Preparation and characterization of laminated thiolated chitosan-based freeze-dried wafers for potential buccal delivery of macromolecules.

This study involves the development and functional characterization of a thiolated chitosan (CS) system for potential buccal delivery of proteins. Thiolated CS was synthesized by conjugating pure CS with thioglycolic acid and dialyzed to remove excess acid. Amount of thiol groups immobilized on CS was determined using L-cysteine calibration curve. The weight average molecular weights of CS and thiolated CS were monitored using gel permeation chromatography. Laminated wafers were obtained by pouring gels (containing bovine serum albumin; BSA, different amounts of glutathione as enzyme inhibitor and mucin to mimic salivary conditions) of the thiolated CS into moulds previously lined with impervious ethylcellulose (EC) films and freeze-dried. The resulting formulations were analyzed using attenuated total reflectance Fourier transform infrared (FTIR) spectroscopy, circular dichroism (CD) and scanning electron microscopy (SEM). The formulations were further characterized for functional buccal mucosa performance using hydration, swelling, mucoadhesion and in vitro drug dissolution studies. FTIR showed successful thiolation of CS's amine functionality, CD confirmed that BSA conformation remained unchanged throughout the gel formulation and freeze-drying process, whilst SEM showed a porous microstructure of the wafers and a uniform EC film laminate with no visible pores or cracks. The functional characterization studies showed that glutathione had significant effects on hydration, mucoadhesion and subsequently drug dissolution and release characteristics, whilst mucin affected the mucoadhesive properties of the wafers. It was concluded that BSA-loaded wafers containing 10% w/w glutathione as enzyme inhibitor was the formulation choice for potential buccal delivery and should be selected for further investigations.

Development and physico-mechanical characterization of carrageenan and poloxamer-based lyophilized matrix as a potential buccal drug delivery system.

CONTEXT AND OBJECTIVES: The buccal mucosa presents a unique surface for non-invasive drug delivery and also avoids first-pass metabolism. The objective of this study was the formulation development of polymeric mucoadhesive lyophilized wafers as a matrix for potential buccal drug delivery. MATERIALS AND METHODS: Differential scanning calorimetry (DSC) was used to develop an optimum freeze-cycle, incorporating an annealing step. The wafers were prepared by lyophilization of gels containing three polymers, κ-carrageenan (CAR 911), poloxamer (P407) and polyethylene glycol 600 (PEG 600). The formulations were characterized using texture analysis (for mechanical and mucoadhesion properties), hydration studies, thermogravimetric analysis (TGA), DSC, X-ray powder diffraction (XRPD) and scanning electron microscopy (SEM). RESULTS AND DISCUSSION: DSC showed the eutectic temperature (12.8 °C) of the system where the liquid solution and pure solids both existed at a fixed pressure which helped determine the freeze-annealing cycle at 55 °C for 7 h. Mechanical resistance to compression, hydration and mucoadhesion studies showed that optimized wafers were obtained from aqueous gels containing 2% w/w CAR 911, 4% w/w P407 and 4.4% w/w PEG 600. TGA showed residual water of approximately 1% and SEM showed a porous polymeric network that made ease of hydration possible. CONCLUSIONS: Lyophilized wafers by freeze-drying gels containing 2% w/w CAR 911, 4% w/w P407 and 4.4% w/w PEG 600 with optimum physico-mechanical properties has been achieved.

Forced Degradation Studies, Elucidation of Degradation Pathways and Degradation Kinetics of Xylopic Acid via LC and LC-MS/MS Analyses.

Stability studies of active pharmaceutical ingredients (APIs) remain an essential quality requirement of the pharmaceutical industry. Stability data of an API could guide in the choice of its processing technique, packaging method and storage conditions. Here, we sought to determine the stability or otherwise of xylopic acid (XA) under various stress conditions as stipulated by the International Conference on Harmonization (ICH). XA is a diterpene kaurene isolate of the African spice, Xylopia aethiopica (Annonaceae) that is credited with diverse biological activities. XA was subjected to various stress conditions (hydrolytic, oxidative, photolytic and thermal) and its degradation products characterized. Seven degradation products were identified and tentatively characterized by LC-MS/MS analysis. The probable degradation pathways for the seven degradation products were then predicted. Using a simple and validated UHPLC-DAD method, the degradation kinetics of XA under the different stress conditions were comprehensively assessed. The degradation of XA under all the stress conditions followed the first order reaction kinetics. XA was found to be less stable in strongly acidic or strongly basic solutions as well as in an oxidizing agent (hydrogen peroxide). The stability of XA was also found to be pH- and temperature-dependent. Its stability was however not affected by UV-light irradiation.

