the responsibility to ensure that products [e.g.
pharmaceuticals, medical devices, food, cosmetics or pest control products]
regulated under the various legislative instruments within its mandate are in
compliance with the relevant statutes [including the Food and Drugs Act, Pest
Control Products Act]. In order to fulfill this responsibility, it is necessary
to be clear as to which product types are covered by each statute. Although
this may seem to be a simple issue at first glance, it is in fact becoming a
significant challenge. This challenge is the result of, not only the
introduction of new product types, but more likely the result of new types of
representations for existing product types, such as: health foods, new product
combinations (e.g. medicated stents), co-packaged products or “convenience
packages”, while we attempt to increase clarity and transparency in the
regulatory process. The presentation will provide specific examples to
illustrate the challenges faced by the regulator, as well as potential pitfalls
in classification decisions and an international perspective on the issue of
product classification.
Calcium Silicate Based
Floating Granular Delivery System for Ranitidine Hydrochloride: A Novel
Approach to Control Oral Delivery Via Gastric Retention
Ashish Jain, Prateek Jain, R.K. Agrawal, Pharmaceutics Division,
Department of Pharmaceutical Sciences, Dr. H. S. Gour University, Saga, India
Purpose: We prepared an intragastric floating granules using porous
calcium silicate (FLR) as a floating carrier, which has floating ability due to
the air included in the pores when they are covered with a polymer. Method:
Floating granules were prepared by dropping a10% (w/v) ethanol solution of
hydroxylpropylmethylcellulose (HPMC) and ethyl cellulose (EC) in four different
concentration ratios (5:95, 10:90,15:85 and 20:80) while the 4.5g ranitidine
hydrochloride adsorb 15g FLR was being agitated in a beaker. After the mixture
was dried in vacuum and sieved (No. 22), we regarded the granules obtained as ranitidine
hydrochloride primary coated granules (RHPCG). After drying, the ethanol
solution of polymer was dropped and dried in vacuum again, and sieving was
carried out to obtain ranitidine hydrochloride secondary coated granules
(RHSCG).Results: The floating property of RHSCG was better than that of RHPCG.
A longer floating time was observed with a lower HPMC concentration in
composition ratio Since HPMC begins to erode when the HPMC ratio of the HPMC/EC
matrix system is over 15%. It was observed by a scanning electron microscope
(SEM) that more pores of FLR in RHSCG were covered with a polymer than those in
RHPCG. RHSCG showed a smaller release rate than RHPCG. It also has a sustained
drug release properties. The serum concentration Cmax of ranitidine hydrochloride
in rats, which received (5:95) HPMC: EC coated formulation RHSCG1 was found to
be 1.56 g/ml after (tmax) 5 hr and AUC0-7 was 7.58 g.hr/ml.
Conclusion: These results suggest that FLR is a useful carrier for the
development of floating and sustained release preparations. Acknowledgements:
The first name author is thankful to All India Council of Technical Education
for financial assistance.
Development of EGF-Conjugated
Block Copolymer Micelles for Actively Targeted Drug Delivery to EGFR-Overexpressed
Cancers
Helen Lee 1, Faquan Zeng 1,
Christine Allen 1,2,3*; 1. Department of
Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy; 2. Department of Chemical Engineering and Applied Chemistry, Faculty
of Applied Science and Engineering; 3. Department of Chemistry, Faculty
of Arts and Science,
PURPOSE: Nano-sized polymeric micelles formed from amphiphilic copolymers
are capable of encapsulating and delivering hydrophobic drugs to tumors by
passive targeting. Recently, much effort has focused on developing actively
targeted drug-loaded micelles via ligand-coupling in order to further increase
drug selectivity towards cancer cells. We have conjugated epidermal growth
factor (EGF) to poly(ethylene
glycol)-block-poly(-valerolactone) (PEG-b-PVL) copolymers for the preparation
of micelles that target drugs to EGF receptor (EGFR) overexpressed cancer
cells. METHODS: Targeted micelles were prepared from EGF-PEG-b-PVL copolymers;
while non-targeted micelles were formed from the PEG-b-PVL copolymers. A
hydrophobic fluorescent probe was loaded into the micelles to serve as a tracer
for cell uptake studies. The cell uptake profiles of the targeted and
non-targeted micelles were evaluated in EGFR-overexpressed breast cancer cells
(MDA-MB-468) by fluorescence-measurements and confocal microscopic analysis.
RESULTS: The targeted and non-targeted micelles were found to have mean
diameters of 32 nm and 45 nm, respectively. Minimal cell uptake was detected in
cells incubated with non-targeted micelles; while incubation of cells with the
targeted micelles resulted in significant intracellular accumulation. In
addition, the presence of EGF reduced the intracellular accumulation of
targeted micelles to a similar level as observed in cells incubated with
non-targeted micelles. Confocal microscopic analysis also revealed that the
targeted micelles mainly localized in the perinuclear region, with occasional
nuclear localization. CONCLUSION: EGF-conjugated micelles were internalized by
MDA-MB-468 breast cancer cells via receptor-mediated endocytosis, which may
allow for more efficient delivery of anticancer agents to EGFR-overexpressed
cancer cells.
Biological and Mechanical
Evaluation of a Polymer-Lipid Blend for Drug Delivery
J. Grant1 V. Vassileva, M.
Piquette-Miller111, C. Allen1*, 1. Leslie Dan Faculty of Pharmacy,
Department of Pharmaceutical Sciences
PURPOSE: We developed a novel blend composed of the natural
polysaccharide chitosan and the lipid egg phosphatidyl-choline (ePC). The biological
and mechanical properties of the chitosan-ePC film were evaluated for use in
localized drug delivery. METHODS: The invitro cytotoxicity was determined in
CHO-K1 fibroblast cells and SKOV-3 human ovarian adenocarcinoma cells. The
degree of protein adsorption to the films was evaluated in fetal bovine serum;
while the in vivo biocompatibility was assessed in healthy CD-1 mice by
examining the chitosan-ePC film macroscopically and histopathologically
following a one-month implantation. The mechanical properties were determined
by dynamic mechanical analysis and Instron testing. Nanoparticles containing
the hydro-phobic anticancer agent paclitaxel were dispersed throughout the
chitosan-ePC film and the drug release profile was analyzed in physiologically
relevant media. RESULTS: A negligible reduction in cell viability and a minimal
degree of protein adsorption to the chitosan-ePC films provided preliminary
indications of the biocompatibility of the chitosan-ePC film. Furthermore, no
observable signs of infection, inflammation, local irritation or fibrous
encapsulation were observed in the CD-1 mice implanted with the chitosan-ePC
film. In addition, the modulus of the chitosan-ePC films was found to decrease
significantly as the amount of lipid increased, thus creating a softer
biomaterial that is suitable for use as an implant. A sustained release of
paclitaxel from the film was achieved over a four-month period. CONCLUSION: The
chitosan-ePC film has shown excellent biocompatibility and mechanical properties,
and may be a promising implant system for the sustained delivery of hydrophobic
agents.
Evaluation of a
Pharmaceutical Care Service for Asthmatic Patients in
Awad AI, Abdelhamid K, Gasmallah A, Department of Pharmacy Practice, Faculty
of Pharmacy, Kuwait University; Department of Pharmacology, Faculty of
Pharmacy, University of Khartoum, Sudan, Department of Internal medicine,
Faculty of Medicine, Al Neelain University, Sudan.
Interest of Community
Pharmacists in Health Promotion in
Awad AI, Al-Thauwaini M, Abahussain E, Department of Pharmacy Practice,
Faculty of Pharmacy, Kuwait University
Purpose: To describe the current practice of community pharmacists with
regard to their provision of health promotion activities, identify their
willingness to participate in health promotion and identify the barriers that
may limit their participation. Methods: The study included 70 community
pharmacies that were selected using stratified and systematic random sampling.
Data were colleted via face-to-face structured interview of the respondents
using a pre-tested questionnaire. Results: The majority of study participants
(65.2%) were strongly involved in counselling patients on health promotion
related to medications, but less involved in counselling them on the other
personal health behaviours. Most of the pharmacists perceived themselves as
very prepared to counsel patients on taking drugs (70%) and less prepared to
counsel them on other personal health behaviours. Half of the participants
(50%) claimed a high level of success in helping patients to change their
behaviour with regard to medications, but not with regard to other personal
health behaviours. The majority of the study participants (74.3%) have the
interest and willingness to participate in continuing education programs so as
to learn more about health promotion. The barriers facing community
pharmacists’ participation in health promotion activities as perceived by
respondents were as follow: lack of pharmacists’ time (60%), lack of patients’
time (50%), lack of information and/or training (32.9%), and lack of privacy or
pharmacy physical design (32%). Conclusions Community pharmacists reported to
achieve considerable success in helping patients to change their behaviour in
relation to medications, but were less confident of their ability to change
personal health behaviours. The majority of the respondents have the
willingness to be a prime source of advice and support on health promotion.
Girish Bende, Sivacharan Kollipara, and Ranendra N. Saha, Pharmacy Group,
Birla Institute of Technology and Science, Pilani, India
Purpose: The objective of this study was to formulate and evaluate
polysorbate-80 coated nanoparticles of Imatinib mesylate using biodegradable
and biocompatible polymeric material - PLGA for effective brain delivery.
Imatinib mesylate (IM) is a selective protein tyrosine kinase inhibitor which
inhibits the Bcr-Abl and cKIT tyrosine kinases in chronic myeloid leukemia and
gastrointestinal stromal tumor, respectively. Although, IM has promising
penitential in the treatment of brain tumors, it shows poor brain penetration
because of efflux transport. Nanoparticulate formulations have the potential to
overcome efflux transport system and also to alter tissue distribution through
enhanced permeation and retention (EPR) effect of several water soluble drugs.
Methods: Emulsification followed by solvent evaporation technique was adopted
for preparation of polysorbate-80 coated PLGA nanoparticles of IM.
Encapsulation efficacy, total drug content and in vitro release profile were
studied using stability indicating HPLC method. Morphological characters of the
nanoparticles like shape, average size, and size distribution were studied
using AFM, PCS, and TEM. Additionally, drug-excipient compatibility studies
were performed using HPLC, FTIR, and DSC. Rat in situ absorption model was used
to study intestinal absorption of pure drug and prepared nanoparticles.
Moreover, in vivo brain uptake was investigated for pure drug and prepared
nanoparticles in Rat model. Results:Nanoparticles have
shown good entrapment efficiency (91.5%) with high drug content. Additionally,
it has provided extended drug release for over 24 hours when studied in
specially designed diffusion cell equipped with dialysis membrane. AFM studies
revealed the spherical shape of particles, while TEM and PCS studies confirmed
mean size of 200 nm with narrow size distribution (poly-dispersity index 0.05).
HPLC, DSC, and FTIR studies confirmed that the drug did not have any physical
or chemical incompatibilities with polymer and other excipients. In situ rat
intestinal absorption study showed significant absorption of prepared
nanoparticles.In vivo brain uptake studies in rat model revealed enhanced
permeation of nanoparticles compared to pure drug. Conclusion: Adopted method
of preparation has provided polysorbate-80 coated PLGA nanoparticles with high
drug content and good entrapment efficiency. Prepared nanoparticles not only
controlled the release but also enhanced spatial delivery of drug to brain.
These nanoparticle formulations can be used for effective brain delivery of
water soluble drugs like IM.
A specific and sensitive
liquid chromatography-mass spectrometry (LC-MS) method for simultaneous
determination of both amiodarone and desethylamiodarone in rat plasma
Anooshirvan Shayeganpour, Vishwa Somayaji and Dion R, Brocks, Faculty of
Pharmacy & Pharmaceutical Sciences, University of Alberta, Edmonton, Canada
Amiodarone (AM) is extensively used for its antiarrhythmic properties. In
humans, the most prevalent circulating metabolite of AM is desethylamiodarone
(DEA). In rat, a commonly used animal model, plasma concentrations of DEA are
low, and near the lower limit of detection of most conventional assays using
ultraviolet detection. Purpose: To develop a sensitive and specific high
performance liquid chromatography-mass spectrometry (LC-MS) assay method for
the determination of AM and DEA in rat plasma. Methods: To 0.1 mL of rat plasma
samples containing 0.03 mL of internal standard (IS) solution (2.5 μg/mL
ethopropazine), 0.3 mL of acetonitrile was added. Tubes were vortex mixed and
centrifuged for 2 min and the supernatant was carefully transferred to new
glass tubes. Then 0.3 mL of HPLC water and 3 mL of hexane were added. Tubes
were vortex mixed for 30 s and centrifuged for 3 min. The top layer was
transferred and evaporated to dryness in vacuo. Residues were reconstituted
using 1 mL of mobile phase and 5-10 μL were injected into the LC-MS system
consisting of a Waters Micromass ZQTM 4000 spectrometer coupled to a Waters
autosampler and pump. A Waters XTerra® MS C18 3.5 μm (2.1×50 mm) column
heated to 45°C was used for separation. The mobile phase consisted of methanol
and 0.2% aqueous formic acid pumped as a linear gradient from 40:60 methanol:formic
acid 0.2% to 90:10 over 15 min, at a constant flow rate of 0.2 mL/min. The
concentrations of IS, DEA and AM were monitored for m/z values of 313.2 (IS),
617.90 (DEA) and 646.0 (AM). Results: In plasma, linearity was present between
the peak height ratios and concentrations over the range of 10-1000 ng/mL for
AM and 2.5-1000 ng/mL for DEA (r2>0.999). For HF and DHF intraday and
interday CV were less than 20%, and mean error was <22%. After
administration of AM to rats and assay of plasma, the eluted peaks representing
AM, DEA and IS were all found to be free of interference. Conclusion: The LC-MS
method was sensitive, specific and appropriate for the simultaneous assay of AM
and DEA in rat plasma.
Role of Peroxisome
Proliferators In Liver Fatty Acid-Binding Protein
(L-FABP) Expression and Oxidative Function.
Jing Yan1, Yu Gong1, Guqi Wang1, Yuewen Gong 1 3, Frank J. Burczynski 1
2, Faculty of Pharmacy, Department of Pharmacology and Therapeutics,
3Department of Internal Medicine, Faculty of Medicine,
INTRODUCTION: L-FABP has been shown to contain significant antioxidant
activity. Pharmacological treatment directed at increasing or decreasing L-FABP
levels was associated with either a decrease or increase in reactive oxygen
species following H2O2treatment. In this study we investigated the role of
peroxisome proliferator-activated receptors (PPARs) in the expression and
antioxidant activity of L-FABP. METHODS: L-FABP expressing 1548-hepatoma cells
were treated with dexamethasone, clofibrate, PPAR alpha antagonist MK886, PPAR
gamma antagonist GW9662 or combination treatment. Oxidative stress was induced
by H2O2incubation. The fluorescent marker, dichlorofluorescein (DCF), was
employed to measure intracellular reactive oxygen species (ROS). RESULTS:
L-FABP expressing 1548-hepatoma cells, treated with dexamethasone or clofibrate
decreased and increased intracellular L-FABP levels, respectively. Expression
of L-FABP was reduced after treatment with MK886 or GW9662. Clofibrate mediated
L-FABP activation was inhibited by both PPAR antagonists. DCF data showed that
ROS levels reflected L-FABP levels. CONCLUSION: Our study shows that PPAR
agonists and antagonist treatment affects cellular antioxidant function. This
study was supported by an operating grant from the Canadian Institute of Health
Research.
Cíntia M. S. Cereda 1, Giovana R. Tófoli 2, Márcio A. Paschoal 1,
Leonardo F. Fraceto 1 3, Daniele R. de Araujo 1, Eneida de Paula 1, State
University of Campinas, Institute of Biology, Department of Biochemistry,
Campinas, São Paulo, Brazil, 2State University of Campinas, Piracicaba Dental
School, Department of Physiological Sciences, Piracicaba, São Paulo, Brazil,
3University of Sorocaba, Department of Pharmacy, Sorocaba, São Paulo, Brazil.
