From co-delivery to synergistic anti-inflammatory effect: Studies on chitosan-stabilized Janus emulsions having chloroquine phosphate and flavopiridol in Complete Freund's Adjuvant induced arthritis rat model.

For the first time, the co-delivery of chloroquine phosphate and flavopiridol by intra-articular route was achieved to provide local joint targeting in Complete Freund's Adjuvant-induced arthritis rat model. The presence of paired-bean structure onto the dispersed oil droplets of o/w nanosized emulsions allows efficient entrapment of two drugs (85.86-96.22 %). The dual drug-loaded emulsions displayed a differential in vitro drug release behavior, near normal cell viability in MTT assay, better cell uptake (internalization) and better reducing effect of mean immunofluorescence intensity of inflammatory proteins such as NF-κB and iNOS at in vitro RAW264.7 macrophage cell line. The radiographical study, ELISA test, RT-PCR study and H & E staining also indicated a reduction in joint tissue swelling, IL-6 and TNF-α levels diminution, fold change diminution in the mRNA expressions for NF-κB, IL-1ß, IL-6 and PGE2 and maintenance of near normal histology at bone cartilage interface respectively. The results of metabolomic pathway analysis performed by LC-MS/MS method using the rat blood (plasma) collected from disease control and dual drug-loaded emulsions treatment groups revealed a new follow-up study to understand not only the disease progression but also the formulation therapeutic efficacy assessment.

Development, systematic optimisation and biofilm disruption activity of eugenol-based nanosized emulsions stabilised with Tween 80.

The aims of this study were to systematically optimise a formula for eugenol emulsions via face-centered central composite design and to assess the activity against two-different bacterial strains (Staphylococcus aureus and Propionibacterium acnes) present at planktonic and biofilm forms. The molecular interaction of excipients, mean particle size (MPS) including zeta potential (ZP), drug entrapment efficiency (DEE) and in vitro drug release of optimised emulsions was done using FT-IR, Malvern Zetasizer, ultracentrifugation technique and membrane-free dissolution model, respectively. The emulsions consisted of 151.3 ± 1.45 nm MPS, -21.3 ± 1.25 mV ZP and 93.98 ± 1.41% DEE values. On storage of emulsions at 25 °C for 3 months, the value of DEE was found to be 72.12 ± 2.82%. The Tween 80 emulsifier film coverage onto the dispersed eugenol droplets of emulsions delayed significantly the drug release (12%-19%) compared to the drug release occurred from pure eugenol. The treatment of planktonic S. aureus and P. acnes with diluted eugenol emulsions showed the minimum inhibitory concentration and minimum bactericidal concentration values at 1.25-2.5 mg/ml whereas it occurred at 10 mg/ml for pure eugenol. Treating the biofilms with eugenol emulsions (1-2 mg/ml) yielded 59-70% minimum biofilm eradication concentration but 10 mg/ml pure eugenol showed 60%. Hence, the eugenol emulsions displayed antibacterial activity and could be projected as an antibiofilm or biofilm disruption agent.

Studies on spray dried topical ophthalmic emulsions containing cyclosporin A (0.05% w/w): systematic optimization, in vitro preclinical toxicity and in vivo assessments.

Cyclosporin A (CsA, 0.05% w/w)-loaded positively charged emulsions were prepared based on castor oil, chitosan, poloxamer 188, glycerin and double-distilled water. To augment the shelf/storage-stability of original emulsions, the solid-dry powder for reconstitution was made by spray drying technique. The screening (Taguchi OA) and optimization (face-centered central composite) designs produced the optimized conditions for spray drying: 40 Nm3/h aspirator flow rate, 15 ml/min feed rate, 115 °C inlet temperature, 10% mannitol and 1.25% trehalose. The % drug entrapment efficiency values of original and reconstituted emulsions ranged from 73.20 ± 0.13 to 71.55 ± 1.25%. At 20 min post-dissolution, two times higher CsA release was seen from reconstituted emulsions than the original emulsions (85.78 ± 1.14 vs. 42.25 ± 1.84%) in simulated tear fluid. Using MTT assay, the reconstituted emulsions with or without CsA produced 94.512 ± 2.12 to 99.941 ± 1.89% cell viability values in HCE-2 cells. No appreciable change in capillary integrity was visualized in HET CAM following reconstituted emulsions treatment. At equivalent 15 µg drug, the in vitro protein denaturation assay showed augmented inhibition value (~ 85%) for tested CsA emulsions compared to diclofenac reference (68.30 ± 2.05) indicating enhanced anti-inflammatory activity. The CsA concentrations in multiple ocular matrices of rabbit eyes determined by the UPLC-MS/MS method attained the therapeutic drug level of 50-300 ng/ml even at 90 min post-topical instillation of both emulsions. Overall, the CsA emulsion eyedrops can be supplied as a spray dried storable intermediate product for reconstitution.