Pharmacokinetics and in vitro liver microsomal enzyme metabolism of Xylopic acid.

Xylopic acid (XA) is a bioactive diterpene kaurene isolate of the Guinea pepper fruit, Xylopia aethiopica (Annonaceae) with numerous well-established biological effects. In this study, we aimed to fill certain scientific voids in terms of the scientific literature on XA, specifically, its pharmacokinetic (PK) parameters and in vitro liver microsomal enzyme metabolism. A new LC-MS/MS method was developed and validated for the determination of the plasma concentration-time profile of XA. The method was found to be accurate, precise, selective and repeatable with lowest limit of quantification (LLOQ) of 10 ng/mL and run time of 15 min. The maximum plasma concentration (Cmax), time at which maximum plasma concentration was attained (Tmax), half-life (t1/2), clearance (CL) and mean residence time (MRT) of XA were 167.03 ± 6.18 ng/mL; 10 h; 13.03 ± 7.33 h; 0.04 ± 0.01 mL/h/kg and 23.83 ± 11.02 h respectively. Six metabolites (M1-M6) were tentatively identified after XA was subjected to in vitro liver microsomal enzyme metabolism. The metabolites were the products of methylation (M1), glucuronidation (M2), deacetylation (M3), glucosylation (M4), hydroxylation and glutamic acid addition (M5) and glutathionylation (M6). The outcome of this study provides useful insights that could guide further research on XA.

Trace Metals in Cannabis Seized by Law Enforcement in Ghana and Multivariate Analysis to Distinguish among Different Cannabis Farms.

For hundreds of years, cannabis has been one of the most known cultivated plants due to its variety of uses, which include as a psychoactive drug, as well as for medicinal activity. Although prohibiting cannabis products, the countries of the African continent are the largest producers of cannabis in the world; a fact that makes the trafficking of cannabis-based illicit drugs a high priority for local law enforcement authorities. The latter are exceedingly interested in the use of chemical analyses for facilitating quantification, identification, and tracing of the origin of seized cannabis samples. Targeting these goals, and focusing on the country of Ghana, the present study used inductively coupled plasma mass spectrometry (ICP-MS) for the determination of 12 elements (Pb, Cu, Ca, Mg, Mn, Zn, Cd, As, Hg, Fe, Na, and K) in cannabis seized by Ghana's law enforcement authorities and soils of cannabis farms. Furthermore, multivariate analysis was applied to distinguish among different cannabis farms and match them with the samples. As a result, 22 seized cannabis samples and 12 other cannabis samples with their respective soils were analyzed to reveal considerable As and Pb concentrations. As and Pb levels in cannabis were found up to 242 ppb for As and 854 ppb for Pb. Multivariate analysis was applied for separating different cannabis farms and seized samples based on elemental analysis, evidently linking the seized samples with two Ghana regions.

Ultraviolet-Visible Spectroscopy and Chemometric Strategy Enable the Classification and Detection of Expired Antimalarial Herbal Medicinal Product in Ghana.

To meet the growing demand for complementary and alternative treatment for malaria, manufacturers produce several antimalarial herbal medicinal products. Herbal medicinal products regulation is difficult due to their complex chemical nature, requiring cumbersome, expensive, and time-consuming methods of analysis. The aim of this study was to develop a simple spectroscopic method together with a chemometric model for the classification and the identification of expired liquid antimalarial herbal medicinal products. Principal component analysis model was successfully used to distinguish between different herbal medicinal products and identify expired products. Principal component analysis showed a clear class separation between all five herbal medicinal products (HMP) studied, with explained variance for first and second principal components as 37.51% and 26.38%, respectively, while the third principal component had 18.74%. Support vector machine classification gave specificity and accuracy of 1.00 (100%) for training set data for all the products. The validation set HMP1, HMP2, and HMP3 had sensitivity, specificity, and accuracy of 1.00. HMP4 and HMP5 had sensitivity and specificity of 0.90 and 1.00, respectively, and an accuracy of 0.98. The support vector machine classification and principal component analysis models were successfully used to identify expired herbal medicinal products. This strategy can be used for rapid field detection of expired liquid antimalarial herbal medicinal products.