Determination of trigonelline
in herbal extract and pharmaceutical dosage form by a validated HPTLC method
Shruti Chopra*, Farhan J. Ahmad, Roop K. Khar, Saiqa Mahdi, Zeenat Iqbal,
Department of Pharmaceutics, Faculty of Pharmacy, Jamia Hamdard, New Delhi,
India
PURPOSE: The objective of the present investigation was to develop a
validated HPTLC method for the determination of Trigonelline in herbal extracts
and in pharmaceutical dosage forms.METHODS: Analysis of trigonelline was
performed on TLC aluminium plates pre-coated with silica gel 60F-254 as the
stationary phase. The mobile phase consisted of n-propanol: methanol: water
(4:1:4 v/v/v). Linear ascending development was carried out in twin trough
glass chamber saturated with mobile phase at room temperature and the mobile
phase was run up to 8 cm. Camag TLC scanner III was
used for spectrodensitometric scanning and analysis in absorbance mode at 269
nm. RESULTS: The system was found to give compact spots for trigonelline (Rf value of 0.46 ± 0.01). The linear regression analysis
data for the calibration plots showed good linear relationship with r2 = 0.9991
± 0.0002 in the concentration range 100-1200 ng per spot. The mean value (±
S.D) of slope and intercept were 4.1312 ± 0.0491 and 208.2135 ± 4.5092
respectively. According to ICH guidelines the method was validated for
precision, recovery, robustness and ruggedness. The mean % recoveries ranged
from 98.29 – 102.19 %. The limits of detection and quantification were 2.27 ng
and 7.58 ng, respectively. The trigonelline content of herbal extracts was
quantified. Drug content estimated from the formulation was well within the
limits (±5% of the labelled content of the formulations). CONCLUSIONS:
Statistical analysis of the data showed that the method is reproducible and
selective for the estimation of trigonelline. Since the proposed mobile phase
effectively resolves trigonelline, the developed HPTLC method can be applied
for identification and quantification of trigonelline in herbal extracts and
pharmaceutical formulations.
JD Clements, M Skrzypiec-Spring, J Sawicka,
3R Schulz, F Jamali; Faculty of Pharmacy and Pharmaceutical Sciences, UofA,
PURPOSE: Inflammatory conditions downregulate cardiac beta1-receptors.
Matrix metalloproteinases (MMPs) are involved in inflammatory processes and
their inhibition can prevent the loss of contractile function. Activation of
cardiac sympathetic nerves, something observed in rheumatoid arthritis
patients, may contribute to reduced cardiovascular drug responsiveness in
humans Pre-adjuvant arthritis (Pre-AA) is an inflammatory model that reduces
cardiovascular response to propranolol. The diminished effect appears to be
restored by, the hydroxymethylglutaryl-CoA reductase inhibitor pravastatin.
Hypothesis: The anti-inflammatory effects of pravastatin will normalize
elevated MMP-2, MMP-9, and norepinephrine (NE) in the myocardium METHODS: Rats
were injected with Mycobacterium butyricum, and after four days oral
pravastatin (6 mg/kg, BID) was given until day 8. In heart homogenates we
detected MMP-2 and MMP-9 by gelatin-zymography, and using competitive ELISA we
detected heart NE. RESULTS: After 8 days, Pre-AA caused a significant
twenty-fold elevation in myocardial MMP-9 activity as compared with healthy
rats (Average band density±SEM; 170±58 vs. 8.3±1.7 units/mg; n=12/group) that
was not attenuated by pravastatin administration (Pre-AA/Placebo vs.
Pre-AA/Pravastatin; 170±58 vs. 141±57 units/mg; n=12/group). Neither MMP-2 nor
NE was influenced by Pre-AA induction or by pravastatin administration.
CONCLUSIONS:The mechanism by which Pre-AA induces
inflammation may involve MMP-9 activation, but how pravastatin assists in
restoring cardiovascular drug non-responsiveness remains to be determined.
Sunita Dahiya, Kamla Pathak , Department of
Pharmaceutics,
Purpose: The purpose of present investigation was to study potential of
water soluble dissolution enhancers polyvinylpyrrolidone K30, polyethylene glycol 400 and propylene glycol on in
vitro dissolution of a poorly water soluble drug meloxicam to develop
multi-component pseudosolid dispersion system. Methods: Preliminary studies
were conducted using a physical mixture of meloxicam and the excipients lactose
and microcrystalline cellulose while solid dispersions were prepared by solvent
evaporation and cogrinding method using aqueous solution of these enhancers.
Based on the data of preliminary studies, a 33factorial design was adopted to
optimize the concentration of solubilizing agents. The concentrations of enhancers polyvinylpyrrolidone K30, polyethylene glycol and
propylene glycol were used as independent variables whereas angle of repose and
in vitro dissolution were used as dependent variables. Full and reduced models
were evolved for the dependent variables and the reduced models were further
validated using two check points. Angle of re-pose < 35˚, percentage of
drug dissolved in 30 min (Q) > 45%, 45 min (Q3045) > 55% and 120 min
(Q120) > 65% were used as constraints for the selection of optimized batch.
Contour plots were presented for the selected dependent variables. Results:
Polyvinylpyrrolidone was found to be more effective in increasing the drug
dissolution, compared with liquid enhancers polyethylene glycol 400 and propylene glycol. The granule flow was adversely
affected when high levels of liquid enhancers were incorporated in the
formulations. Wet-ability studies were conducted to measure wetting time of
selected batches and the improved dissolution was attributed to improved
wetting and solubilizing effects of adjuvants from the pseudosolid dispersions
of meloxicam. More than a five fold increase in drug dissolution was observed
in some batches compared with pure drug powder. The similarity factor (f2) for
the selected batches were in the range 50-100 which were in accordance with the
SUPAC FDA guidelines. Conclusion: Cogrinding of meloxicam with aqueous solution
of dissolution enhancers offered potential advantages such as reduced cost,
improved safety and environment friendliness as compared to solvent evaporation
method which used dimethyl formamide. The studies revealed that optimum levels
of solubility enhancers based on systematic statistical approach could be
appropriately blended with pharmaceutical adjuvants to demonstrate
significantly higher release rates of meloxicam in an economical and cost
effective manner.
DEVELOPMENT OF CONTROLLED
RELEASE PLATFORM FOR HIGH DOSE GASTRO RETENTIVE DRUG DELIVERY
Anupama Diwan, Kanchan Kohli, Sanjula Boboota, Javed Ali, Vipin Dhall,
M.Zaffer; Dept. of Pharmaceutics, Faculty of Pharmacy, Jamia Hamdard, Hamdard
Nagar, New Delhi.
Purpose: This study was performed to prepare controlled release gastro
retentive (GR) formulation for high dose drugs. Ciprofloxacin Hydrochloride had
been selected as model drug. It is highly water-soluble drug with an absorption
window at upper jejunum with dose of 598.47 mg. Methods: The matrix based
controlled release formulation strategy for GR tablets were used. The drug is
mixed with polymers and half of the amount of sodium bicarbonate and then the
mix was granulated with isopropyl alcohol. Various polymer combinations have
been tried viz: HPMC, Ethylcellulose, Sodium alginate, Xanthum gum, Kollidon SR
and their combination thereof. An optimized platform for high dose GR
formulation has been prepared. Results:Even with low
compression forces tablets of astonishing hardness has been obtained with
Kollidon SR and xanthum gum combination. Also these formulations have shown
better floating behavior (18 Hr) as compared to HPMC based floating tablets,
that sink after certain time. The formulation was achieved with lowest ratio of
polymer to drug as low as 1:3. Similarly, amount of Sodium Bicarbonate to total
tablet weight was achieved in ratio of 1:5. Dissolution profiles, 30% of the
drug were released in first hr and 70% of the drug was released in next 8-10
Hrs. The total tablet weight was less than1g, the objective set for high dose
formulation. Conclusion: Formulation of Ciprofloxacin Hydrochloride GR tablets
in Kollidon SR and Xanthum gum combination were better than HPMC based formulations.
A floating time of more than 18Hrs was easily achieved. Dissolution data proves
24 Hrs controlled release profile.
MODULATION OF CLASS III
ANTIARRHYTHMIC PROPERTIES OF d-SOTALOL BY GLUCOSE CONCENTRATION
Ricard G, Simard C, Daleau
P, Drolet B, Faculté de pharmacie, Université Laval, Centre de recherche,
Hôpital Laval, Québec, QC, Canada
Purpose: Prolongation of the QT interval on the electrocardiogram by
blocking cardiac potassium channels is a well-known property of Class III
antiarrhythmics, such as d-sotalol.Excessive QT prolongation is a trigger for
ventricular arrhythmias (torsades de pointes). Interestingly, QT prolongation
has also been associated with both hypo- and hyperglycemia seen in diabetes. We
hypothesized that low and/or high glucose levels potentiate the effect
ofd-sotalol. Methods: Chinese hamster ovary cells were transfected with HERG,
encoding IKr,a major repolarizing current of the human
heart and the target of nearly all QT-prolonging drugs, including d-sotalol.
Using the whole cell patch-clamp technique, the modulating effect of glucose on
the IKr-blocking properties of d-sotalol was measured. Results: Both low (1 mM)
and high (20 mM) glucose reduced IKr amplitude when compared to glucose 5 mM (normoglycemia),
at baseline. Moreover, while d-sotalol 25 M blocked IKr by 50±4% (n=5) at
glucose 5 mM, block was 74±2% (n=5) at glucose 1 mM and 65±4% (n=5) at glucose
20 mM. Conclusion: This suggests that low and high blood glucose not only
reduce baseline IKramplitude, but potentiate the IKr-blocking properties of
d-sotalol, further increasing its QT-prolonging effect and thereby increasing
the risk for torsades de pointes. This is a Merck poster. Work also supported
by FRSQ, Québec Heart Institute and HSFC.
Down-regulation of aryl
hydrocarbon receptor-regulated genes by inflammation: the role of nitric oxide
Negar Gharavi and Ayman O.S. El-Kadi, Faculty of Pharmacy and
Pharmaceutical Sciences,
Purpose: We previously demonstrated that tumour necrosis factor- (TNF-)
and lipopolysaccharide (LPS) down-regulate aryl hydrocarbon receptor
(AHR)-regulated genes such as cytochrome P450 1a1 (Cyp1a1) and NADPH: quinone
oxidoreductase (Nqo1) gene expression, yet the mechanisms involved remain
unknown. The correlation between the inflammation-mediated suppression of
AHR-regulated genes and the TNF- or LPS-induced nitric oxide (NO) production
especially in murine hepatoma Hepa1c1c7 cells has been questioned; therefore we
investigated whether NO is involved in the modulation of Cyp1a1 and Nqo1 by
TNF- or LPS in Hepa 1c1c7 cells. Methods: For this purposeHepa1c1c7 cells were
incubated withTNF- or LPS in the absence or presence of the selective inducible
nitric oxide synthase (iNOS) inhibitor, L-N6-(1-iminoethyl) lysine (L-NIL)
(1mM) and peroxynitrite decomposer, iron tetrakis (n-methyl-4'-pyridyl)
porphyrinato (FeTMPyP) (5 M). The amount of NO produced by various
concentrations of TNF- (1, 5, 10 ng/ml) and LPS (1, 5 g/ml) in the absence or
presence of L-NIL were measured by fluorometric HPLC assay. Cyp1a1 and Nqo1
mRNAs were assessed by Northern blot analysis. Cyp1a1 and Nqo1 activities were
measured using 7-ethoxyresorufin and 2,6-dichlorophenolindo-phenol as
substrates, respectively.Results: A significant dose-dependent increase in NO
production was observed by various concentrations of TNF- and LPS which was
completely inhibited by L-NIL. Furthermore, TNF-alpha and LPS significantly
induced iNOS expression in a dose-dependent manner. Both TNF- and LPS strongly
repressed the constitutive expression and the -naphtho-flavone-mediated
induction of Cyp1a1 and Nqo1 at mRNA and activity levels which were
significantly prevented by L-NIL. However, FeTMPyP did not affect TNF- and LPS-mediated
down-regulation of Cyp1a1 and Nqo1 at both mRNA and activity levels.
Conclusion: These results showthat NO, but not peroxynitrite, may be involved
in the down-regulation of AHR-regulated genes mediated by TNF- or LPS.
Acknowledgement: N.G. was supported by Rx & D HRF/CIHR Graduate Research
Scholarship in Pharmacy
Leticia Ely, Raimar Löbenberg,Warren H. Finlay,
Wilson H.-Y. Roa. Faculty of Pharmacy and
Pharmaceutical Sciences; Department of Mechanical Engineering; Department of
Radiation Oncology,
IN VIVO SKIN PERMEATION OF
SODIUM NAPROXEN FORMULATED IN PLURONIC F-127 GELS: EFFECT OF AZONE AND
TRANSCUTOL
José Juan Escobar-Chávez, Miriam López-Cervantes, David
Quintanar-Guerrero, Adriana Ganem-Quintanar, División de Estudios de Posgrado
(Tecnología Farmacéutica), Facultad de Estudios Superiores Cuautitlán –
Universidad Nacional Autónoma de México Cuautitlán Izcalli, Estado de México,
México
PURPOSE: This work focuses on the effects of Azone and Transcutol®®
formulated in PF-127 gels, on the human skin penetration of sodium naproxen in
vivo.METHODS: PF-127 gel formulations were prepared (containing 3 % w/v of
sodium naproxen). Formulation II (F-II) had ~24.8% Transcutol® and Formulation
III (F-III) had a mixture of ~1.7 % Azone®/24.8% Transcutol®. The human
subjects were dosed topically on the ventral forearm. The sodium naproxen
distribution across the stratum corneum (SC) was determined by tape stripping
and quantified by HPTLC. FTIR/ATRandTEWL were used to
evaluate the effect of some of the components of the gel formulations on SC
permeability properties. RESULTS: It was clearly confirmed that the presence of
enhancers promoted sodium naproxen permeation. The combination of Azone and
Transcutol®® (F-III) produced an approximately 2 to 3-fold increase in drug
penetration with respect to F-II, suggesting a synergistic effect between
Azone® and Transcutol®. Treatment of the skin with F-II increased the value of
TEWL to 20.6 ± 1.7 g/h.m2, and with F-III, TEWL reached 33.5 ± 2.6 g/h.m2. A
synergistic effect was proposed for the Azone/Transcutolmixture (F-III). A
significant shift toward a higher wavenumber was observed for the methylene
(CH2) group vibration at 2850 cm-1, only in the case of the Azone/Transcutol®
mixture (F-III).CONCLUSIONS: The experiments demonstrated that the inclusion of
Transcutol®or Azone/Transcutol®® promotes sodium naproxen permeation across the
skin. However, the Azone®/Transcutol combination appears to act
synergistically.
Kinetic Studies of
Pentachlorophenol (PCP) 4-Monooxygenase
Erin Fiege, Rena Okrainetz, Yunyou Su and Jim Fang and Jian Yang, College
of Pharmacy and Nutrition, University of Saskatchewan, Saskatoon, Saskatchewan
Are Steady-State Studies Necessary for Approval of
Generic Modified Release Products?
Anat Fields, Jingsong He, Priya Misra, Yu Chung Tsang; Apotex Inc.,
Purpose: To evaluate the importance of multiple-dose studies for
bioequivalence evaluation of generic modified-release drug products at
steady-state. Methods: Intra-subject variability for AUC and Cmax, as well as
test to reference ratio of geometric means (GMR) for these parameters were
compared in 10 pairs of single-dose and steady-state bioequivalence studies
under fasting conditions. Results: Intra-subject variability at steady-state
was lower than that after a single dose in 9 out of 10 cases for both AUC (mean
17.4% vs. 13.0%) and Cmax (mean 21.4% vs. 15.9%). These differences were
statistically significant for both AUC (p = 0.0068) and Cmax (p=0.0137)
(Wilcoxon Signed Ranks Test). The formulation differences in AUC at steady
state were not different than those after a single dose, while average GMR
values of Cmax appeared closer to 100% at steady state, compared to single
dose. Conclusion: The intra-subject variability is smaller at steady-state than
after a single-dose, suggesting that a single-dose design provides a more
discriminatory condition for assessing the in-vivo performance of two products.
