Development of Cetylpyridinium-Alginate Nanoparticles: A Binding and Formulation Study.

In this study the development of stable polyelectrolyte-surfactant complex nanoparticles composed of alginate and cetylpyridinium chloride (CPC), with and without ZnCl2, for therapeutic use, is investigated. The mechanism of CPC binding by alginate was analyzed using a cetylpyridinium cation (CP(+)) selective membrane electrode. The cooperative nature of the interaction between CP(+) and alginate was underlined by the sigmoidal shape of the binding isotherms. The presence of salts was shown to weaken interactions and, moreover, ZnCl2 reduced the cooperativity of binding. The CP(+) cations in the form of micellar associates acted as multivalent crosslinkers of the alginate chains where stable dispersions of CP-alginate nanoparticles were formed in water at CP(+)/alginate charge ratios from 0.2 to 0.8. Characterization of the nanoparticles showed hydrodynamic diameters from 140 to 200nm, a polydispersity index below 0.2, a negative zeta potential and spherical morphology. The entrapment efficiency of CPC was ∼94%, the loading capacity more than 50% and prolonged release over 7days were shown. The formulations with noted charge ratios resulted in stable CP-alginate nanoparticles with a potential of treating periodontal disease.

Comparative bioavailability and antimicrobial activity of cetylpyridinium chloride mouthrinses in vitro and in vivo.

PURPOSE: Commercialized cetylpyridinium chloride (CPC) mouthrinses were compared for antimicrobial substantivity/bioavailability in an in vitro disk retention assay (DRA) and clinical antimicrobial activity in vivo in the plaque glycolysis and regrowth method (PGRM). METHODS: Formulations compared in this testing included commercially available CPC mouthrinses: Crest Pro Health (CPH), (containing 700 ppm formulated CPC); Colgate Total Puerto Rico (CT450), (containing 450 ppm formulated CPC); Colgate Total US (CT750), (containing 750 ppm formulated CPC); and Scope Mouthwash (SCP), (containing 450 ppm formulated CPC). A water control (CTR) was included in one of the PGRM clinical trials. Two separate clinical PGRM studies employed a controlled, double-blind, randomized, crossover design where qualified adult PGRM panelists were supplied with acclimation NaF dentifrice for use throughout the trials. On treatment days, subjects sampled baseline plaque and then rinsed with assigned mouthrinse following morning toothbrushing. Treated plaque samples were collected 15 and 45 minutes after rinsing. Sampled plaques were vortexed, normalized for biomass and incubated under standard conditions to assess glycolysis. pH response of treated plaques in incubation buffers were compared to baseline untreated plaque values and an Area Under Curve (AUC) composite/aggregate analysis of glycolysis inhibition was used for treatment comparisons. A laboratory disk retention substantivity/bioavailability assay measured adsorption affinity of CPC in mouthrinse for anionic cellulose disks in vitro. RESULTS: Clinical PGRM studies showed significant differences in antibacterial clinical efficacy of commercialized mouthrinses. Combining clinical study results reveals rank ordered efficacy CPH > CT750 > SCP > CT450 > CTR. Comparison of DRA to PGRM glycolysis showed a linear relation highlighting correlation of CPC bioavailability to clinical antimicrobial performance of CPC mouthrinses.

In vitro CPC retention and VSC adsorption by IPM oil droplets: possible mechanisms of action of a two phase mouthwash.

Two phase oil-water mouthwash has been previously shown to efficiently bind oral microorganisms, relying on their cell surface hydrophobicity. The aim of the present in vitro study was to test the cetylpyridinium chloride (CPC) retention and volatile sulfide compounds (VSCs) adsorption abilities of the oil droplets created by mixing of a two phase oil-water solution. VSC adsorption was assayed using a salivary incubation assay and garlic powder solutions, and demonstrated using microscopic sulfide assay. CPC retention was assayed by kinetic and endpoint measurement of Streptococcus salivarius outgrowth using microplate (ELISA) reader. Results showed that the isopropyl myristate (IPM) oil droplets in the two phase solutions were able to adsorb 68-80% of VSCs. CPC at a concentration of 0.05% was most affectively retained by the oil droplets showing a significantly increase in residual antibacterial activity against Streptococcus salivarius. These results taken together, suggests that VSC adsorption and CPC retention by IPM oil droplets may be two additional mechanisms in the activity of the two phase mouthwash formulation.