Oral thin films as a remedy for noncompliance in pediatric and geriatric patients.

Pediatric and geriatric patients experience swallowing difficulties for traditional oral dosage forms, such as tablets. Further, microbial contamination, chemical stability, unpleasant taste and swallowing large volumes of fluids have led to low therapeutic efficacy and patient noncompliance. The emergence of oral thin films has resulted in dramatic improvements in compliance and drug therapy outcomes in pediatric and geriatric patients. Oral thin films do not require water for administration, are readily hydrated upon contact with saliva, adhere to the mucosa and disintegrate ideally under 1 min. This article provides an overview of oral thin films, modern trends in their formulation and characterization, available commercial products, information to fill knowledge gaps and future potential and economic prospects of oral thin film technology, with emphasis on their use in the pediatric and geriatric patient groups.

The Challenges and Knowledge Gaps in Malaria Therapy: A Stakeholder Approach to Improving Oral Quinine Use in the Treatment of Childhood Malaria in Ghana.

BACKGROUND: The study was undertaken to elicit the knowledge, views, and perceptions of key stakeholders on malaria, its bioburden, and treatment options, in order to ascertain the knowledge gabs and challenges, especially in the use of oral quinine in childhood malaria. METHODS: A cross-sectional survey was conducted using a well-structured Likert Scale and self-administered questionnaire. The principal site of the study was a government-run children's hospital located in the Ashiedu Keteke Sub-Metro of Accra. The study population included health workers, parents, and guardians or care givers. The participants were 300, purposively selected, and consisted of both men (41%) and women (59%) who were twenty years and above, whether employed (42%), self-employed (37%), or unemployed (21%). RESULTS: Majority of the participants (78%) demonstrated above average knowledge of malaria. However, their awareness of the causes, modes of transmission, signs, and symptoms as well as preventive mechanisms of malaria did not result in low incidence of malaria. About 77% of the respondents agreed they would seek treatment within 24 hours once signs and symptoms are detected. Though close to 50% undertook home treatment of malaria, majority eventually sought treatment at hospital or clinic. Above 92% of respondents knew that quinine is used to treat malaria and agreed its bitter taste greatly affects compliance, especially in children. Consequently, 95% of the respondents would be glad if its bitter taste is masked. CONCLUSION: The study demonstrated the availability of substantial knowledge of the devastating effects of malaria, especially in children. Therefore, there is the need to ensure the availability and utilization of effective paediatric formulations in the fight against malaria. From this study, fast dissolving oral thin film with a good mouth feel, would be the formulation of choice for quinine.

Functional characterisation and permeation studies of lyophilised thiolated chitosan xerogels for buccal delivery of insulin.

Stable and mucoadhesive, lyophilised, thiolated chitosan xerogels, loaded with insulin for buccal mucosa deliv- ery, in place of the currently used parenteral route have been developed. The xerogels were backed with impervious ethyl- cellulose laminate to ensure unidirectional release and also loaded with enzyme inhibitor to enhance insulin permeability across the buccal mucosa. Characterisation of xerogels using(1) HNMR confirmed the degree of deacetylation of the syn- thesised thiolated chitosan. The amount of thiol groups immobilised on the modified chitosan was quantified by Ellman's reaction and molecular weight monitored by gel permeation chromatography. The stability of the secondary structure of insulin was examined by attenuated total reflectance Fourier transform infra-red spectroscopy and circular dichroism. In vitro and ex vivo permeation studies were undertaken by using EpiOral ™ and sheep buccal membrane respectively. Insu- lin released from thiolated chitosan xerogels, loaded with aprotinin (enzyme inhibitor and permeation enhancer) showed a 1.7-fold increase in permeation through EpiOral ™ buccal tissue construct compared to the pure drug. However, permea- tion was decreased for xerogels containing the enzyme inhibitor glutathione. Further, aprotinin containing xerogels en- hanced insulin permeation through sheep buccal membrane and demonstrated good linear correlation with the permeation data from the EpiOral ™ study. The results show the potential application of lyoph ilised thiolated chitosan xerogels con- taining aprotinin with improved mucoadhesion, penetration enhancing and enzyme inhibition characteristics for buccal mucosa delivery of macromolecules such as insulin.