This observation, in addition to the Cmax trend towards smaller formulation
differences at steady-state in comparison to a single-dose, suggests that when
a generic product passes all the bioequivalence criteria in a single-dose study
with no notable formulation differences in pharmacokinetic parameters, a
multiple-dose study at steady-state provides no additional value in
bioequivalence assessment and therefore should not be required for regulatory
approval.
In vitro inhibitory effect of
African medicinal and food plants on human cytochrome P450 3A subfamily.
Amegnona Agbonona,b, Kwashie Eklu-Gadegbeku b,
Kodjo Aklikokoub, Messanvi Gbeassor. b, Vipin D.P. Nairc, Isadore Kanferc, John
Thor Arnasona,Brian C. Fosterd;aDepartment of Biology, University of Ottawa,
Ottawa, Canada; bDepartment of Physiology/Pharmacology, Faculty of Sciences,
University of Lomé, Lomé, Togo;cFaculty of Pharmacy, Rhodes University, South
Africa; dFaculty of Medicine, University of Ottawa and Therapeutics Products
Directorate, Health Canada, Ottawa, Canada
Purpose: African Medicinal plants (AMPs) derived from roots, leaves or
bark are an important component of traditional medicine (TM). These products,
frequently administered by oral route could affect the bioavailability of drugs
or other AMPs. Some of these products are also available as commercially
available formulated products. Hence it is important to test these AMPs as they
are prepared and used in their traditional and commercially available form.
This study was undertaken to evaluate the extracts of AMPs on CYP3A4, CYP3A5
and CYP3A7 metabolic activity in vitro. Methods: The potential for these
botanical species to affect human cytochrome P450 3A-mediated metabolism was
determined using in vitro bioassays with the commercially available microsomes.
Results: The results indicated that extracts of Aframomum cuspidatum,Aframomum melegueta,Harrisonia abyssinica,Hypoxis
rooperi(traditional and commercial forms) and Piper guineense inhibited the
ability of 3A4, 3A5 and 3A7-mediated metabolism. Phyllanthus amarus showed high
inhibition on 3A5 and 3A7. The extracts of Corchorus olitorius,Solanum macrocarpon,Talinum triangulare and Morinda lucida
inhibited CYP3A4 and CYP3A5 less than 20%. The activity of CYP3A7 was inhibited
more than 30% by these same extracts. Conclusion: Frequently traditional
medicines are polyherbal preparations, and it is thought that some of the
plants present in a given preparation are used to increase the effectiveness or
decrease the potential toxicity effect of others. Little scientific evaluation
has been undertaken on these “co-administered medicinal plants”. We conclude
that oral administration of AMPs and particularly the co-administered plants
may alter the disposition of other AMPs and conventional drugs.
Sustained Release Drug
Delivery System for Peptides and Proteins Using Thermo-Responsive polymers
R. Dinarvand, F. Dorkoosh, A, Afshar Ghahremankhani, Pharmaceutics
Department, Faculty of Pharmacy, Tehran University of Medical Science, Tehran,
Iran
Purposes: recently developing of peptide and protein drugs in sustained
release forms widely have been studied. In this work,
studying of sustained release delivery system of calcitonin as a protein with
biodegradable thermo-responsive copolymers based on PL(G)A-PEG-PL(G)A
has been investigated. Copolymers were synthesized by ring opening method.
Materials and Method: The triblock copolymers with different ratio of Glycolide
to D,L-Lactide according to Zenter method with little
change were synthesized. Polymers structure and molecular weight were evaluated
by 400 MHz H NMR and GPC. Purified copolymers were dissolved in acetate buffer
(pH=4) and calcitonin as a model protein was loaded into vial containing
polymer solution. Loaded vial incubate at 37ºC for 2 min and then release media
was added to vial. The released of calcitonin was determined by HPLC method.
Results: Copolymers were characterized by H NMR and GPC. Molecular weight of
copolymers obtained by GPC and calculated from H NMR spectrums were found
5100-7200. Calcitonin release was evaluated for 7-10 days. Although 20±3% of
calcitonin was released in 24 hours but, the release profile was linear up to
80 hours. Calculation shows the mechanism of release is based on diffusion.
Conclusion:thermo-response copolymer may be useful for
short time sustained release of peptides and proteins.
Determination
of fluconazole in human plasma using high performance liquid chromatographic
method.
K.Veeran Gowda, U. Mandal, W.D. Sam Solomon, P. Senthamil Selvan, D.
Senthil Rajan, S. Agarwal, A. Bose, A.K. Sarkar, D. Ghosh, T.K. Pal; *Dept. of
Pharmaceutical Technology, Jadavpur University, Kolkata, India
Purpose: To describe a validated High performance liquid chromatographic
(HPLC) method for the determination of Fluconazole in human plasma. Methods:
Fluconazole in human plasma was extracted using dichloromethane as solvent by
liquid extraction technique. Mobile phase consisting of 10 milli mole phosphate
buffer, methanol and acetonitrile mixture (70:10:20) was used at the flow rate
of 1 ml/min on a C18 column. The eluate was monitored using an UV detector set
at 210 nm. Peak area ratio of the analyte to internal standard (Metoprolol) was
used for the quantification of plasma samples. The method was validated for its
accuracy, precision, linearity, sensitivity and specificity. Results and
conclusion: Theretention time of the analyte and internal standard were 5.7 and
8.1 min. respectively. The validation of the proposed method was carried out. The
method was specific and sensitive with a quantification limit of 0.05μg/ml
and detection limit of 0.02 μg/ml in plasma. The
mean absolute recovery for Fluconazole using the present plasma extraction
procedure was 75.8%. The intra and inter-day coefficient of variation and
percent error values of the assay method were all in the acceptable range.
Calibration curves were linear (r2 > 0.999) from 0.05 to 8
μg/ml. The method was found to be simple, sensitive reproducible
and specific. The suitability of the method was confirmed in the bioequivalence
study of Fluconazole in human volunteers.
The Role of Superantigen
Producing Staphylococcus aureus and Streptococcus in Multiple Sclerosis
M. Namaka1, A. Gomori1, F. Esfahani, L. Wong1, M. Doupe, A. Gupta, Y.
Gong, M. Klowak, T. Du, R. Hizon4, M. Mulvey. Health Sciences Centre,
Department of Neurology, Winnipeg, Canada; University of Manitoba, Faculty of
Pharmacy, Winnipeg, Canada; St. Boniface Research Centre, Department of Family
Medicine, Winnipeg, Canada; National Microbiology Laboratory, Antimicrobial
Resistance and Nosocomial Infections, Winnipeg, Canada
Background: Epidemiologic studies suggest that environmental factors,
such as infectious diseases, may be associated with the underlying pathogenesis
of Multiple Sclerosis (MS). This theory has evolved from the association
between geographical location and prevalence of MS. Henceforth, patients
residing in specific geographical locations may be predisposed to certain
organisms such as S. aureus and/orStreptococcus pyogene that trigger MS.
Purpose:The primary objective of this study is to determine if nasal carriage
rates for S. aureus and/or Streptococcus pyogene correlate with acute
exacerbations of MS. Methods: Study participants (n=240) recruited into the
study will be divided into 3 main groups that include: naïve control (n=80),
active control (n=80) and an acute exacerbation group (n=80). Participants from
each main group will be further subdivided into four subgroups of 20 that correspond
to each of the four seasons. Polymerase chain reactions (PCR) will be used to
determine the toxin genotype for all S. aureus and/or Streptococcusisolates.
Pulsed-field gel electrophoresis (PFGE) will be conducted on all isolates to
determine the molecular relatedness of strains. Results:Nasal carriage rates for S. aureus and/orStreptococcus pyogene are increased in MS
patients compared to controls. Nasal carriage rates of these organisms further
increase during an MS attack. Seasonal variability appears to influence nasal
colonization. Conclusion: The research identifies a novel mechanism by which
antimicrobial treatments may be used as adjunctive therapy with conventional
treatments to reduce MS exacerbations.
Concomitant blockade of
leukotriene B4 and platelet-activating factor receptors underline important
roles of lipid mediators in acute inflammation
Leila Hamdan, Julie
Lefebvre, Pierre Borgeat and Sylvie Marleau. Faculté de pharmacie, Université
de Montréal, Montréal, Canada, Centre de recherche en Rhumatologie et
Immunologie, Centre de recherche du CHUQ(CHUL) and Université Laval, Québec,
Canada
Purpose. A number of studies
reported the involvement of either leukotriene B4 (LTB4) or platelet-activating
factor (PAF) in reperfusion injury. The aim of the present study was to
determine the potentially co-operative effect of these mediators in regulating
polymorphonuclear neutrophils (PMN) trafficking and plasma extravasation in
remote tissues following 2 hours of bilateral hind limb ischemia and 4 hours of
reperfusion. Methods.Rabbits were pre-treated orally with selective BLT1 (BIIL
284, 0.1 mg/kg) and/or PAF (WEB 2086, 10 mg/kg) receptor antagonists. Whole
blood chemiluminescence, a measure of ROS generation, tissue
oedema and myeloperoxidase activity, a marker of PMN accumulation, were assessed. Results. In the lung, PMN accumulation was
reduced by 47 and 36% by BIIL 284 and WEB 2086 compared to vehicle-treated
rabbits, respectively, whereas the inhibitory effect of combined drug
administration was 96 ± 2% (P<0.01). Additive inhibitory effects of combined
PAF and LTB4antagonists administration were observed for lung tissue oedema (53
± 5%, P<0.001) and whole blood ROS generation (65 ± 2%, P<0.01). Similar
inhibition of PMN accumulation and tissue oedema were observed for the liver
and the intestine. Conclusion. Our results support
that LTB4 and PAF play a critical role in the regulation of PMN accumulation at
remote sites following hind limb ischemia and reperfusion. The inhibitory
effect on PMN trafficking is accompanied by a reduced systemic production of
ROS by leukocytes. Supported by the Canadian Institutes of Health Researc
Evidence-Based Review of the
Natural Health Product Hops (Humulus lupulus)
Jaklin Iskander, Heather Boon; Leslie Dan Faculty of Pharmacy,
Purpose: The Canadian public’s interest in complementary and alternative
medicine (CAM) has led to the need for a reliable source of
No Effect of the Flaxseed
Lignan, Secoisolariciresinol Diglucoside, on TTriglyceride Levels in a
Hypertriglyceridemic Rat Model
Jason Jobse, Gloria Woo, Jane Alcorn;
Background: Human clinical studies indicate possible flaxseed effects on
triglyceride homeostasis. The flax component mediating possible changes in
triglyceride levels is unknown. Purpose: Our study’s purpose was to investigate
the effects of the flaxseed lignan, secoisolariciresinol diglucoside (SDG), on
triglyceride homeostasis in a hypertriglyceridemic rat model. Methods: Male
Sprague-Dawley rats (n = 10) fed 10% fructose in water were dosed daily with
SDG at 0 (vehicle), 4.4 or 8.8 mol/kg body weight by oral gavage for two weeks.
An additional rat group (control) was provided tap water and underwent daily
sham (vehicle) oral gavage. Baseline (saphenous venepuncture) and 2 week
(cardiac puncture) blood samples were taken (isoflurane) and analyzed for
triglycerides, phospholipids and non-esterified fatty acids using standard
diagnostic kits. After the 2 week blood sample, rats were killed humanely and
livers and retroperitoneal fat were removed, weighed and stored for analysis.
Real time RT-PCR assays assessed hepatic mRNA levels of two key transcription
factors regulating triglyceride homeostasis, SREBP-1c and PPAR-.Results and
Conclusions: 10% fructose in water significantly increased serum triglyceride
values. In the SDG 0 mol/kg group, fructose decreased hepatic PPAR- mRNA levels
to 66% of control and increased SREBP-1c mRNA levels ~16-fold relative to
control. SDG caused no significant changes in serum and hepatic triglycerides,
serum non-esterified fatty acids and phospholipids, rate of weight gain,
hepatic and retroperitoneal fat tissue weights, and hepatic mRNA levels of
SREBP-1c or PPAR-. In conclusion, SDG had no effect on triglyceride homeostasis
in our rat model of hypertriglyceridemia.
Determination of Hypoxoside
and Quality Control of Commercial Formulations of African Potato (Hypoxis
hemerocallidea) using Capillary Zone Electrophoresis
Vipin. D. P. Nair and I.
Kanfer*, Division of Pharmaceutics, Faculty of Pharmacy,
Purpose: Hypoxoside is a norlignan di-glucoside present in the corms of
African Potato (AP), Hypoxis hemerocallidea, a popular African traditional
medicine used for its nutritional and immune boosting properties. A highly
specific analytical method involving Capillary Zone Electrophoresis (CZE) was
developed for the quantitative analysis of hypoxoside, a major constituent in
AP and this method was subsequently applied for the quality control of
commercially available AP products. This technique has specific advantages over
commonly used techniques such as HPLC, particularly with respect to the use of
relatively non-toxic aqueous buffers thereby obviating the need for more expensive
and relatively toxic HPLC grade organic solvents. Methods: A CZE method was
developed and validated for the determination of the marker compound,
hypoxoside, using a 25 mM sodium tetraborate buffer (pH 9.2). A detection
wavelength of 260 nm was used and samples were loaded hydrodynamically onto an
uncoated fused silica capillary (71cm x 50 m i.d). Sulfafurazole (SF) was used
as an internal standard. Results: Theelectrophoretic separation of hypoxoside
and SF were achieved within 12 min. Linearity of the method was established
throughout the range of 5-60 g/ml and the assay provided a high degree of
accuracy (100 ± 3%). The recovery of the method was found to be 100 ± 5% and
the % RSD of the intraday and interday precision was better than 5.2 and 2.5%
respectively. The limits of detection (LOD) and quantification (LOQ) were
calculated to be 0.5 and 2 g/mlrespectively.Conclusion: This method was used
for the assay and quality control of commercially available products containing
AP. In addition, the method was shown to be stability-indicating, confirmed
from stress testing of hypoxoside.
ANTI-INFLAMMATORY ACTIVITY OF
VARIOUS EXTRACTS OF GARCINIA MANGOSTANA BY INHIBITION OF NITRIC OXIDE
PRODUCTION FROM MOUSE MACROPHAGE RAW 264.7 CELL LINE.
SURESH KUMAR,
Purpose: Inflammation is a body’s response to the damage caused to its
cells by infectious, chemical and physical stimuli. Biological stimuli of
inflammation include endotoxin (Lipo-polysaccride) released by gram-negative
bacteria. The inflammatory cascade initiated by LPS results in release of
various potent inflammatory mediators such as TNF-, IL-1, IL-12 and nitric
oxide by macrophage cells. In our present study we demonstrated the ethyl
extract and acetone extract prepared from pericarp of Garcinia mangostana
showing down regulation of nitric oxide production by murine macrophage RAW
264.7 cell line on stimulation with purified LPS. Methodology: The murine
macrophage 264.7 cell lines was procured from ATTC and cultured by standard
protocol. The nitric oxide estimation was performed by Griess assay. Results:
Our experiment shows concentration ranging from 0.906g/ml to 15.625g/ml of
ethyl acetate extract and 3.906g/ml to 31.125g/ml of acetone extract prepared
from pericarp of Garcinia mangostana significantly inhibited nitric oxide
production by murine macrophage 264.7 cell line when
stimulated by 5g/ml of purified LPS. Conclusion: We conclude from our results
that ethyl extract and acetone extract prepared from the pericarp of Garcinia
mangostana showing potent anti-inflammatory activity by inhibiting the nitric
oxide production which is giving us a lead towards new drug development for
treatment of inflammatory condition. The knowledge of exact mechanism of action
needs further investigation.