Treatment with a novel topical nanoemulsion (NB-001) speeds time to healing of recurrent cold sores.

BACKGROUND: Current topical therapies for cold sores are only marginally beneficial due to poor skin penetration. We assessed the safety and efficacy of a novel topical antiviral nanoemulsion (NB-001) with high tissue bioavailability. OBJECTIVES: The primary endpoint was the time to lesion healing. METHODS: 482 subjects with recurrent cold sores were randomized to self-initiate treatment with either vehicle or NB-001 (0.1%, 0.3% or 0.5%) at the first signs or symptoms of a cold sore episode. Lotion was applied 5 times per day, approximately 3 to 4 hours apart, for 4 days. Time to lesion healing was correlated with NB-001 bioavailability determined in human cadaver skin. RESULTS: Subjects treated with 0.3% NB-001 showed a 1.3-day improvement in the mean time to healing compared to vehicle (P=0.006). This was consistent with human cadaver skin data indicating that the 0.3% nanoemulsion had the highest bioavailability, compared to 0.1% and 0.5% emulsions. No significant safety or dermal irritation concerns or systemic absorption were noted with any of the doses. CONCLUSIONS: Topical NB-001 (0.3%) was well tolerated and highly efficacious in shortening the time to healing of cold sores. The improvement in time to healing was similar to that reported for oral nucleoside analogues, but without systemic exposure. Topical agents for recurrent herpes labialis (cold sores) reduce healing time by one half day, compared to oral therapies that speed healing by a day or more. A topical antiviral nanoemulsion was well tolerated and improved cold sore healing time by over a day compared to vehicle control. Nanoemulsion (NB-001) could represent a more efficacious topical treatment for recurrent cold sores.

Efficacy of a high bioavailable cetylpyridinium chloride mouthrinse over a 24-hour period: a plaque imaging study.

PURPOSE: To evaluate the antiplaque benefits of a 0.07% high bioavailable, alcohol-free cetylpyridium chloride (CPC) rinse used after toothbrushing versus toothbrushing alone. METHODS: A digital plaque image analysis technique was used to quantify in situ plaque formation in a subject population carrying out modified hygiene using standard fluoridated dentifrice or standard dentifrice augmented with 30 seconds mouthrinsing with an alcohol-free mouthrinse containing 700 ppm CPC. RESULTS: Comparison of plaque formation 24 hours following "last hygiene" revealed that brushing followed by CPC mouthrinse use provided a statistically significant decrease in plaque coverage on teeth averaging 42% as compared with brushing only. Moreover, toothbrushing with a standard dentifrice in the morning resulted in 34% less plaque when subjects used the CPC mouthrinse 24 hours prior to examination. CLINICAL SIGNIFICANCE: These results support the strong retention and lasting antiplaque efficacy of high bioavailable CPC mouthrinse and suggest that the plaque biofilms formed during CPC use are susceptible to more efficient debridement.

Silica coatings on clarithromycin.

Pre-crystallized clarithromycin (6-O-methylerythromycin A) particles were coated with silica from the tetraethyl orthosilicate (TEOS)-ethanol-aqueous ammonia system. The coatings had a typical thickness of 100-150 nm and presented about 15 wt.% of the silica-drug composite material. The properties of the coatings depended on reactant concentration, temperature and mixing rate and, in particular, on the presence of a cationic surfactant (cetylpyridinium chloride). In the presence of cetylpyridinium chloride the silica coatings slightly decreased the rate of pure clarithromycin dissolution.