Multifunctional medicated lyophilised wafer dressing for effective chronic wound healing.

Wafers combining weight ratios of Polyox with carrageenan (75/25) or sodium alginate (50/50) containing streptomycin and diclofenac were prepared to improve chronic wound healing. Gels were freeze-dried using a lyophilisation cycle incorporating an annealing step. Wafers were characterised for morphology, mechanical and in vitro functional (swelling, adhesion, drug release in the presence of simulated wound fluid) characteristics. Both blank (BLK) and drug-loaded (DL) wafers were soft, flexible, elegant in appearance and non-brittle in nature. Annealing helped to improve porous nature of wafers but was affected by the addition of drugs. Mechanical characterisation demonstrated that the wafers were strong enough to withstand normal stresses but also flexible to prevent damage to newly formed skin tissue. Differences in swelling, adhesion and drug release characteristics could be attributed to differences in pore size and sodium sulphate formed because of the salt forms of the two drugs. BLK wafers showed relatively higher swelling and adhesion than DL wafers with the latter showing controlled release of streptomycin and diclofenac. The optimised dressing has the potential to reduce bacterial infection and can also help to reduce swelling and pain associated with injury due to the anti-inflammatory action of diclofenac and help to achieve more rapid wound healing.

An integrated buccal delivery system combining chitosan films impregnated with peptide loaded PEG-b-PLA nanoparticles.

Peptide (insulin) loaded nanoparticles (NPs) have been embedded into buccal chitosan films (Ch-films-NPs). These films were produced by solvent casting and involved incorporating in chitosan gel (1.25% w/v), NPs-Insulin suspensions at three different concentrations (1, 3, and 5mg of NPs per film) using glycerol as plasticiser. Film swelling and mucoadhesion were investigated using 0.01M PBS at 37°C and texture analyzer, respectively. Formulations containing 3mg of NPs per film produced optimised films with excellent mucoadhesion and swelling properties. Dynamic laser scattering measurements showed that the erosion of the chitosan backbone controlled the release of NPs from the films, preceding in vitro drug (insulin) release from Ch-films-NPs after 6h. Modulated release was observed with 70% of encapsulated insulin released after 360h. The use of chitosan films yielded a 1.8-fold enhancement of ex vivo insulin permeation via EpiOral™ buccal tissue construct relative to the pure drug. Flux and apparent permeation coefficient of 0.1µg/cm(2)/h and 4×10(-2)cm(2)/h were respectively obtained for insulin released from Ch-films-NPs-3. Circular dichroism and FTIR spectroscopy demonstrated that the conformational structure of the model peptide drug (insulin) released from Ch-films-NPs was preserved during the formulation process.

Development and characterisation of chitosan films impregnated with insulin loaded PEG-b-PLA nanoparticles (NPs): a potential approach for buccal delivery of macromolecules.