Novel use of an in vitro
method to predict the stability of block copolymer based nano-containers
Hamidreza Montazeri Aliabadi, Parvin Mahdipoor, Dion Brocks, and Afsaneh Lavasanifar;
Faculty of Pharmacy and Pharmaceutical Sciences,
Purpose: To design an in vitro experiment that can assess the stability
of polymeric micellar formulations of hydrophobic drugs such as cyclosporine A (CyA).
Methods: Poly(ethylene oxide)-b-poly(-caprolactone) (PEO-b-PCL) blockcopolymers
with respective PEO and PCL molecular weights of 5000 and 13000 g/mol were
assembled to polymeric nano-containers and used for the encapsulation of CyA by
a co-solvent evaporation method. Particles were characterized for their average
diameter and CyA loading efficiency using light scattering and HPLC,
respectively. Measurement of the unbound drug fraction was used to compare the
stability of PEO-b-PCL formulations of CyA with 0.082 (low content, LC) and
0.229 mg/mg (high-content, HC) drug to polymer loading, to that of commercially
available intravenous CyA (Sandimmune®). Blood was collected from
Sprague-Dawley rats by cardiac puncture. Red blood cells (RBC) separated from
rat blood samples were re-suspended in either Sørenson’s phosphate buffer (pH
7.4) or rat plasma and incubated in quadruplicate with the polymeric or
commercial CyA formulations (at a final concentration of 5 μg CyA/mL in
blood) at 37°C for one hour. Samples of whole blood and centrifuged plasma or
buffer were analyzed for CyA by HPLC. The unbound fraction of CyA (fu) was
calculated by equations published by Schuhmacher et al. (J Pharm Sci, 2000, 89:
1008-21). The level of significance was set at p=0.05. Results: The CyA fu as
part of Sandimmune®formulation was 45.6%. The fu of CyA from LC nanocarriers
was significantly lower (36.1%) than that fu observed for Sandimmune®. The HC
nanocarriers had fu of only 10.6%, which was significantly lower than the
unbound fraction for both Sandimmune®and LC nanocarrier groups. Conclusion:
Nanocarriers based on PEO-b-PCL copolymers are capable of changing the protein
binding pattern of CyA. Higher drug contents can stabilize the encapsulation of
CyA in the polymeric nanocarrier further. The plasma protein binding method has
potential utility as a predictor of in vivo micellar stability.
A
Comparison of the Quality of Published Articles Sponsored by Pharmaceutical
Companies to Those Prepared by Independent Research Institutions.
Purpose: To determine whether scientific quality of pharmaceutical
company-sponsored (PCS) articles/scientific meeting abstracts differs from
those sponsored by independent research institutions (non-pharmaceutical
company; NPC).Methods: The quality scores of PCS vs NPC articles was compared
in 27 therapeutic areas (TAs). Scores were obtained through the Thomson Message
Mapping System (TMMS), using independently validated processes for evaluators
to assess strengths/weaknesses of methodology, statistics, results, discussion,
and analysis. The algorithm calculates a quality score for each article.
Comparisons of mean quality scores were made using an unpaired t-test. Results:
11,534 articles/scientific abstracts were analyzed: 4,176 PCS; 460 NPC; and
6,898 no specific sponsorship. There were no differences (N.S.) between mean
quality scores for PCS vs NPC in 20 TAs. A difference
(p<0.05) in favor of PCS articles was observed in 5 TAs
(climacteric/menopause, hepatitis B, osteoarthritis, pertussis, smoking
cessation) and in favor of NPC articles in 2 TAs (ADHD, ulcerative colitis).
Across all 27 TAs a difference (P= 0.0064) was found in favor of PCS vs NPC. A
difference (p= <0.0001) was also found in favor of PCS vs all other
articles. Conclusion: For 74% of TAs, there was no difference between the PCS
and NPC articles with respect to mean quality scores; PCS published information
was of quality similar to that of independent research. Overall, across a broad
range of TAs, PCS sources were generally of superior quality than NPS; these
findings provide a new insight into recent controversies regarding the
potentially incomplete publication of research by pharmaceutical companies.
The Effect of Infliximab on
Hepatic CYP Enzymes and Pharmacokinetics of Verapamil in AdjuvantArthritis Rats
Spencer Ling, Ayman El-Kadi, and Fakhreddin Jamali; Faculty of Pharmacy
and Pharmaceutical Sciences,
Purpose. Clearance of verapamil is
reduced under inflammatory conditions due to increased protein binding and
inhibition of hepatic metabolism. Infliximab reduces pro-inflammatory mediators
and has been shown to reverse the effects of inflammation on sotalol response
in the rat. Since inflammation is also implicated in reduced drug clearance we
hypothesized that infliximab treatment would reverse the effects of inflammation
on drug metabolism and clearance. We examined hepatic cytochrome P450 content
and pharmacokinetics of verapamil in rats treated with infliximab during the
pre-arthritic (pre-AA) phase of adjuvant arthritic (AA) disease. Methods. Pre-AA was induced in male Sprague-Dawley rats with
a tail base injection of M.butyricum. Animals were monitored for symptoms of
arthritis, and levels of the pro-inflammatory mediators serum nitrite and C-reactive protein (CRP). On day 6, rats were administered
single sc dose of infliximab (10 mg/kg). On day 14, a single iv dose of racemic verapamil (2 mg/kg) was administered, and S- and R-verapamil
concentrations were determined by stereospecific HPLC. Hepatic CYP content and
verapamil protein binding were also measured. Results. Serum nitrite levels were significantly elevated in pre-AA and AA phases of
disease. Infliximab treatment did not suppress nitrite levels or reverse the
effects of AA on pharmacokinetic indices. Total CYP and CYP3A contents,
however, were significantly restored in AA rats treated with infliximab. Conclusion. Infliximab partially restores hepatic CYP enzyme
levels, but not sufficiently to reverse the inflammation-induced reductions in
verapamil clearance and free fraction.
Pharmacokinetics and Biodistribution
Profiles of the Micelle Forming Block Copolymer Poly (ethylene glycol)-block-Poly(caprolactone) Following Systemic Administration
Jubo Liu, Faquan Zeng,
Christine Allen. Department of Pharmaceutical Sciences, Leslie Dan
Faculty of Pharmacy; Department of Chemical Engineering and Applied Chemistry,
Faculty of Applied Science and Engineering; Department of Chemistry, Faculty of
Arts and Science, University of Toronto, Toronto, Canada
PURPOSE: At present drugs relying on block copolymer micelles for
formulation are under clinical trial evaluation for the treatment of various
cancers. However, only a few studies have examined the in vivo fate of the
copolymer micelles and unimers. In addition, the effect of the copolymer dose
administered on the in vivo behavior of these systems remains relatively
unexplored. Further in vivo evaluation of copolymer unimers and micelles will
aid in advancing micelles as a more viable drug formulation strategy. METHODS:
Poly (ethylene glycol)-b-poly (caprolactone) (MePEG-b-PCL) copolymers were
synthesized and characterized. The physico-chemical properties of micelles
formed from this series of copolymers were also evaluated. 3H-labeled
MePEG5000-b-PCL5000 micelles were i.v. administered to Balb/C mice at copolymer
doses of 250mg/kg, 2.5mg/kg and 0.2mg /kg in order to examine the distribution
kinetics of 1) copolymer assembled as thermo-dynamically stable micelles 2)
copolymer assembled as thermodynamically unstable micelles and 3) copolymer
single chains, respectively. The biodistribution of the copolymer in the major
organs was investigated and the main pharmacokinetic parameters were
determined. RESULTS: The copolymer assembled as micelles is found to be
effectively trapped within the plasma. The formation of micelles has also been
found to inhibit the cellular uptake of this copolymer in the main elimination
organs. CONCLUSION: The micelle system formed from MePEG5000-b-PCL5000
copolymer was found to be a kinetically stable drug delivery system with a long
circulation lifetime. Therefore, if a drug is well-retained within the micelle
core, this micelle system could serve as a true delivery carrier leading to a
prolonged circulation lifetime for the encapsulated drug.
DISSOLUTION STUDY OF KANAMYCIN
FORMULATED IN A TRANSDERMAL PATCH
Miriam López-Cervantes; José Juan Escobar-Chávez; David
Quintanar-Guerrero; Adriana Ganem-Quintanar; División de Estudios de Posgrado
(Tecnología Farmacéutica), Facultad de Estudios Superiores Cuautitlán –
Universidad Nacional Autónoma de México, Cuautitlán Izcalli, Estado de México,
México
Purpose The present work focuses on the development of a transdermal
patch containing kanamycin, as a complementary treatment for mycetoma due to
Actinomadura madurae. MethodsThe drug was included in an Eudragit-E-100 matrix, forming a patch by pouring a solution in a mold and
evaporating the solvent. Patches were then covered with an occlusive layer of
ethyl cellulose. Triacetin was used as plasticizer. Two types of patches were prepared:
i) Free kanamycin, ii) Kanamycin adsorbed onto silica gel. Patches were
evaluated by their thickness, rupture force, rupture distance, bioadhesivity,
bioadhesivity when dampened, water uptake, gaseous interchange and release rate
studies. Results The composition of the optimal patch
was as follows: Ethyl cellulose 24.4%, Eudragit-E-100 48.8%, triacetin 14.6%,
erythromycin 2.4%, kanamycin adsorbed onto silica gel 9.8%. It was shown to
have the following properties: Thickness 0.574+0.08 mm, rupture force 9314+350,
rupture distance 9.09+1.5 mm, bioadhesivity 86.67+31,bioadhesivity when
dampened 32.8+7.82,water absorption after 2h: 0.029+0.014 g, gaseous
interchange 3.73+2.87g/hmggg2. The dissolution studies indicated that, when
kanamycin was in its free form, 7.2% was released after 24h, and 3.2% when it
was adsorbed (in both cases, the amount was quite enough to produce a 2.3 cm
inhibition diameter in an antibiogram assay). Conclusion The patch described in the results showed good technological properties to be used
as a release system. Permeation studies will be performed with this patch in
healthy skin and in skin affected by Actinomadura madurae.
Cytotoxicity of magnetite
nanoparticles surface-modified with polyethylene glycol triblock copolymers
Framin Mark, Dr. Urs O. Häfeli
Purpose The objective of this study was to investigate the toxicity of
magnetite nanoparticles surface-modified with polyethylene glycol (PEG)
triblock copolymers and of the polymer itself on prostate cancer cells (PC3)
and human umbilical vein endothelial cells (HUVEC). We hypothesized that
magnetite coated with longer tail block lengths would be less toxic. The
overall goal is to use biocompatible and non-toxic magnetic particles as a
potential magnetic drug delivery vehicle for in vivo applications. Methods
Magnetite dispersions were prepared by coating the surfaces of magnetite
nanoparticles with a diameter of 8.8±2.7 nm with a hydrophilic triblock
copolymer having two PEG endblocks and containing controlled concentrations of
carboxylic acid functional groups (PEG-COOH-PEG) in the central segment. An in
vitro 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium
bromide (MTT) assay was used to assess the cell viability. Confocal microscopy
was applied to follow the intracellular fate of magnetic particles at different
time points. ResultsAll pure triblock copolymers as well as the coated
magnetite nanoparticles exhibited concentration dependent toxic effects. The
pure polymers, however, were several folds more toxic than the magnetic
nanoparticles. Within each group, the shorter the PEG tail lengths, the
significantly more toxic they were (15K < 5K < 2K < 0.75K). Confocal
microscopy confirmed that the polymer was more toxic than the magnetite
nanoparticles.Conclusion Magnetic nanospheres coated with PEG triblock
copolymers of 5K and 15K containing 20-50% of iron oxide seem relatively
biocompatible and thus might be useful for magnetic drug delivery. NB. This
project was funded by Merck Frosst to the Summer Student Research Program,
Faculty of Pharmaceutical Sciences, UBC.
A Sensitive
and Specific Liquid Chromatography/Mass Spectrometry Method for Quantitative
Analysis of Cucurbitacin I in Non-Biological Samples and Rat Plasma.
Ommoleila Molavi,AnooshirvanShayeganpour,Vishwa Somayaji, Samar Hamdy,
Dion R. Brocks and John Samuel, Faculty of Pharmacy and Pharmaceutical
Sciences, University of Alberta, Edmonton, Canad
Purpose: to develop a liquid chromatography/mass spectrometry (LC/MS)
method for quantitative analysis of cucurbitacin I, anti-cancer agent that
inhibit JAK2/STAT3 signaling pathway, in non-biological solvents and rat plasma
samples. Methods: Standard samples of cucurbtiacin I were prepared from stock
solution of the compound (1mg/mL) in methanol. 4-hydroxybenzophenone was used
as an internal standard (I.S.). Extraction of cucurbitacin I and I.S. from rat
plasma was performed using acetonitrile/dichloromethane. LC-MS analyses were
performed using Waters Micromass ZQTM4000 spectrometer coupled to Waters
separationsmodule. Chromatographic separation was achieved using C18 3.5
μm (2.1×50 mm) as the stationary phase and a mixture of acetonitrile: 1%
formic acid in water with the ratio of 20:80 and linear gradient to 40:60 for
13 minutes at constant flow rate of 0.2 mL/min as mobile phase. Mass
spectrometer was operated in negative ionization mode and analytes were
quantified with single ion recording (SIR) at m/z 559 for cucurbitacin I and
m/z 196 for I.S. Results: The standard curves over the concentration range of 5-10000
ng/mL for non-biological samples and 10-1000ng/mL for rat plasma samples were
validated, yielding calibration curves with R2> 0.99. Intra- and inter-day
coefficient of variation and mean intraday error were less than 20% at plasma
concentration extending from 10-1000 ng/mL. Conclusion: The developed assay is
sensitive and highly specific for quantitative analysis of cucurbitain I and it
can be used for pharmacokinetics studies.
A validated high-performance
thin-layer chromatographic method for determination of gatifloxacin from
polymeric nanoparticles
Sanjay K. Motwani*, Shruti Chopra, Roop K. Khar, Farhan J. Ahmad, Kanchan
Kohli, S. Talegaonkar; Department of Pharmaceutics, Faculty of Pharmacy, Jamia
Hamdard, New Delhi, India
PURPOSE:Development and validation of an
instrumental high-performance thin layer chromatographic (HPTLC) method for
quantification of Gatifloxacin from biodegradable polymeric nanoparticles.
METHODS:The method employed TLC aluminium plates
pre-coated with silica gel 60F-254 (20 cm x 10 cm with 200 μm thickness, E
Merck,
Preliminary assessment of
interactions between selected alcoholamines and model skin sebum components
Witold Musial, Aleksander Kubis; Drug Form Technology Unit, Department of
Applied Pharmacy,
PURPOSE. The aim was to evaluate
the interactions between the components of model skin sebum and selected
alcoholamines proposed for cleansing activity in the pilosebaceous unit area. MATERIALS AND METHODS. The rate and depth of penetration
into the lipophilic bead imitating pilosebaceous unit lumen was measured using
optical and multiple light scattering analysis devices for alcoholamines
penetration activity assay. The activity differentiation of 0.5% aqueous
alcoholamines solutions with a potential cleansing effect on the pilosebaceous
unit was performed.RESULTS. The depth of aminomethylpropanol penetration
increased from 0.080 mm after 15 min to 3.049 mm after 72 h. The depth of
aminomethylpropendiol penetration increased with time from 0.148 to 4.064
respectively, of diisopropanolamine from 0.481 to 4.626, triethanolamine from
0.236 to 4.342, triisopropanolamine from 0.275 to 2.392 and trometamol from
0.338 to 4.580. The products of alcoholamines reaction with the model skin
sebum components are easily dispersed in water. CONCLUSIONS. The rate of alcoholamines reaction with the model skin sebum depends on the
alcoholamine, being the highest in the case of diisopropanolamine, decreasing
to minimum for triisopropanolamine. Selected alcoholamines would be applied for
in vivo research.