Soft antimicrobial agents: synthesis and activity of labile environmentally friendly long chain quaternary ammonium compounds.

A series of soft quaternary ammonium antimicrobial agents, which are analogues to currently used quaternary ammonium preservatives such as cetyl pyridinium chloride and benzalkonium chloride, were synthesized. These soft analogues consist of long alkyl chain connected to a polar headgroup via chemically labile spacer group. They are characterized by facile nonenzymatic and enzymatic degradation to form their original nontoxic building blocks. However, their chemical stability has to be adequate in order for them to have antimicrobial effects. Stability studies and antibacterial and antiviral activity measurements revealed relationship between activity, lipophilicity, and stability. Their minimum inhibitory concentration (MIC) was as low as 1 microg/mL, and their viral reduction was in some cases greater than 6.7 log. The structure-activity studies demonstrate that the bioactive compounds (i.e., MIC for Gram-positive bacteria of <10 microg/mL) have an alkyl chain length between 12 and 18 carbon atoms, with a polar headgroup preferably of a small quaternary ammonium group, and their acquired inactivation half-life must be greater than 3 h at 60 degrees C.

An in vitro evaluation of the availability of cetylpyridinium chloride and chlorhexidine in some commercially available mouthrinse products.

OBJECTIVES: To assess the comparative activity of mouthrinses containing cetylpyridinium chloride (CPC) and chlorhexidine using the propensity to cause extrinsic staining in vitro as the outcome variable. METHODS: Saliva-coated clear acrylic specimens were exposed to cyclical staining regimens of either CPC or chlorhexidine followed by tea. Water and 0.2% chlorhexidine were used as negative and positive controls respectively. Staining cycles were repeated until an optical density of > 2 was reached by one of the products. RESULTS: For CPC there was a highly significant difference in staining between the products. Two CPC products performed numerically little better than water. For the chlorhexidine products the 0.2% formulation produced the most staining although little more than the UK version of the 0.1% rinse. The French 0.1% rinse produced by the same manufacturer as the UK formulation showed markedly reduced staining potential although significantly greater than water. CONCLUSION: This study, supported by previous in vitro and in vivo studies, indicates discrepancies in the availability of CPC and chlorhexidine in some mouthrinse products. Importantly, this may have an effect on the potential of some rinses to provide the expected plaque inhibitory activity.

Design and characterization of mucoadhesive buccal patches containing cetylpyridinium chloride.

Mucoadhesive patches for delivery of cetylpyridinium chloride (CPC) were prepared using polyvinyl alcohol (PVA), hydroxyethyl cellulose (HEC) and chitosan. Swelling and bioadhesive characteristics were determined for both plain and medicated patches. The results showed a remarkable increase in radial swelling (S(D)) after addition of the water-soluble drug (CPC) to the plain formulae. A decrease in the residence time was observed for PVA and chitosan-containing formulae. Higher drug release was obtained from PVA patches compared to HEC ones, while both are non-ionic polymers. A considerable drop in release was observed for chitosan formulae after the addition of water-soluble additives, polyvinyl pyrrolidone (PVP) and gelatin. Ageing was done on PVA formulae; the results showed there was no influence on the chemical stability of CPC, as reflected from the drug content data. Physical characteristics of the studied patches showed an increase in the residence time with storage accompanied with a decrease in drug release. This may be due to changes in the crystal habit of the drug as well as to slight agglomeration of the polymer particles.

Utility of MTT assay in three-dimensional cultured human skin model as an alternative for draize skin irritation test: approach using diffusion law of irritant in skin and toxicokinetics-toxicodynamics correlation.