Analytical Quality-by-Design-Based Systematic Optimization of RP-HPLC Method Conditions to Analyze Simultaneously Chloroquine Phosphate and Flavopiridol in Stress-Induced Combined Drug Solution and Pharmaceutical Emulsions.

A design of experiments (DoE)-driven RP-HPLC method conditions was employed to analyze simultaneously chloroquine (CQ) phosphate and flavopiridol (FLAP) in emulsions and solution. After subjecting the various critical method attributes to preliminary risk assessment and screening by Pareto-chart-based fractional factorial design, the 17 runs were produced in Box-Behnken design for optimization. Analysis of variance, lack of fit, prediction equations, 3D response surface plots and contour plots were used to evaluate the critical analytical attributes such as retention time, tailing factor and theoretical plate count. The optimized RP-HPLC method conditions include 262 nm as detection wavelength, 37°C temperature for column, 20-µl injection volume, 1-ml/min flow rate and mobile phase mixture [70:30 ratio of 0.4% triethylamine in methanol&sodium phosphate buffer (11 mM, pH 3.0)]. The studied validation parameters were found within the ICH-prescribed limits. Exposing the combined drug solution at oxidative stress condition resulted to diminish the FLAP recovery value (53.39 ± 0.86) and arrival of an extra chromatographic peak. However, the % drug entrapment efficiency values of 96.22 ± 2.47 and 85.86 ± 3.66, respectively, were noticed for CQ phosphate and FLAP in emulsions. Thus, DoE-driven approach could be helpful for systematically optimizing RP-HPLC method conditions.

Dual Delivery of Cyclosporin A and Etodolac Using Polymeric Nanocapsules in a Rabbit Eye Model: Ocular Biodistribution and Pharmacokinetic Study.

Purpose: Commercially available eye drops are loaded only with a single drug. By using the polymeric nanocapsules, dual delivery of 0.05% w/w cyclosporin A (CsA) and 0.2% w/w etodolac (Edc) was achieved. An ultraperformance liquid chromatography/tandem mass spectrometry (UPLC-MS/MS) method was developed for determining simultaneously the biodistribution and pharmacokinetic profile of CsA and Edc in ocular tissues. Methods: After one single drop instillation of nanocapsules into healthy right eyes of rabbits, the eyeballs were enucleated at 5, 15, 30, 60, and 90 min time periods to separate the 5 different ocular tissues. A liquid/liquid extraction method was used for ocular sample extraction using darunavir as internal standard. Using 3 diverse conditions such as bench-top, autosampler, and freeze-thaw, the stability of the analytes at 3 quality control samples in ocular tissues was also checked. Results: Intra- and interday precisions for both CsA and Edc in multiple ocular tissues were <10.32%, and the accuracy was <11.98%. The % bias and % RSD values for CsA and Edc were found within the acceptable limit of ±15%. The highest Cmax values were attained in cornea for both the drugs at 60 min postinstillation time point. Despite molecular size and structural differences, both CsA and Edc after liberation from nanocapsule drops can permeate into the tissues of the anterior as well as posterior segments of the eye. Conclusion: The biodistribution and pharmacokinetic data might help and strengthen our understanding of synergetic anti-inflammatory activity of CsA and Edc from nanocapsules after its ocular topical application for managing keratoconjunctivitis sicca.

Application of Design of Experiments® Approach-Driven Artificial Intelligence and Machine Learning for Systematic Optimization of Reverse Phase High Performance Liquid Chromatography Method to Analyze Simultaneously Two Drugs (Cyclosporin A and Etodolac) in Solution, Human Plasma, Nanocapsules, and Emulsions.

The objectives of current investigation are (1) to find out wavelength of maximum absorbance (λmax) for combined cyclosporin A and etodolac solution followed by selection of mobile phase suitable for the RP-HPLC method, (2) to define analytical target profile and critical analytical attributes (CAAs) for the analytical quality by design, (3) to screen critical method parameters with the help of full factorial design followed by optimization with face-centered central composite design (CCD) approach-driven artificial neural network (ANN)-linked with the Levenberg-Marquardt (LM) algorithm for finding the RP-HPLC conditions, (4) to perform validation of analytical procedures (trueness, linearity, precision, robustness, specificity and sensitivity) using combined drug solution, and (5) to determine drug entrapment efficiency value in dual drug-loaded nanocapsules/emulsions, percentage recovery value in human plasma spiked with two drugs and solution state stability analysis at different stress conditions for substantiating the double-stage systematically optimized RP-HPLC method conditions. Through isobestic point and scouting step, 205 nm and ACN:H2O mixture (74:26) were selected respectively as the λmax and mobile phase. The ANN topology (3:10:4) indicating the input, hidden and output layers were generated by taking the 20 trials produced from the face-centered CCD model. The ANN-linked LM model produced minimal differences between predicted and observed values of output parameters (or CAAs), low mean squared error and higher correlation coefficient values in comparison to the respective values produced by face-centered CCD model. The optimized RP-HPLC method could be applied to analyze two drugs concurrently in different formulations, human plasma and solution state stability checking.