Validation of a liquid
chromatography tandem mass spectrometry assay method for the determination of
nateglinide in human plasma
Adrien Musuku, Gina deBoer, Sarah Bororand,CANTEST BioPharma Services,
Purpose. Development and
validation of an electrospray negative ion LC/MS/MS assay method for the
quantitative determination of nateglinide in human plasma. Methods. Nateglinide and the internal standard (d5-nateglinide) were extracted from 0.1
mL plasma by protein precipitation. Nateglinide was chromatographically
separated on a ProntoSIL120-C18-EPS (4.6 50 mm, 3 m)column using isocratic elution with 20/80 (v/v) 0.1% formic acid/acetonitrile as
mobile phase at a flow rate of 1.0 mL/min. Detection and quantitation were
carried out by ESI-MS/MS monitoring the transitions m/z 316.2 to m/z 164.1
(nateglinide) and m/z 321.0 to m/z 169.0 (d5-nateglinide). Results. The method was validated over a concentration range from 0.10 to 10.00 μg/mL using a linear calibration curve with a
weighing factor of 1/x. Inter-batch precision (%CV) for standards ranged from
1.9 to 2.7. Inter-batch accuracy (%RE) ranged from -2.4 to 1.6,
indicating an acceptable goodness-of-fit. Inter-batch assay precision (%CV) for
quality control samples, based on individual batch means, ranged from 2.6 to
3.8 over four concentration levels, 0.10, 0.30, 4.00 and 9.00 μg/mL. Inter-batch
assay accuracy (%RE) results for quality control samples ranged from -0.6 to
2.6 over four concentration levels. The mean correlation coefficient was
0.9997±0.0002. The mean assay recovery for nateglinide was 79.0±2.8%.
Freeze/thaw stability was established at -40C and -70C for three cycles at each
temperature. Conclusions. An accurate and rapid
analytical assay was developed and successfully applied to the measurement of
nateglinide in human plasma samples.
Qualification of a liquid
chromatography tandem mass spectrometry assay method for the determination of
scopolamine in human plasma
Adrien Musuku, Pricilla Chee, Sarah Bororand, CANTEST BioPharma Services,
Purpose. Qualification of an
LC/MS/MS assay method for the quantitative determination of scopolamine in
human plasma using atmospheric pressure electrospray ionization in positive ion
mode. Methods. Scopolamine and the internal standard
(atropine) were extracted from 0.20 mL human plasma by deproteination followed
by liquid/liquid extraction using chlorobutane as extraction solvent. The
analyte was chromatographically separated on a ACE 3 AQ (4.6 x 50mm, 3.0μm
particle diameter) column using gradient elution with 60% acetonitrile to 40% 1
mM ammonium acetate as initial mobile phase followed by LC/MS/MS analysis.
Detection and quantitation were carried out by ESI-MS/MS monitoring the
transitions m/z 304.2 to m/z 138.1 (scopolamine) and m/z 290.2 to m/z 124.2
(atropine). Results. The method was validated over a
concentration range from 0.05 to 10.00 ng/mL using a linear
calibration curve with a weighting of 1/x. Inter-batch precision (%CV) for
standards ranged from 2.3 to 11.0. Inter-batch accuracy (%RE) ranged
from -5.9 to 4.7, indicating an acceptable goodness-of-fit. Inter-batch assay
precision (%CV) for quality control samples, based on the individual batch
means, ranged from 2.4 to 7.4 over four concentration levels. Inter-batch assay
accuracy (%RE) results for quality control samples ranged from -2.6 to 3.7 over
four concentration levels. The mean (n = 4) correlation coefficient was 0.9984
± 0.0021. In-process stability was established for 10 hours. Conclusions. An accurate, sensitive and rapid analytical assay was developed and
successfully applied to the measurement of scopolamine in human plasma samples.
Validation of a liquid
chromatography tandem mass spectrometry assay method for the determination of
levetiracetam in human plasma
Adrien Musuku, Gina de Boer, Sarah Bonorand, Grace van der Gugten,CANTEST BioPharma Services,
Purpose. Validation
of an LC/MS/MS assay for the quantitative determination of levetiracetam in
human plasma using atmospheric pressure electrospray ionization in positive ion
mode. Methods. Levetiracetam and the internal
standard (d5-levetiracetam) were extracted from 50.00 μL plasma by protein
precipitation. The analyte was chromatographically separated on a Zorbax
Eclipse XDB-C8 (4.6 x 50mm, 3.5μm particle diameter) column using gradient
elution with 95:5 0.1% formic acid in water and 100% methanol (v/v) as initial
mobile phase followed by LC/MS/MS analysis. Detection and quantitation were
carried out by ESI-MS/MS monitoring the transitions m/z 171.0 to 126.0
(levetiracetam) and m/z 176.0 to 131.0 (d5-levetiracetam). Results. The method was validated over a concentration range from 0.10 to 25.00 μg/mL using a linear calibration curve with a
weighing factor of 1/x2. Inter-batch precision (%CV) for standards
ranged from 1.9 to 3.7. Inter-batch accuracy (%RE) ranged from -4.2 to 5.0,
indicating an acceptable goodness-of-fit. Inter-batch assay precision (%CV) for
quality control samples, based on individual batch means, ranged from 1.3 to
2.6 over four concentration levels, 0.10, 0.30, 10.00 and 20.00 μg/mL.
Inter-batch assay accuracy (%RE) results for quality control samples ranged
from -2.9 to 4.2 over four concentration levels. The mean (n=5) correlation
coefficient was 0.9991 ± 0.0002. The mean assay recovery for levetiracetam was
88.8 4.4. Freeze/thaw stability was established at -40C and -70C for three
cycles at each temperature. Conclusions.An accurate and rapid analytical assay
was developed and successfully applied to the measurement of levetiracetam in
human plasma samples.
MICROENCAPSULATION
OF THE SOLID DISPERSIONS OF GRISEOFULVIN—AN APPROACH TO OBTAIN A CONTROLLED AND
COMPLETE RELEASE.
Navin Kumar Satyadas, Narayana Acharyulu, Anitha Dudhani, D
Satyanarayana, Rajesh Dudhani, Nitte Gulabi; Shetty Memorial Institute of
Pharmaceutical Sciences, Mangalore Karnataka, India.
Prajapati V.D.,Zinzuwadia M.M. and Jani
G.K.;Department of Pharmaceutics, L. M. College of Pharmacy, Affiliated by Gujarat
University, Ahmedabad, Navrangpura, Gujarat, India;*Department of
Pharmaceutics, Maliba Pharmacy College, Gopal Vidyanagar, Affiliated by Veer
Narmad South Gujarat University, Tarsadi, Gujarat, India
Purpose: To evaluate the role of Citric acid (CA) and Eudragit E 100 in
delivering multiple coated 5-aminosalicylic acid (5-ASA) tablets intact to the
colon. Methods: Tablets (diameter, 10.5 mm; weight, 300 mg) containing 66.67 %
5–ASA and 10 % CA prepared wet granularly. The aim achieved by imparting compression
coat of HPMC 6 cps and Avicel PH 112 (ratio, 1: 2) on them (diameter, 12.9 mm;
weight, 440 mg) following sequential coat of Eudragit E 100 (diameter, 13.7 mm;
weight, 466 mg), HPMC 6 cps (diameter, 14.3 mm; weight, 485 mg) and Eudragit S
100 (diameter, 15.2 mm; weight, 515 mg) respectively. To mimic gastric,
duodenal, ileac and ascending colon transits of 5-ASA from final coated
tablets, sequential in vitro dissolution (total time, 8 h; lag time, 5 h)
studied using 0.1 N HCl (pH, 1.2) and phosphate buffers (pH, 6.0; 7.2, 6.4)
respectively, colon targeted delivery in vivo by roentgenography. Results:
Tablet’s intactness observed in 0.1
Amphiphilic gels as a
potential carrier for topical delivery in treatment of Psoriasis
Vure Prasad, Rajeev Mishra, P.S.R.Murthy and P. R. Mishra; Pharmaceutics
division, Central Drug Research Institute,
Purpose: In this study an attempt was made to develop Amphiphilic gels as
novel carriers solely consisting of non-ionic surfactants bearing cyclosporine
A, a drug used in the treatment of Postural psoriasis. Amphiphilic gels were
characterized for microstructure, gelation temperature. In-vitro drug release
was performed and suitable formulations were evaluated for antipsoriatic
activity using mouse tail test. Methods: Gels consisting of cyclosporine were
prepared by mixing the solid gelator (Spans) with the liquid phase (liquid
Tweens) and heating them at 60°C to form a clear isotropic sol phase. The sol
phase was cooled to form an opaque semisolid at room temperature. Gel microstructure
was examined by microscopy while the gelation temperature was measured by
differential scanning calorimetry (DSC). In vitro release of amphiphilic gels
were evaluated using rat skin mounted in a Franz diffusion cell and it was
further evaluated in vivo using a mouse tail test. This method uses topical
treatment of a mouse tail with anti-psoriatic drugs to enhance orthokeratotic
cell differentiation in the epidermal scales. This characteristic is used for
direct measurement of drug efficacy in an animal model. Results: Gel
microstructures consisted mainly of clusters of tubules of gelator molecules
that had aggregated upon cooling of the sol phase, formed a 3D network
throughout the continuous phase. At temperatures near the skin surface
temperature, the gels softened considerably. The release studies also supported
drug release from the gel and accumulation in the dermis. This delivery vehicle
of cyclosporine showed morphometric quantification of the conversion of
parakeratotic into orthokeratotic regions in mouse tail scales. Conclusion: The
use of Amphiphilic gels was demonstrated as an ideal vehicle for topical use of
cyclosporine and was corroborated by histological studies in animals.
Cyclodextrin as Enhancemer
for Transdermal Delivery of Rofecoxib
Rawat Swati and Jain Sanjay K.1. Y.B. Chavan College of Pharmacy, Rouza Bagh,
Aurangabad, (M.S.) India, 1Department of Pharmaceutical Sciences, Dr. Harisingh
Gour Vishwavidalaya, Sagar (M.P.), India
Purpose: The purpose of the study was to determine the in vitro as well
in vivo efficiencies of 1% carbopol hydrogel different preparations containing
the rofecoxib (R), a COX–2 inhibitor anti-inflammatory drug with -cyclodextrin
(CD). The release of plain drug was then compared with release of from trans
dermal gel containing physical mixture of drug and CD, inclusion complex of
drug and CD obtained by kneading method and in situ complex of drug and CD
obtained by reacting the drug and CD within the gel. All solid inclusion
complex abstained were then characterized by XRD, IR DSC and SEM, where as the
in situ complex formation was evidenced only by release and permeation studies.
Although many other derivative of CDs such as HP-CD and R-CDare better skin
permeating enhancers but in this study CD was employed because of ease of
availability and low cost. Method: Solid inclusion complex was prepared by
kneading method. Gel formulations were prepared by incorporating pure drug,
physical mixture, inclusion complex and separately drug and CD dispersed in propyleneglycol
(to get in situ complex) to the gel base. Prepared gels were subjected to
physical evaluation for its viscosity, pH and drug content. In vitro drug
release and in vitro drug permeation experiments were carried out on Franz
Diffusion Cell using cellophane membrane and human cadaver skin respectively.
Selected formulations were evaluated for anti-inflammatory activity using the
carrageenin –induced paw oedema in rats. Results: DSC and IR studies indicate
the complexation where as X-RD studies indicate conversion of crystalline drug
to porous, spherical and fluppy structures. The release rates when compared
were found to be highest with gel containing inclusion complex than the gels
containing pure drug, physical mixture and the in situ complex. A lag period
was observed with all formulations. Physical stability was performed by freeze
thaw cycling. The complex containing in situ complex was more stable.
Conclusion: the overall data suggest that the prepared hydrogel of rofecoxib is
highly efficient transdermal vehicle for the delivery of the drug at the site
of action.
Versatile depo-carrier for
controlled protein delivery
Manju rawat,Deependra singh and S.P.Vyas. Department of Pharmaceutical Sciences, Dr. H.S. Gour University,
Sagar (M.P.);
Purpose: Therapeutic peptides and proteins demand an effective delivery
system due to chemical and structural complexities. Methods: In the present
study, IFN- , model protein was stabilized by conjugation with methoxy
-polyethylene glycol (mPEG, MW 5000D) and characterized. In processstability
studies of pegylated IFN - (IFN - - mPEG- 5000) exhibited better stability when
exposed to chloroform: diethyl ether (1: 1 ratio) mixture as well as variable
vortexing time as compared to native IFN – . Pegylated IFN- were formulated as
multivesicular liposomes (MVLs) forutilization as a delivery system for optimum
use. MVLs were prepared by modified reverse phase evaporation method utilizing
double emulsification technique followed by evaporation of organic solvents
from chloroform-ether spherules suspended in water. Three sets of MVLs were
prepared by varying lipid ratio. Formulated PEG-IFN- MVLs was then
characterized for shape, size, vesicle count, encapsulation efficiency and
in-vitro release rate. Results: MVLs prepared were in the size range of 15-20
m. Entrapment efficiencies of three formulations were in the range of 65-78%.
In vitro release profile of IFN - mPEG- 5000 containing MVLs in the PBS
(PH-7.4) showed initial burst release with sustained and nearly complete
release over a period of one week. In contrast plain IFN -mPEG 5000 showed
higher initial burst release i.e. 35% followed by almost complete loss of
protein. Conclusion: Thus, it is evident from this study that MVLs could be a
promising delivery system for controlled delivery of proteins together with
protein modification approach i.e. pegylation.
COMPARATIVE STRUCTURAL AND
FUNGICIDAL STUDIES OF MONO-METHYL PHTHALATE AND ITS TIN(IV)
DERIVATIVES.
Wajid Rehman a, Musa Kaleem Balocha, Amin
Badshah and Saqib Ali. Department of Chemistry, Gomal University,
Dera Ismail Khan, Pakistan; Department of Chemistry, Quaid-e-Azam University,
Islamabad, Pakistan
PURPOSE: The aim of the work is to synthesize, characterize and
investigate the fungicidal properties of some organotin(IV)
compounds with Mono-methyl phthalate. METHODS: The compounds were characterized
by various spectroscopic techniques including 1H-13C-119Sn-NMR, FT-IR and 119Sn
Mössbauer studies. DISSCUSSIONS: On the basis of the spectroscopic techniques
all the complexes show penta coordination with trigonal bipyramidal environment
around the tin. The synthesized compounds were tested against a number of plant
pathogenic fungi. The fungicidal data reveals that the tri-phenyltin(IV)
compound proved to be powerful fungicide. Comparison between the fungicidal
activity of the tri-alkyltin(IV) compounds show that the tri-phenyl tin(IV)
complex is most active against all plant pathogens, rest of the complexes also
exhibit significant antifungal activity but less than the former one.
Growth, Extraction and
Isolation of Novel Jadomycins
Taryn R. Reid, Charles N. Borissow, David L. Jakeman; College of
Pharmacy, Dalhousie University, Halifax, Canada
Purpose: The Jadomycins are modified angucycline antibiotics. Previous
studies of angucycline anticancer antibiotics show activity against human
carcinoma cells. These potential anticancer agents are prepared by fermentation
and are isolated by a series of chemical isolation techniques. The objective of
this research project was to feed two commercially available, non-natural amino
acids (R- and S-phenylglycine) to Streptomyces venezuelae ISP5230, isolate and
purify novel Jadomycins and characterize the compounds using mass spectrometry
and N MR spectroscopy.