PURPOSE: A cytotoxicity assay using a three-dimensional cultured human skin model, Living Skin Equivalent-high (LSE-high) was evaluated as an alternative to the Draize skin irritation tests using animals. A relation between the cytotoxicity and calculated concentration of an irritant in skin was also evaluated. METHODS: Colorimetric thiazoyl blue (MTT) conversion assay and a surfactant, cetylpyridinium chloride (CPC), were selected as a cytotoxicity assay and a model irritant. The fraction of dead cell number in the MTT assay or the Draize irritation score (in vitro and in vivo irritation data, respectively) was treated as a function of CPC concentration in the viable skin of LSE-high and guinea pig. Separately, in vitro permeations of CPC through the LSE-high or excised guinea pig skin were determined to calculate the average concentration of CPC in the viable skin using the Fickian diffusion theory. The obtained relations between the irritation scores and CPC concentration were evaluated by the Emax model (Hill equation). RESULTS: CPC concentration showing 50% irritation (IC50) was similar for the MTT assay (18.9%) and Draize test (12.3%), and a good relationship (r = 0.981) was observed between the fraction of dead cell number and the Draize score. In contrast, IC50, 1.32%, for the MTT assay in LSE-high was much lower than that using guinea pig skin. We then corrected the results for the MTT assay using a ratio of IC50 in guinea pig skin against LSE-high, resulting in a good relation between both MTT results in guinea pig skin and LSE-high. CONCLUSION: The present results suggest that the MTT assay using LSE-high may be utilized as an alternative for the Draize test in animals for evaluating skin irritation.

Buccoadhesive erodible disk for treatment of oro-dental infections: design and characterisation.

Buccoadhesive erodible disks of cetylpyridinium chloride were prepared using different bioadhesive polymers along with excipients like mannitol. The purpose of designing the erodible disk was to obviate the need for removal of exhausted device. The optimized disk containing 5.0 mg of cetylpyridinium chloride, 2.0 mg of magnesium stearate and 6.0 mg of mannitol along with sodium carboxy methyl cellulose DVP and hydroxypropylmethylcellulose K4M in the ratio of 1:3 was found to release the drug for a period of over 6.0 h without getting dislodged. Maximum in vitro drug release was found to be 94.78% in 6.0-h study. In situ release characteristics were evaluated using a 'flow-through assembly', which simulated the conditions of the human buccal cavity. The drug concentrations in the in situ samples were found to be above minimum inhibitory concentration (MIC) of the drug. The bioadhesive performance and the surface pH of the disks were satisfactory. Cetylpyridinium chloride disks were tested against microorganisms commonly found in oro-dental infections namely Candida albicans, Staphylococcus aureus, Escherichia coli and Streptococcus mutans. The disk as well as the in situ samples showed inhibition of growth of microorganisms. Approval was taken from Jamia Hamdard Review Board (Ethical Board) to perform in vivo studies in healthy human volunteers. In vivo evaluation of buccoadhesive disks revealed adequate comfort, taste, and non-irritation and none of the volunteers reported severe dry mouth/severe salivation or heaviness at the place of attachment. Salivary concentrations were maintained above MIC for 8.0 h. Correlation was found between the drug concentration in situ and concentration of drug in saliva collected in healthy human volunteers. The correlation was found to be positive with a correlation coefficient of 0.9596. It was found to be statistically significant at 5% confidence level (P<0.05).

Buccoadhesive tablets for the slow delivery of cetylpyridinium chloride: design and in vitro/in vivo analysis.

A buccoadhesive slow-release system constituted by a monolayer tablet was developed for the administration of cetylpyridinium chloride (CPC). Tablet composition was based on a mixture of a bioadhesive polymer with conventional excipients. Three different polymers (Methocel K4M, Noveon AA1, Carbopol 974P) in three different concentrations (10%, 20%, 30%) were used. The release profile and the adhesion properties of nine formulations were evaluated in vitro. Based on these results three formulations containing 10% w/w of the tested polymers were chosen to assess, with preliminary in vivo studies, the compliance and the residence time in the month of the systems. Tablets containing Noveon AA1 showed the best performances.