Orally Administered Drug Solubility-Enhancing Formulations: Lesson Learnt from Optimum Solubility-Permeability Balance.

Although oral drug delivery is considered as most acceptable route for administering the active pharmaceutical ingredients to patients of all age-groups with the exceptions of bed-ridden patients and infants, the extent and rate of drug reaching the systemic circulation (in other word, drug bioavailability) always depends on many factors such as drug solubility in gastrointestinal fluids and drug permeation into intraluminal epithelial membrane structure, absence (fasting state) and presence (fed state) of food materials in the gastrointestinal tract, and individual variations in gastric emptying time. Taking the most influential factors like drug solubility and its permeability into consideration, these two factors play a pivotal role and even act as the litmus test for the formulation scientists who involve in oral dosage form development. It is very clear that there should be an optimum solubility and permeability balance to be reachable for getting the desired drug bioavailability to exert the intended therapeutic activity. The objectives of current review are (1) to provide an overview of two-different categories of poorly water soluble API molecules, (2) to describe briefly three-different case studies taken from drug solubility-enhancing formulations dealing with interplay between solubility and permeability, and (3) to showcase selected examples of solubility-permeability interplay phenomena arising out from the various orally administrable dosage forms. The lessons learnt from the past and current literatures are certainly encouraging to go ahead for oral dosage form development but with the prior knowledge about the possible existence of solubility-permeability interplay/tradeoff phenomenon.

Protecting the Normal Physiological Functions of Articular and Periarticular Structures by Aurum Nanoparticle-Based Formulations: an Up-to-Date Insight.

Taking the articular and periarticular structures as a litmus test for gold-based nanoformulations, the potential of gold nanoparticles in protecting the normal physiological functions of these structures particularly in geriatric patients is one of the research areas of current interest. Aside from its use to make the traditional and fashionable ornaments for human usage, the gold metal is also known for its rich therapeutic activity. This is especially true when the gold is converted from its bulk form into nanosized form before its administering into the human body. Since it is the age of nanocomponents in medical and pharmaceutical research areas, this review is therefore mainly focused on nanoparticulate systems consisting of aurum. Accumulating research reports nevertheless show concrete evidence indicating the potential of gold-based nanoformulations to manage joint syndromes such as osteoarthritis and rheumatoid arthritis. This review embarks from preparation techniques and characterization methods to therapeutical application potentials of gold-based nanoformulations.

Systematic Optimization, In Vitro Drug Release, and Preliminary Nonclinical Toxicity Assessment of Nonphospholipid-Based Topical Ophthalmic Emulsions Containing 0.05 or 0.1% w/w Cyclosporin A for Dry-Eye Syndrome Management.

The objectives of the present investigations are (1) to envisage a risk assessment plan for nonphospholipid-based topical ophthalmic emulsions with the help of failure mode and effect analysis (FMEA), (2) to screen the risky formulation and process variables by the Taguchi design, (3) to optimize systematically an emulsion formula by face-centered central composite design (CCD), (4) to incorporate cyclosporin A (0.05 or 0.1% w/w) into the optimized emulsions and predict the in vitro drug release kinetic via a particle diffusion-controlled mathematical model equation, and (5) to assess the emulsion's toxicity using in vitro hemolysis study. Through the risk priority number (RPN) scores of FMEA, half-normal and Pareto charts of the Taguchi design, 3D-response surface graphs, and overlay plots of CCD, the emulsion formula was systematically optimized. Irrespective of the two different drug loadings into optimized emulsions, the drug entrapment efficiency values ranged from 73.20 ± 0.13 to 74.42 ± 0.15%. The film diffusion or ion-exchange process fails to interpret the in vitro drug release kinetic profile. A permissible percentage hemolysis value of above 10% but below 25% guidance was observed for emulsions with or without cyclosporin A. The systematically optimized phospholipidless ophthalmic emulsions could further be exploited commercially for managing dry-eye syndrome.