Investigation on Niosomes
with Zidovudine as a Carrier for Treating HIV Infection
V. Sankar, K. Ruckmani, R. Saraswathi, M. Ramanathan; Research Scholar, Department
of Pharmaceutical Engineering and Technology, Bharathidasan University,
Tiruchy; PSG College of Pharmacy, Peelamedu Coimbatore, India
PURPOSE: Targeting the drugs to HIV infected cells is an important
challenge today. Zidovudine an anti HIV drug has poor selectivity to
macrophages and manifests dose dependent hematological toxicity. Developing a
site specific formulation can minimize these unwanted effects and dose can also
be reduced. Hence the present study is designed to investigate niosomes for the
transport of antiviral drug Zidovudine. METHODS: Niosomes were prepared by hand
shaking and rotary evaporator method using surfactant: cholesterol in the ratio
1:1, 1:3 and 1:5. Formulations were also prepared including Polyethylene glycol
(PEG). Prepared niosomes were sonicated using Probe Sonicator. Vesicle size,
size distribution was determined using optical microscopy and entrapment
efficiency of drug was determined by solvent method. In vitro release studies
were carried out by dialysis method. RESULTS: Niosomes prepared using rotary
evaporator was reproducible when compared with the niosomes prepared by ether
injection method. Sonicated vesicles are found to be stable when compared with
unsonicated vesicles. The mean size of the vesicles was found to be in the
range of 13- 20, which is the accepted diameter for niosomal injectable.
Increase in the concentration of surfactants increases the drug entrapment by
10%. Similar observations were made with PEG. The drug release from PEG coated
niosomes was slow during the initial hours in case of 1:1 and 1:3 ratio. Eighty
percentage of drug was released over a period of 24 hours which confirms
sustain release. Erratic release was observed from ratio 1:5. CONCLUSION: PEG
coated niosomes containing Zidovudine can be an effective carrier for extending
the life term of HIV infected patients by reducing the severity of infection at
reservoir site.
Development and
characterization of bi-layered multicomponent system of nimesulide and
tizanidine
Swarnlata Saraf , Kamlesh Dashora and S. Saraf
Purpose: The rational fixed dose combination of nimesulide (NIM) and
tizanidine (TIZ) are available in the market for the relief of inflammations,
multiple sclerosis, myofascial pain, neuropathic pain and cerebral spasticity.
The aim of this study was to develop a bi-layered system which is able to
maintain plasma concentration without the need of frequent dosing and less side
effects unlike in case of conventional dosage form and applicable for long term
therapy. Till today, none of the sustained multicomponent formulation of NIM
and TIZ is available in the market. Methods: Bi-layered system composed with
matrix of core NIM and TIZ microparticles with additional immediate release
layer of NIM complexed of beta-cyclodextrin. Various tablet formulation were
prepared with varying concentration of TIZ microparticles and fixed dose of
nimesulide. Physical characteristics and in- vitro release pattern were studied
in alkaline phosphate buffer pH 6.8. Various kinetic models viz., Higuchi and
Korsmeyer-Peppas were applied to know drug release pattern. Results: The
prepared microparticles (ethyl cellulose: drug ratio i.e., 1:1,2:1 and 3:1) were free flowing (angle of repose< 30
degree) with the particle size varying from 215.38±11.52 to 227.36 ±12.89
respectively. The matrix tablet containing 2:1 polymer: drug ratio of TIZ
microparticles and NIM showed parallel release kinetic pattern after 2 hr for
more extended period (beyond 18 h) than conventional tablets. Conclusion: The
corresponding rate constant (K1), regression coefficient (r) and exponent
coefficient (n) of NIM and TIZ were found to be 0.129,0.134,0.9923,
0.9917,0.789 and 0.784 respectively, which indicates anomalous transport and
diffusion controlled mechanism.
Second Derivative
Spectrophotometric Method for the Estimation of Atenolol and Hydrochlorthiazide
in Combined Dosage forms
Swarnlata Saraf, S.Saraf and Gopal Garg; Institute of Pharmacy,
Purpose: The combination of atenolol and hydrochlorthiazide has been
emerged as one of the widely prescribed combination in
single dosage forms as an anti-hypertensive agent. The literature revealed that
no method of simultaneous estimation by uv-spectrophotometer
of both the drugs in tablet dosage forms have been reported. Hence a
simple, rapid, accurate, economical and sensitive second derivative
Spectrophotometric simultaneous method for the determination of atenolol and
hydrochlorthiazide in tablet formulation has been developed. Method: The
Shimadzu Pharmaspec 1700 UV-visible spectrophotometer was used for the
experimental purpose. The absorption maximum was found 274.5 nm and 323 nm
respectively for atenolol and hydrochlorthiazide in 0.1N NaOH. Atenolol shows
zero absorbance at 229.5 nm and hydrochlorthiazide at 234 nm respectively for
second order derivative spectrophotometery.Results: Atenolol obeyed the Beer
Lambert’s law at 234 nm in the concentration range 4-28 g/ml and
hydrochlorthiazide 2-20 g/ml at 229.5 nm. The method was employed for the
estimation of drugs in marketed formulations the result showed the close
proximity to the percentage of label claim (98.95-99.98%). The low value of
standard deviation and relative standard deviation show the accuracy and
precision of the method. The method was validated with the recovery study
(99-101%) and the result show there is no interference with the excipients.
Conclusion: From the above results it can be concluded that the proposed method
is very sensitive and accurate. This method can be employed for the
determination of atenolol and hydrochlorthiazide in combined dosages forms as well
as in bulk drugs.
FORMULATION OF TARGETED
TERBUTALINE SULPHATE MICROCAPSULES OF YEAST FOR ACUTE AND CHRONIC ASTHMA
R. Saraswathi; K.Nagalakshm; P. Priyadharshini; K. Anitha; V.Shankar; K.
N. Krishnan.
PURPOSE: The purpose of the work was to assess the ability of the yeast
cell to act as a microcapsule and to formulate terbutaline sulphate
microcapsule using yeast cells and preparation of dry powder inhalations of
these microcapsules for acute and chronic asthma. Saccharomyces cerevisiae was
the first micro organism chosen for micro encapsulation and its epithelial
adherence is due to the property of being stripped down human cell. METHODS: A.
PRETREATMENT OF YEAST CELLS: A suspension of fresh yeast was treated with over
night with sodium azide(2 gram), a respiratory
inhibitor which is used to prevent the cell from performing any energy
dependent processes. The cells were autoclaved at 121oc for 20 mins. By this
process it was ensured that the yeast cells were made to lose its viability
completely. hence possibility of fungal infection to occur; B. PREPARATION OF
MICROCAPSULES: Drug, yeast and water were taken in the ratio of 1:2:4 and
agitated with a magnetic stirrer for 4 hours at a temperature of 400 c. the
cells were then centrifuged for 10 mins. The supernatant solution was decanted
and the cells were washed 5 times with distilled water and freeze dried for 48
hrs. The above procedure was repeated with 0.2 gm, 0.4 gm of the same drug with
yeast and water in the same ratio of 1:2:4 at a temperature of 350c, 300c and
250c. a total of 12 samples were done using this
process. RESULTS: The freeze dried samples of micro encapsulated drug with
yeast cells were evaluated with parameters such as: yield, entrapment
efficiency was analyzed using UV spectra at 295 nm; photographs of micro
encapsulated drug using confocal microscope. Among the 12 samples the 6th
sample constituted with 0.2 gm of drug, 0.4 gm of yeast and 6 ml of water
maintained at 350c showed higher encapsulation yield of 14 micrograms. The
release of the drug from the microcapsules showed first order kinetics
sustained action. The dry powder mixture of the encapsulated yeast with spray
dried lactose was prepared and powder characteristics studies are being
undertaken.CONCLUSION: The present study envisages the epithelial adherence
property of the yeast cells and its drug encapsulation capacity. So the formulated
dry powder inhalation expected to be very effective, targeting the inflamed
bronchial epithelial cells. The optimization of the microcapsules to diluent
ratio in the dry powder inhalation is being carried out using invitro models.
Exploitation of Some
Traditional Plant Drugs for Anti-fertility Activity
S.K. Sharma, Neeru Vasudeva; Division of Pharmacognosy, Faculty of
Pharmaceutical Sciences,
Purpose: One approach pursued to identify new anti-fertility agents is
the search of their presence in natural sources; for which we have made an
attempt to scientifically authenticate the traditional use of some
anti-fertility plant drugs namely,Achyranthes aspera
Linn. (Amaranthaceae) roots, Daucus carota Linn. (Umbelliferae)
seeds and Hibiscus rosa-sinensisLinn. (Malvaceae) roots were selected
for their scientific authentication of their traditional use. Methods:The coarsely powdered drugs were extracted with ethanol
(95%) by hot continuous extraction method and concentrated. The extracts were
subjected to post-coital anti-fertility studies and estrogenic and
anti-estrogenic studies. Post-coital anti-fertility studies: the extracts were
administered at two different doses 200mg/kg and 400mg/kg to the female albino
rats in the proesterous stage from day one to 7 of pregnancy. The number of
implantation sites was counted on day 10. Estrogenic and anti-estrogenic
activity: colony-bred immature female albino rats, 21-23 days old, weighing
between 35 and 45 g, were selected for this activity. The extracts were
administered at the dose of 400 mg/kg for 7 days. On day 8 the uteri were
dissected out and weighed. Histological studies of the dissected were also
performed. Results:The ethanol extracts ofDaucus
carota Linn. seeds and Hibiscus rosa-sinensis Linn. roots showed significant anti-implantation activity at the
dose of 400 mg/kg body weight and Achyranthes asperaLinn. roots showed significant anti-implantation activity at the dose of 200 mg/kg body
weight. All the three extracts acted as weak estrogens. The ethanol
extract ofDaucus carota Linn. seeds when given
along with ethinyl estradiol acted as anti-estrogenic and the ethanol extracts
of Achyranthes asperaLinn. and Hibiscus rosa-sinensis
Linn. roots potentiated the effect of ethinyl
estradiol. Conclusion:The three plant drugs have weak
estrogenic activity thus authenticating traditional use.
A Study of the Antifungal and
Antibacterial Activity of Some Essential Oils
Sumitra Singh and Surendra K Sharma; Pharmacognosy Discipline, Faculty of
Pharmaceutical Sciences,
Purpose: Essential oils have been used since ancient times to alleviate
various ailments like flatulence and colic discomfort, as appetizers, and as perfumery.
Very recently their use has been exploited as an antiseptic, stimulant,
expectorant, diuretic, etc. Today indiscriminate use of antibiotics has led to
resistance of microbes, hence attention is being given
to plant derived antimicrobials. The present study is aimed to evaluate the
antifungal and antibacterial properties that inhibit or kill resistant
organisms. Methods: The essential oils were obtained from the leaves of
Cymbopogan nardusLinn./Rendle.(Gramineae), Mentha arvensis Linn.(Labiateae), Mentha
spicata Linn.(Labiateae), Seeds of Anethum sowa Kurz.(Umbelliferae), fresh
pericarp of Citrus limon Linn./Burn.f. (Rutaceae) by water steam distillation
and dried over anhydrous sodium sulphate. The antimicrobial activity was tested
by agar diffusion method by measuring zone of inhibition and minimum inhibitory
concentration by turbidity method using Spectrophotometer at max530 nm.
Results: All the essential oils in pure form showed antibacterial activity
against Escherichia coli NCIM 2065, Staphyloccocus aureus NCIM 2901, S.
aureusNCIM 2079, Bacillus cereus NCIM 2322, B. cereusNCIM 2106, B. coagulans
NCIM 2030 and Proteus mirabilis NCIM 2241 and antifungal activity against
Candida albicans ATCC 10231, Aspergillus
Synthesis and antimicrobial
activity of a new series of N1-substituted 1H-indazol-3-yl-acetic acid
D.G. Dalvi1, Y.V. Pore1, B.S. Kuchekar1, S.B. Bhise1, A.A. Shingavi. 1.
Purpose: Indazole derivatives possess a wide range of pharmacological
activities like anti- inflammatory, antimicrobial, aldose reductase inhibitor,
nitrous oxide inhibitor. In order to obtain new potent therapeutic agent we
have synthesized a series of indazole derivatives containing substituted
anilino methyl group at 1 position.Method: The titled compounds were
synthesized by the reaction of malonic acid, formic acid, ammonium formate, and
2-nitro benzaldehyde to obtain the 3-amino-3- (2-nitrophenyl) propionic acid
derivative (1). This was further reacted with hydrazine hydrate at 80oC using
raney nickel as catalyst to yield indazole 3-acetic acid (2). Various
substituted anilines were treated with indazole 3-acetic acid in presence of
formaldehyde, dioxan, and hydrochloric acid to give N1substituted indazole 3-acetic
acid (3a-3l). Scheme: All these compounds were recrystallized and obtained in
satisfactory yield. Structures of synthesized compounds were confirmed by
spectral analysis. Melting points were taken in open capillary tube and are
uncorrected. Reaction was monitored by TLC. Results: All the compounds (3a-3l)
were screened for antimicrobial activity against E.coli (EC), S.aureus (SA),
B.substilis (BS), S.pyrogenes (SP)’ K.pneumoia (KP) and P.vulgaris (PV) by cup
plate method. After 24 hr of incubation at 37 0Cthe zone of inhibition were
measured in mm. The activity was compared with the standard antibiotic. Results
are expressed in tabular form. Figures indicate zone of inhibition in mm. Conclusion:
The data expressed in tabular form indicates that the compounds (3c, 3f, 3g,
3h, 3i, 3j, 3k) have shown good antimicrobial activity against the
microorganisms. These compounds are found to be more effective against E.coli
and 3j was found to be effective against B.substilis,comparable to standard antibiotic. From these results, it can be concluded that the
electron withdrawing substituents on benzene ring of aniline moiety at N1
influences the antimicrobial activity. More number of compounds are necessary to be synthesized and their structure activity
relationship is required to be studied.
Preparation and
Characterization of Collagen based dual delivery system for effective wound
healing.
Deependra Singh, Swarnlata Saraf, S.Saraf;
Purpose: Wound is a pathological condition involving disruption of normal
anatomical structures and function. We aimed to develop biocompatible collagen
based delivery system to hasten and effectively facilitate the wound healing process.
Method: Combined delivery system was prepared by loading alginate microspheres
of proteolytic enzyme (serratiopeptidase) on gentamicin impregnated collagen
(GIC) sheet. GIC sheet was prepared by soaking the collagen sheet in (10%)gentamicin solution. The serratiopeptidase loaded alginate
microspheres were prepared by internal gelation method. Combined system was
prepared by patting the serratiopeptidase microspheres on the application
surface of GIC sheet. In vivoperformance of combined delivery system was
evaluated on albino rats in terms of physical, histological, cytological and
biochemical assessment of wound healing. Results: Optimized serratiopeptidase
alginate microspheres exhibited particle size in the range of 70-75m with
around 80 % loading. Microspheres showed initial burst release of 35% in first
two hours followed by extended 83% release in 72 hours. GIC sheet retained
substantial antimicrobial activity when tested for effectiveness on different
strains of bacteria by agar diffusion method. Animal studies showed well-formed
granulation tissue by day 7. Comparatively significant increase in percent
wound reduction, protein content and Hydroxyproline content was observed
(P<0.001). Histological studies further supported effective healing by
increase in neutrophils along with proliferating fibroblasts and macrophages.
Conclusion: The prepared biocompatible dual delivery system can prove to be an
effective system for rapid wound repair in terms of better tissue debridement,
neovascularization, increased chemotaxis for fibroblasts and macrophages,
removal of microbes from wound site and effective contraction.
Are the Current
Bioequivalence (BE) Requirements Unnecessarily Stringent for the Approval of
Generic Proton Pump Inhibitor (PPI) Products?