Evaluation of Cetylpyridinium Chloride-Containing Mouthwashes Using In Vitro Disk Retention and Ex Vivo Plaque Glycolysis Methods.

The Disk Retention Assay (DRA) is an in vitro method developed to measure the available level of cetylpyridinium chloride (CPC) in mouthwash formulations. This method is based on the binding of the cationic CPC molecule to the anionic surface of a cellulose filter disk. Aqueous CPC solutions demonstrate a linear response (A545) for concentrations up to 0.3%. Higher levels of CPC showed no increased response in the assay. Among common oral product ingredients, at relevant concentrations, surfactants are the primary compounds which inhibit CPC detection and hence, chemical availability. Poloxamer-407 decreased CPC availability to 60% at 0.1%, to 10% at 0.5%, and to 24-33% for 0.2-0.4%. Polysorbate-80 decreased CPC availability to 30% at 0.1% and 6% at 0.25%. A range (4-54%) of available levels of CPC were determined for several commercial products containing 0.045-0.05% nominal levels of CPC indicating significant formulation excipient influence. A plaque glycolysis (PG) assay was used to determine the biological activity of all mouthwash products analyzed by DRA. An experimental series of mouthwash formulations having nominal CPC levels of 0-0.10% demonstrated a good correlation (r2 = 0.955) between the calculated available level of CPC (DRA) and inhibition of plaque glycolysis. The calculated available level of CPC from select commercial mouthwash products, also fit the established correlation with biological activity. The combination of DRA and plaque glycolysis methods are valuable tools which can be used during development to maximize the biological activity of CPC mouthwash formulations prior to clinical evaluation.

Primary interactions of three quaternary ammonium compounds with blastospores of Candida albicans (MEN strain).

The absorption of three quaternary ammonium compounds (QAC), cetylpyridinium chloride, cetrimide and benzalkonium chloride, onto the surface of blastospores of Candida albicans (MEN strain) was examined at room temperature. Equilibrium uptake occurred in less than 30 seconds for cetylpyridinium chloride and cetrimide whereas 5 min contact time was required for benzalkonium chloride. The adsorption of all three agents may be mathematically described as Langmuirian and hence a concentration-dependent formation of drug-monolayer on the surface of the blastospore occurred. From this the number of molecules adsorbed onto the surface of a single blastospore was calculated to be 1.33 x 10(12), 3.17 x 10(12) and 2.32 x 10(12) for cetylpyridinium chloride, cetrimide and benzalkonium chloride, respectively. These dissimilarities are most likely due to differences in the orientations of both the cationic nitrogen atom and the accompanying lipophilic portions of each QAC at the blastospore surface. Relating these observations to the known antiadherence effects of cetylpyridinium chloride and cetrimide, it can be concluded that monolayer coverage of the blastospore surface with QAC does not account for the observed reduced adherence. This suggests that the anti-adherence effects are due to either direct interaction with, or steric blockade of, adhesions on the blastospore surface.

Bioadhesive lozenge for the improved delivery of cetylpyridinium chloride.

Conventional lozenges produce a high initial release of drug in the oral cavity, which rapidly declines to subtherapeutic levels, and requires multiple daily administration with associated problems of systemic toxicity and compliance. Various multilayer compacts containing cetylpyridinium chloride were evaluated in vitro using release into simulated saliva (buffer pH 6.6). The drug loading, the wax content of the active layer, and the composition of the bioadhesive layer were important variables affecting product performance. Following preliminary in vivo studies, the release of a three-layered device containing drug in a nonadhesive and flavored waxy exposed layer was studied in six humans using HPLC and was shown not to be affected by location within the mouth. In comparison with a proprietary lozenge formulation, the device produced more uniform and effective levels of drug (approximately 20 micrograms.mL-1), with adequate comfort, taste, and irritancy over a period of 3 h.