Mucoadhesive chitosan based films, incorporated with insulin loaded nanoparticles (NPs) made of poly(ethylene glycol)methyl ether-block-polylactide (PEG-b-PLA) have been developed and characterised. Blank-NPs were prepared by double emulsion solvent evaporation technique with varying concentrations of the copolymer (5 and 10%, w/v). The optimised formulation was loaded with insulin (model protein) at initial loadings of 2, 5 and 10% with respect to copolymer weight. The developed NPs were analysed for size, size distribution, surface charge, morphology, encapsulation efficiency and drug release. NPs showing negative (ζ)-potential (<-6 mV) with average diameter> 300 nm and a polydispersity index (P.I.) of ≈ 0.2, irrespective of formulation process, were achieved. Insulin encapsulation efficiencies of 70% and 30% for NPs-Insulin-2 and NPs-Insulin-5 were obtained, respectively. The in vitro release behaviour of both formulations showed a classic biphasic sustained release of protein over 5 weeks which was influenced by pH of the release medium. Optimised chitosan films embedded with 3mg of insulin loaded NPs were produced by solvent casting with homogeneous distribution of NPs in the mucoadhesive matrix, which displayed excellent physico-mechanical properties. The drug delivery system has been designed as a novel platform for potential buccal delivery of macromolecules.

Effect of membrane dialysis on characteristics of lyophilised chitosan wafers for potential buccal delivery of proteins.

The effect of membrane dialysis on the characteristics of chitosan based lyophilised wafers was investigated. Gels loaded with BSA, glycerol and d-mannitol were lyophilised with or without membrane dialysis and characterised by X-ray diffraction, attenuated total reflectance Fourier transform infra red spectroscopy, circular dichroism, scanning electron microscopy, hydration capacity, in vitro mucoadhesivity and drug dissolution. The dialysed wafers demonstrated enhanced mucoadhesion and drug release properties while newly formed sodium acetate in the undialysed wafers caused increased crystallinity with poor mucoadhesion and drug release properties. Removal of sodium acetate by membrane dialysis is essential for obtaining optimised wafers for potential application to the buccal mucosa surface.

In vitro characterisation of chitosan based xerogels for potential buccal delivery of proteins.

Chitosan and thiolated-chitosan based xerogels have been prepared by lyophilising aqueous gels of the polymers incorporating glycerol, d-mannitol and BSA with an annealing process. Analytical characterisation was by circular dichroism, infrared spectroscopy, X-ray diffraction and scanning electron microscopy. Swelling capacities of 1,110 ± 23.3% and 480 ± 18.2% were obtained for the chitosan and TG-chitosan xerogels respectively. Thiolation caused improved in vitro mucoadhesive properties by demonstrating peak adhesive force of 4.5 ± 0.7 and 5.8 ± 0.2N, and total work of adhesion of 6.5 ± 1.0 and 19 ± 0.8 mJ for chitosan and thiolated-chitosan xerogels respectively. In vitro drug dissolution studies using Bradford's assay showed BSA release of 91.5 ± 3.7% and 94.4 ± 7.3% from the chitosan and thiolated-chitosan xerogels respectively. These results demonstrate the potential of developing lyophilised thiolated-chitosan xerogels with optimised mucoadhesion characteristics for buccal mucosa delivery of protein based drugs.

Development and physico-mechanical characterisation of lyophilised chitosan wafers as potential protein drug delivery systems via the buccal mucosa.

Lyophilised wafers from chitosan have been developed as potential protein drug delivery systems via the buccal mucosa. Wafers were prepared by lyophilising aqueous gels of the polymer incorporating varying concentrations of glycerol as plasticizer and d-mannitol as cryoprotectant. The different formulations were characterised by their physico-mechanical properties in order to select the optimum system for further development. The optimised formulation with 6.5 mg each of both plasticizer and cryoprotectant was loaded with bovine serum albumin and lyophilised with or without annealing. Differential scanning calorimetry was used to determine the appropriate lyophilisation cycle by evaluating thermal events before lyophilisation and possible phase separation of bovine serum albumin after lyophilisation. Texture analysis was employed to investigate the in vitro mucoadhesive properties in tensile mode, residual moisture content by thermo-gravimetric analysis while hydration capacity and drug release studies were performed in 0.1 M phosphate buffered saline. Microscopic architecture and crystallinity were examined using scanning electron microscopy and X-ray diffractometry respectively. The ease of hydration, in vitro mucoadhesive characteristics, microscopic architecture and BSA release were influenced by the annealing process. A 7 h cumulative percentage drug release of 91.5% and 80.1% was observed for the annealed and non-annealed wafers respectively. The results showed the potential of employing lyophilised chitosan wafers for buccal mucosa delivery of protein based drugs.