Yu Chung Tsang. Apotex Inc.,
Purpose. All PPIs are commercially
available as enteric coated drug products in
Decreased expression of the
low density lipoprotein receptor (LDLr) in human embryonic kidney cells using
RNA interference
Carlos Leon, Guosong Qiu, Hana Kolac, John S.
Hill and Kishor M. Wasan. Division of Pharmaceutics and
Biopharmaceutics, Faculty of Pharmaceutical Sciences, University of British
Columbia, Vancouver, Canada; CAPTURE Centre, St. Paul’s Hospital, University of
British Columbia
Purpose: To develop an RNA interference approach
to down regulate LDLr expression within a human embryonic kidney cell line
(293T). Materials and methods: siRNA cloning: To generate the constructs, two
pairs of complementary oligonucleotides, were annealed (LDLr-792 and LDLr-973).
They were cloned into the pSHAG vector that directs the in vivo synthesis of
siRNA. Positive clones acquired both kanamycin resistance and a new restriction
site. An annexin V construct (AxV) was used as a siRNA control. siRNA transfection: 293T cells cultured in complete DMEM
were seeded in pre-coated (Poly Lysine) plates to enhance adherence. Three days
after transfection, the cells were washed and frozen. Western blot and RT-PCR:
Cells were lysed in RIPA buffer, protein was quantified and the lysates were
resolved by SDS-PAGE and transferred to nitrocellulose. Immunoblotting was
carried out with commercial anti LDLr, SR-BI, annexin V and actin antibodies.
RNA was isolated to prepare cDNA. Real time RT-PCR was performed to amplify the
LDLr and GAPDH genes. Results: When compared with transfected control cells,
the LDLr-792 and LDLr-973 constructs were associated with a reduction in LDLr
protein expression of 70% and 50%, respectively. Interestingly, the cells
transfected with AxV showed a higher LDLr protein expression, while annexin V
expression was reduced by 70%. No differences were observed in SR-BI expression
consistent with the specificity of the down-regulation effect. Quantitation of
the LDLr mRNA by real time RT-PCR in LDLr-792 transfected cells indicated a
reduction of 60% compared to control cells consistent with the immunoblot
results. Conclusions: We have developed a tool to decrease LDLr expression in
multiple cell lines. We will use this model to test the role of the LDLr in the
internalization of drugs that interact with LDL. Acknowledgements: Funding was
provided by a grant from CIHR to K.M.W and a grant from the
Influence
of Lipid Excipients, Capryol PGMC and Gelucire 44/14 on P-glycoprotein (P-gp)
activity in Human
Andrea Thamboo, Kristina Sachs-Barrable, Stephen D. Lee, and Kishor M.
Wasan, Faculty of Pharmaceutical Sciences, University of
Purpose: The objective of this study was to determine the influence of
lipid excipients, Capryol PGMC and Gelucire 44/14, on P-gp activity in Caco-2
cells. Methods: To determine non-toxic concentrations for Capryol PGMC and
Gelucire 44/14 on Caco-2 cells, LDH, MTS and BCA assays were performed to
determine cell plasma membrane integrity, mitochondrial respiration, and total
protein concentration respectively. For P-gp efflux experiments, Caco-2 cells
were seeded into 12-well plates. After cells reached 90% confluency, they were
incubated overnight with previously determined non-toxic excipient
concentrations and HBSS (control). The following day, a final concentration of
5M RH-123 was added to each well and incubated for 3 hours. Plates were then
washed with PBS and HBSS was added. RH-123 efflux was measured after certain
time points. Cells were lysed with 1% Triton X-100 and RH-123 and BCA protein
was determined. For Transwell studies, cells were incubated overnight with
excipients on apical and basolateral sides. A final concentration of 5M RH-123
was added only onto the basolateral side and RH-123 efflux (basolateral to
apical transport) was measured after certain time points. At the end of the
experiment, RH-123 content and BCA total protein in lysed cells were measured.
The Dunnett test was used to measure statistical significance. Results: LDH,MTS
and BCA (N=6) assays all correlated, showing that concentrations 0.1 v/v% Capryol
PGMC and 0.02 w/v% Gelucire 44/14 are non-toxic to Caco-2 cells. Cells treated
with 0.02 w/v% Gelucire 44/14 showed significant reduction (p<0.001) in the
percent of RH-123 effluxed compared to the control, whereas lower
concentrations for Gelucire 44/14 showed no significant reduction. Capryol PGMC
showed no statistical difference between normalization of percentage RH-123
effluxed from Caco-2 cells treated with 0.1, 0.05 or 0.025 v/v% Capryol PGMC to
the control (HBSS). In the Transwell studies (basolateral to apical transport),
0.1, 0.05 and 0.025v/v% Capryol PGMC and 0.02 and 0.01w/v% Gelucire 44/14
showed significant increase (P< 0.01, 0.01, 0.05, 0.01 and 0.01
respectively) in the ratio of absorbance of RH-123 to milligrams of protein
compared to the control.Conclusion: Our findings suggest that Caco-2 cells
treated with lipid excipients, Capryol PGMC or Gelucire 44/14 at non-toxic
concentrations may decrease the amount of RH-123 effluxed compared to untreated
control cells, suggesting a reduction in P-gp activity. Acknowledgments: This
project was funded by a Canadian Institute of Health Research Operating grant
to KMW. A portion of this work was published at the 2005 AAPS Annual Meeting
held in
COMPARISON OF PHYSICOCHEMICAL
DATA VS DISSOLUTION DATA TO ESTABLISH IN VITRO/IN VIVO CORRELATIONS
Hai Weil, Izzy Kanfer, Marie
Di Maso, and Raimar Loebenberg. Faculty of Pharmacy and
Pharmaceutical Sciences, University of Alberta, Edmonton, Canada; Faculty of
Pharmacy, Rhodes University, South Africa; 3Pharmaceutical Research &
Development group, Merck Frosst, Canada
Purpose: To differentiate the physicochemical properties of different
glyburide powders from different sources using material characterization
methods. To predict the oral absorption of these powders using physicochemical
data or dissolution data as input functions into the advanced compartmental
absorption and transit model (ACAT). Methods: The material characterizations
include SEM, X-Ray, Raman Spectrum, DSC, Particle Size Distribution Analysis,
Surface Area and True Density. Solubility of glyburide was determined in
different dissolution media at different pH values. The dissolution behaviors
of two glyburide formulations (3.5 mg and 5 mg) were tested using apparatus 2,
USP 28. The dissolution tests were performed using a multi-pH gradient method.
The prediction of the fraction dose absorbed for each formulation was performed
using GastroPlusTM. The simulations were compared with clinical data. Results:
The crystal forms of the different glyburide powders were similar; however,
significant differences in morphology, surface area and particle size were
determined. The solubility of the glyburide was pH-dependent. The particle size
had significant influence on the simulations when only the solubility data were
used. Both physicochemical and dissolution data could be used to successfully
predict two 3.5 mg formulations. For a 5 mg formulation only the
physicochemical data were able to predict the oral absorption. This was due to
incomplete dissolution of this product of about 60%. Conclusion:
Physicochemical data can be used to predict the oral absorption of glyburide.
Dissolution data can only be used as input function if the in vitro dissolution
reflects the in vivo dissolution.
Structural Similarity between
Human Bitter Taste Receptors and Histamine H1-Receptor
Jian Yang and Jim Fang;
Purpose Bitter taste is the self-protection mechanism against poisonous
substances evolved in mammals. In human, more than thirty bitter taste
receptors (T2Rs) have been identified. Use of bitter substances to relieve
inflammation-like symptoms has been used in traditional Chinese medicine. In the
current study, we investigated the potential relationship between bitter taste
and anti-inflammatory activity for H1-antihistamines. Methods
Thethree-dimensional models of representative human bitter taste receptors
(T2R5, T2R14, T2R16, T2R43 and T2R61) and histamine H1-receptor were built by
homology modeling method using program MODELLER. H1-antihistamines were docked
into the ligand binding sites of both the bitter taste receptors and the
histamine H1-receptor by the docking software AUTODOCK. ResultsThe bitter taste
receptors exhibit very high structural similarities to the histamine
H1-receptor. The root-mean-square deviations among the bitter taste receptors
and between the bitter taste receptors and H1-receptor are less than 1.5 Å. A
hydrophobic binding pocket similar to the H1-receptor ligand binding pocket is
present in the bitter taste receptors. However, two basic residues Lys76 and
Lys78, which can interact with polar functions groups of substrates, are
adjacent to the hydrophobic binding pocket, implicating the bitter taste
receptors may have a broader substrate spectrum than the histamine
H1-receptor.Conclusion This study suggests that
H1-antihistamines are likely to bind to the bitter taste receptors.
Hemodynamic effects of
diltiazem in different rat models following repeated subcutaneous injections in
vivo
Pollen K.F. Yeung, Angelita Alcos, Jinglan Tang, William L. Casley. Pharmacokinetics
& Metabolism Laboratory, College of Pharmacy, Dalhousie University,
Halifax, Canada; Centrefor Biologics Research, Biologics and Genetic Therapies
Directorate, Health Products and Food Branch, Health Canada, Banting Research
Centre, Ottawa, Canada
Purpose: To compare the hemodynamic effect of diltiazem in different rat
models following repeated subcutaneous injection. Methods: MaleSD, SHR, and WKY
rats (Charles River Laboratories, n = 6 – 10 per group) weighing between 300 -
450 g were used. Each rat received either saline (control) or 5 mg/kg of
diltiazem
A sensitive and specific HPLC
assay of cladribine for pharmacokinetic studies in rats
Pollen K.F. Yeung,Ameer Jaraar,Carrie Ferguson, Soulatchana Narayanan;
Pharmacokinetics and Metabolism Laboratory, College of Pharmacy, Faculty of
Health Professions, Dalhousie University, Halifax, Canada
Background and Purpose: Claribine is a prototype of the nucleoside
anticancer agents. To develop a sensitive and specific HPLC assay for
cladribine in plasma for pharmacokinetic studies. Methods: Cladribine and the
internal standard AZT were purchased from Sigma-Aldrich Chem. The HPLC system
consisted of a Shimadzu LC-9A pump, a 3 m,250 x 2.0 I.D. high speed C18 column
(Jupiter), preceded by a 5 m 4 x 4 mm i.d. C18 guard column (Licrocart®), an
Agilent Model 1050 UV-VIS detector and a 3395 Integrator. The mobile phase was
made up of 0.01M pH5 KH2PO4: methanol: acetonitrile (90:5:5). The system was
operated at ambient temperature with a flow rate of 0.3 mL/min, and UV
wavelength at 265 nm, and an operating pressure of ca. 1.56 kpsi. Extraction of
cladribine and AZT from plasma was achieved by solid phase extraction using 100
mg/mL C18 SPE columns (Extra-sep). The assay was validated for sensitivity,
precision, specificity and application for pharmacokinetic study in rats.
Results: Under these conditions, the average retention times of cladribine and
AZT were 13.5 and 21 min, respectively, and recoveries were > 80%. Standard
curves based on absolute on column injection of each compound were linear from
2.5 to 15 ng, with regression coefficient (r2) 0.99 or greater. Sensitivities
based on absolute injection were < 1 ng on column. Using a 50 uL plasma
sample size, the intra-assay variations at 0.1 ug/mL were 7%, and inter-assay variations over a period of 3 months were 17%. The
assay was used to study a single dose pharmacokinetic study of cladribine in
rats after a 2 mg/kg dose. Conclusions: The described HPLC assay has adequate
sensitivity and specificity to study pharmacokinetics of cladribine in rats
(Supported in part by a Nova Scotia Health Research Foundation Innovation Grant
and a Dalhousie Science Co-Op Program student scholarship to Ameer Jaraar).
Design, Synthesis and
Anticonvulsant Activity of New 1,3,4-Oxadiazole
Derivatives as Benzodiazepine Receptor Agonists
Afshin Zarghi *, Avideh Ahadian, and Hamid R. Khojastehpoor; Department
of Medicinal Chemistry, Faculty of Pharmacy,
PURPOSE: A series of new 2-substituted-5-(2-benzylthio or
benzyloxyphenyl)-1,3,4-oxadiazoles was designed and
synthesized as anticonvulsant agents . METHOD: In order to synthesis of
compounds 2-benzylthio or benzyloxyphenyl acid hydrazides were converted to
2-amino-5-(2-benzylthio or benzyloxyphenyl)-1,3,4-oxadiazoles using cyanogen
bromide in methanol (75-84%).Conformational analysis of the 2-amino-5-[2-(p-fluoro)-benzylthio
or benzyloxyphenyl)- 1,3,4-oxadiazole and estazolam was preliminarily performed
by MMX force field method implemented in PCMODEL 6.0 software. The conformers
were optimized further by AM1 calculation using MOPAC 6.0 program.Global energy
minima conformers of the designed compounds were superimposed on corresponding
conformer of estazolam molecule, which was considered as a reference BZD
agonist. The compounds were characterized by 1H nuclear magnetic resonance,
infrared, mass spectrometry and CHN analysis. The BZD activity of the
synthesized compounds was determined through the evaluation of the ability of
the compounds to protect mice against convulsion induced by a lethal dose of
PTZ and electroshock as two routine models. RESULT:Most of the synthesized compounds showed anticonvulsant activity in both models. The
benzyloxy phenyl -1,3,4-oxadiazoles had more
anticonvulsant activity in compared with benzylthiophenyl derivatives. The
fluoro substituent at para position of benzyloxy or benzylthio moiety gave the
most active analogue in both series of compounds. CONCLUSION: Some
2-amino-5-aryl- 1,3,4-oxadiazoles with a simple
non-rigid structure in which the flexible second out-of-plane aromatic ring,
benzylthio or benzyloxy group, has a suitable substituent could show
benzodiazepine activity.
A Validated New UV
Spectrophotometric Method for Determination of Ascorbic Acid in Its
Effervescent Dosage Forms
Wenming Zeng, Frank Martinuzzi, and Alexander MacGregor; Department of
Research and Development, Toronto Institute of Pharmaceutical Technology,
Purpose. The objective of this
work was to develop and validate a new UV spectrophotometric method for
determination of ascorbic acid (vitamin C) in its effervescent dosage forms. Methods. Ascorbic acid was first found to dissolve in
methanol, and its solubility in it was measured to be 81.0 mg/ml at room
temperature (22oC). The stability of ascorbic acid in mehanol at the room
temperature was determined to be only 0.7% of oxidation within 1 hour compared
with 46.0% of oxidation at the same period in de-ionized water. Ascorbic acid
was found to have a maximum wavelength of 245 nm in methanol. Then, methanol
was used to prepare analytical samples of effervescent vitamin C tablets for
the UV determination of ascorbic acid, finding no interferences in the UV
region from other substances because of their insolubility in methanol. Results. The analytical curve is linear (R2 = 0.9997) in
range from 0.01 to 0.03 mg/ml. The recovery of ascorbic acid ranges from 98.3
to 101.4%. The method repeatability test results meet the relevant acceptance
criteria (RSD < 2%). The analytical results from different operators in
different days are in good agreement. This method is specific for ascorbic
acid. The dilution factors and the small change in the sonication time have no
any effects on the recovery of ascorbic acid. The analyte stablity in sample
preparation is also validated. Conclusions. The
validated method can be used for the routine determination of ascorbic acid for
the dosage uniformity testing of effervescent vitamin C tablets with various
strengths. This method is rapid, accurate and reliable, and methanol is not
expensive and easily available, which can save much time and money for the
manufacturers who produce the effevescent vitamin C products.
Tablet Formulation
Development for a Poorly Water Soluble New Chemical
Entity
Kai Zhang, AndreaToth, Kay Koch-Gaynor, Wlodek Karolak, Myrna Dela-Cruz,
David Valentini, Phyllis Dawson, Mehran Maleki; GlaxoSmithKline, Pharmaceutical
Development, Mississauga, Canada
Purpose: To develop a tablet formulation using a conventional wet
granulation method for new chemical entity X (NCE X), a BCS Class II compound
(low aqueous solubility < 0.1 g/mland high permeability). Only micronized
NCE X with D (v,50) of 1.7 m was used. Methods: A HPLC
method was used for solubilizer screen. A conventional wet granulation method
was employed for tablet manufacturing. All excipients used were common
materials available commercially. Dissolution profiles of tablets in simulated
intestinal fluid (SIF) were assessed for formulation optimization. Results:Among all the investigated solubilizers, sodium dodecyl
sulfate (SDS) emerged as the most efficient candidate for improving NCE X’s
aqueous solubility. Formulations incorporated with SDS confirmed this finding.
In addition, tablet dissolution profiles also indicated povidone (PVP) acting
as a possible solubility enhancer. Consequently, studies by Design of
Experiments (DOE) were performed to investigate three possible factors (NCE X,
SDS, and PVP) at multiple levels for formulation optimization. In all the
formulations investigated, Formulation F14 was the best in terms of both
dissolution rate and equilibrium concentration, whose dissolution profile was
better than that from a gelucire capsule formulation with proven sufficient
exposure in dog. A DMPK study in dog evaluating F14 tablet formulation against
the gelucire capsule formulation indicated that the exposure for the F14 Tablet
formulation was not significantly different from that obtained using the
gelucire capsule formulation. Conclusions: A tablet formulation was developed
successfully with sufficient exposure in dog. The tablets were developed with
common excipients and manufactured by a conventional wet granulation method.
Dendritic cell targeting of
MUC-1 breast cancer peptide expressed on bacterial surface
Jany H. Zhang, Pravin K. Bhatnagar, Welson W. Wang, John F. Nomellini,
John Smit, and Mavanur R. Suresh; Faculty of Pharmacy and Pharmaceutical
Sciences, University of Alberta, Edmonton, Canada and *Department of
Microbiology and Immunology, University of British Columbia, Vancouver, British
Columbia, Canada
Purpose: Whole bacterial cells can stimulate a T cell-mediated response;
therefore, a bacterial formulation of Caulobacter crescentus displaying both
MUC-1 breast cancer peptide and a Protein G binding domain was developed for
targeting to dendritic cells (DCs). Since DCs are the most potent antigen
processing cells, a specific antibody targeting system could enhance and/or
stimulate an active specific immune response against cancer. Methods:Anti-DEC-205 antibody was produced by HB290 cells, purified
on a Protein G column, and biotinylated. The C. crescentus bacteria constructs
were grown in PYE media and characterized for MUC-1 and the Protein G domain by
direct whole cell ELISA. Results: The ELISA results detected that the activity
of both MUC-1 and the Protein G binding domain of the 4x MUC-1 construct was
intense. In contrast, very little activity was detected for either domain of
the 1x MUC-1 construct. Although the 2x MUC-1 construct had strong activity for
the Protein G domain, there was weak reactivity for MUC-1. A difference in
activity was detected when the concentrations of anti-MUC-1 and anti-DEC-205
were varied for the 4x MUC-1 construct. The preliminary simulated in vivo study
demonstrated that anti-DEC-205 bound to the Protein G domain of the 4x MUC-1 construct was minimally displaced when other antibodies were
added. Conclusion: The Protein G binding domain of the 4x MUC-1 C. crescentus
construct could be used via an antibody-based targeting system, to present the
MUC-1 breast cancer antigen or any other antigen integrated into the bacterial
S-layer, to DCs.
Non-invasive Assessment of
the In Vivo Pharmacokinetics of Liposomes Using CT and MR Imaging
Jinzi Zheng, Mike Dunne, David A. Jaffray, Christine Allen2; Department
of Medical Biophysics, University of Toronto, Toronto, Ontario, Canada; Faculty
of Pharmacy, University of Toronto, Toronto, Ontario, Canada; Departmentof
Radiation Physics, Princess Margaret Hospital, University Health Network,
Toronto, Ontario, Canada; Department of Radiation Oncology, University of
Toronto, Toronto, Ontario, Canada.
Purpose: Traditional evaluation of the pharmacokinetics of drugs and
materials involves invasive tissue and plasma sampling. These methods require
the use of large numbers of animals which introduces animal-to-animal
variability and limits the number of time points that may be sampled. The
present study is aimed to explore the use of non-invasive techniques such as
computed tomography (CT) and magnetic resonance (MR) imaging to effectively
track the in vivo circulation pathway of contrast agent modified lipid carriers
in a rabbit model. Methods: A Female New Zealand White rabbit was administered
liposomes encapsulating 200 mg/kg iodine and 16 mg/kg gadolinium. The rabbit
was imaged in CT and MR at specific time points (up to 7 days) following
liposome injection. The relative signal intensity increases measured in the two
imaging modalities at the rabbit aorta were correlated with the iodine and
gadolinium concentrations present in blood as measured by HPLC and ICP-AES
analyses. Results: A linear concentration prediction range was found in both CT
and MR with correlation coefficients of 0.9. Specifically, a clinical CT system
operating at 120 kV and 200 mA was able to detect plasma iodine concentrations
ranging from 100 g/mL to 500 g/mL, while a clinical 1.5 Tesla MR system was
able to measure plasma gadolinium concentrations ranging from 20 g/mL to 150
g/mL. Conclusion:This pilot study showed the potential
of employing CT and MR imaging to non-invasively map the distribution of
liposome carriers in vivo. The successful refinement of this imaging-based
pharmacokinetics assessment tool may facilitate delivery carrier design and
optimization in both the preclinical and clinical settings. Acknowledgement:A portion of the following work will be presented (oral
presentation) at the Nanotech 2006 Symposium that will take place on May 7-11,
2006 in
Pharmaceutical Care Services
in Hospitals of
Awad AI, Al-Ebrahim S, Abahussain E, Department of Pharmacy Practice,
Faculty of Pharmacy, Kuwait University
Purpose: To describe the current pharmacy practice in the general
hospitals based on self-reported practice by pharmacists, explore the awareness
of the pharmacists of the pharmaceutical care concept, identify their willingness to its implementation and the barriers that may
limit the implementation. Methods: Eighty hospital pharmacists working in four
general governmental hospitals were approached to be included in the study.
Data were colleted via face-to-face structured interview of the respondents
using a pre-tested questionnaire. Results: The response rate was 76.3%. Thirty
five (57.4%) of the respondents had frequently performed interventions on the
prescription through interacting with the medical doctors. Eighteen (29.5%)
were frequently contacted by doctors requesting information about drugs. Thirty
two (52.5%) had frequently provided patient counselling. Forty six (75.4%) of
the respondents claimed that they were aware of the pharmaceutical care
concept. Thirty five (76.1%) and 39.1% of those claiming to be aware of the
pharmaceutical care concept indicated that its main focus is the patient and
the appropriate objectives of the concept, respectively. Thirty (65.2%) of them
claimed that they had already implemented the pharmaceutical care services in
their practice. All respondents demonstrated willingness to implement the
pharmaceutical care practice in their hospitals. The main barriers perceived by
the participants were lack of time (78%), lack of staff (71.2%), and lack of
educational programs and training (44.1%). Conclusion: The current practice of
hospital pharmacists needs further improvement in relation to interaction with
doctors and patient counselling. The lack of uniformity in the responses
regarding the focus and objectives of pharmaceutical care indicates a lack of
appropriate understanding in this matter.All respondents have shown high
willingness towards the implementation of the pharmaceutical care services in
their practice.
Normal Pharmacodynamic
Response and Pharmacokinetics of Verapamil in Rheumatoid Arthritis Patients
Treated with Infliximab
Spencer Ling, Richard Z. Lewanczuk, Anthony
S. Russell, Brendan Ihejirika, and Fakhreddin Jamali. Faculties of Pharmacy and Medicine,
Purpose. Potency of the
cardiovascular drug verapamil is reduced in patients with rheumatoid arthritis
(RA) despite elevated plasma drug concentrations. Excess pro-inflammatory
mediators are associated with suppression of drug clearance and down-regulation
of various cardiovascular receptors. Infliximab is an anti-TNF monoclonal
antibody that reduces the levels of a multitude of pro-inflammatory cytokines
and has been shown to reverse the effects of inflammation on pharmacodynamic
response to sotalol in the inflamed rat. We examined whether RA patients who
are under treatment with infliximab still demonstrate reduced response to
verapamil and decreased verapamil clearance. Methods. Twelve RA patients on infliximab therapy were matched with twelve healthy
volunteers. Serum levels of interleukin-6 (IL-6), nitrite (NO2-),TNF and C-reactive protein (CRP) were measured to assess
degree of inflammation. Verapamil (80 mg) was administered orally, and then
ECG, blood pressure and verapamil enantiomers concentrations were measured for
12 h. Results. Serum TNF and CRP concentrations were significantly greater in
infliximab treated patients compared with healthy controls (TNF, p<0.001;
CRP, p=0.04). Serum nitrite and IL-6 concentrations were not significantly
different from controls. No significant differences in pharmacokinetics were
observed between control and infliximab treated subjects. Pharmacodynamic
response to verapamil was also not significantly different between the two
groups in any of the measured parameters. Conclusion. Patients with RA who are treated with infliximab demonstrated similar plasma
drug concentration and PR-interval response to verapamil when compared with
healthy volunteers despite elevated TNF and CRP concentrations.
Pro-inflammatory mediators nitrite and IL-6, appear to
be reduced by infliximab treatment.
Validation of a liquid
chromatography tandem mass spectrometry assay method for the determination of
sertraline in human plasma
Adrien Musuku, Gina deBoer, Grace van der Gugten,CANTEST BioPharma Services,
Purpose. Validation of an LC/MS/MS
assay method using atmospheric pressure electrospray ionization in the positive
ion mode for the measurement of sertraline in human plasma. Methods. Sertraline and the internal standard (d3-sertraline) were extracted from 1.0 mL
human plasma by liquid/liquid extraction using 1-chlorobutane as extraction
solvent. The analyte was chromatographically separated on a Phenomenex LunaB
C18 (2) (3 x 50 mm, 5 m) column using a gradient elution with 0.05% aqueous TFA
and 0.05% TFA in acetonitrile as the initial mobile phase, followed by LC/MS/MS
analysis. Detection and quantitation were carried out by ESI-MS/MS monitoring
the mass transitions from m/z 306 to 159 (sertraline) and m/z 309 to 159
(d3-sertraline). Results. The method was validated
over a concentration range from 0.25 to 100.00 ng/mL using a
linear calibration curve with a weighting of 1/x2. Inter-batch precision
(%CV) for standards ranged from 1.6 to 2.5. Inter-batch accuracy (%RE) ranged
from -1.2 to 1.3, indicating an acceptable goodness-of-fit. Inter-batch assay
precision (%CV) for quality control samples, ranged from 0.7 to 3.1 over five
concentration levels. Inter-batch assay accuracy (%RE) results for quality
control samples ranged from -2.9 to 14.3. The mean correlation coefficient was
0.9997 ± 0.0001. Assay recovery for sertraline was 77.4 % (%CV of 6.9), 65.4 %
(%CV of 8.9) and 72.8 % (%CV of 8.4) at 0.75 ng/mL, 30.00 ng/mL and 80.00 ng/mL
respectively.Conclusions. An accurate, sensitive and rapid LC/MS/MS analytical
assay was validated and successfully applied to the measurement of sertraline
in human plasma samples.
Validation of a liquid
chromatography mass spectrometry assay method for the determination of
trimethoprim and sulfamethoxazole in human plasma
Adrien Musuku, Luis Sojo, Sarah Bonorand, Grace van der Gugten,CANTEST BioPharma Services,
Purpose. To
develop and validate an LC/MS method for the simultaneous determination of
trimethoprim and sulfamethoxazole in human plasma. Methods. Trimethoprim, sulfamethoxazole and sulfisoxazole (internal standard), were
extracted from 0.1 mL human plasma by liquid/liquid extraction using ethyl
acetate as extraction solvent. The analytes were chromatographically separated
on a Zorbax Extend C18 (4.650 mm, 3.5 m) column using gradient elution with 95%
0.1% formic acid in water and 5% acetonitrile as initial mobile phase.
Quantitation was carried out by monitoring selected ions at m/z 291
(trimethoprim), m/z 254 (sulfamethoxazole) and m/z 268 (sulfisoxazole). Results. The method was validated over a concentration range
from 0.10 to 10.00 μg/mL (trimethoprim) and 1.00 to 100.00 μg/mL
(sulfamethoxazole) using a quadratic calibration curve weighted 1/x2 for
trimethoprim and 1/x for sulfamethoxazole. Inter-batch precision (%CV) for
standards ranged from 1.6 to 4.1 (trimethoprim) and 1.2 to 3.0
(sulfamethoxazole). Inter-batch accuracy (%RE) ranged from -6.7 to 7.1
(trimethoprim) and -3.7 to 2.1 (sulfamethoxazole). Inter-batch assay precision
(%CV) for quality control samples ranged from 1.5 to 5.3 (trimethoprim) and 1.1
to 3.8 (sulfamethoxazole). Inter-batch assay accuracy (%RE) for quality control
samples ranged from -5.5 to 3.2 (trimethoprim) and -2.2 to 2.2
(sulfamethoxazole). The mean correlation coefficients were 0.9983±0.0010 (trimethoprim)
and 0.9998±0.00001 (sulfamethoxazole). The mean assay recovery was 79.2±2.6%
(trimethoprim) and 99.4±3.6% (sulfamethoxazole). Freeze/thaw stability was
established at -40C and -70C for three cycles at each temperature. Conclusions. An accurate, sensitive and rapid analytical
assay was applied to measure trimethoprim and sulfamethoxazole in clinical
plasma samples.
Validation of a liquid
chromatography tandem mass spectrometry assay method for the determination of
verapamil and norverapamil in human plasma
Adrien Musuku, Gina de Boer, Meng Yu, CANTEST BioPharma Services,
Purpose. Validation
of an LC/MS/MS method for the simultaneous determination of verapamil, and its
metabolite, norverapamil, in human plasma. Methods. Verapamil, norverapamil and the internal standard, d6-verapamil, were extracted
from 0.1 mL human plasma by liquid/liquid extraction. The analytes were
chromatographically separated on a reverse-phase Zorbax Extend-C18 (4.650 mm,
3.5 m) column using gradient elution with initial mobile phase of 70% 10mM
ammonium acetate in water and 30% acetonitrile. ESI mass spectra were acquired
in positive ion mode with multiple reaction monitoring. Quantitation was
carried out by monitoring mass transitions at m/z 455 to m/z 165 (verapamil),
m/z 441 to m/z 165 (norverapamil) and m/z 461 to m/z 165 (d6-verapamil). Results. The method was validated over a concentration range
from 1.00 to 200.00 ng/mL for both analytes.
Inter-batch precision (%CV) for standards ranged from 3.1 to 7.3 for verapamil
and 4.4 to 8.3 for norverapamil. Inter-batch accuracy (%RE) ranged from -3.8 to
5.7 for verapamil and -3.9 to 4.3 for norverapamil. Inter-batch assay precision
(%CV) for quality control samples ranged from 1.1 to 3.9 for verapamil and 3.2
to 6.1 for norverapamil. Inter-batch assay accuracy (%RE) results for quality
control samples ranged from -5.6 to 1.2 for verapamil and -3.6 to 3.5 for
norverapamil. The mean correlation coefficients were 0.9979 ± 0.0014
(verapamil) and 0.9977 ± 0.0014 (norverapamil). Freeze/thaw stability for three
cycles was established at -40C.Conclusions. An accurate, sensitive and rapid
analytical assay was developed and applied to the measurement of verapamil and
norverapamil in clinical samples.
Published by the Canadian Society for Pharmaceutical Sciences.
Copyright © 1998 by the Canadian Society for Pharmaceutical Sciences.
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