Fixed-Dose Combination Formulations in Solid Oral Drug Therapy: Advantages, Limitations, and Design Features.

Whilst monotherapy is traditionally the preferred treatment starting point for chronic conditions such as hypertension and diabetes, other diseases require the use of multiple drugs (polytherapy) from the onset of treatment (e.g., human immunodeficiency virus acquired immunodeficiency syndrome, tuberculosis, and malaria). Successful treatment of these chronic conditions is sometimes hampered by patient non-adherence to polytherapy. The options available for polytherapy are either the sequential addition of individual drug products to deliver an effective multi-drug regimen or the use of a single fixed-dose combination (FDC) therapy product. This article intends to critically review the use of FDC drug therapy and provide an insight into FDC products which are already commercially available. Shortcomings of FDC formulations are discussed from multiple perspectives and research gaps are identified. Moreover, an overview of fundamental formulation considerations is provided to aid formulation scientists in the design and development of new FDC products.

Potential pharmacokinetic interactions with concurrent use of herbal medicines and a ritonavir-boosted COVID-19 protease inhibitor in low and middle-income countries.

The COVID-19 pandemic sparked the development of novel anti-viral drugs that have shown to be effective in reducing both fatality and hospitalization rates in patients with elevated risk for COVID-19 related morbidity or mortality. Currently, nirmatrelvir/ritonavir (Paxlovid™) fixed-dose combination is recommended by the World Health Organization for treatment of COVID-19. The ritonavir component is an inhibitor of cytochrome P450 (CYP) 3A, which is used in this combination to achieve needed therapeutic concentrations of nirmatrelvir. Because of the critical pharmacokinetic effect of this mechanism of action for Paxlovid™, co-administration with needed medications that inhibit or induce CYP3A is contraindicated, reflecting concern for interactions with the potential to alter the efficacy or safety of co-administered drugs that are also metabolized by CYP3A. Some herbal medicines are known to interact with drug metabolizing enzymes and transporters, including but not limited to inhibition or induction of CYP3A and P-glycoprotein. As access to these COVID-19 medications has increased in low- and middle-income countries (LMICs), understanding the potential for herb-drug interactions within these regions is important. Many studies have evaluated the utility of herbal medicines for COVID-19 treatments, yet information on potential herb-drug interactions involving Paxlovid™, specifically with herbal medicines commonly used in LMICs, is lacking. This review presents data on regionally-relevant herbal medicine use (particularly those promoted as treatments for COVID-19) and mechanism of action data on herbal medicines to highlight the potential for herbal medicine interaction Herb-drug interaction mediated by ritonavir-boosted antiviral protease inhibitors This work highlights potential areas for future experimental studies and data collection, identifies herbal medicines for inclusion in future listings of regionally diverse potential HDIs and underscores areas for LMIC-focused provider-patient communication. This overview is presented to support governments and health protection entities as they prepare for an increase of availability and use of Paxlovid™.

Permeation of Phytochemicals of Selected Psychoactive Medicinal Plants across Excised Sheep Respiratory and Olfactory Epithelial Tissues.

The intranasal route of drug administration offers an opportunity to bypass the blood-brain barrier and deliver compounds directly into the brain. Scientific evidence exists for medicinal plants (e.g., Centella asiatica and Mesembryanthemum tortuosum) to treat central nervous system conditions such as anxiety and depression. The ex vivo permeation of selected phytochemicals (i.e., asiaticoside and mesembrine) has been measured across excised sheep nasal respiratory and olfactory tissue. Permeation studies were conducted on individual phytochemicals and C. asiatica and M. tortuosum crude extracts. Asiaticoside exhibited statistically significantly higher permeation across both tissues when applied alone as compared to the C. asiatica crude extract, while mesembrine permeation was similar when applied alone or as M. tortuosum crude extract. Permeation of all the phytocompounds was similar or slightly higher than that of the drug atenolol across the respiratory tissue. Permeation of all the phytocompounds was similar to or slightly lower than that of atenolol across the olfactory tissue. In general, the permeation was higher across the olfactory epithelial tissue than across the respiratory epithelial tissue and therefore showed potential for direct nose-to-brain delivery of the selected psychoactive phytochemicals.

In vitro and ex vivo experimental models for evaluation of intranasal systemic drug delivery as well as direct nose-to-brain drug delivery.

The intranasal route of administration provides a noninvasive method to deliver drugs into the systemic circulation and/or directly into the brain. Direct nose-to-brain drug delivery offers the possibility to treat central nervous system diseases more effectively, as it can evade the blood-brain barrier. In vitro and ex vivo intranasal models provide a means to investigate physiological and pharmaceutical factors that could play a role in drug delivery across the nasal epithelium as well as to determine the mechanisms involved in drug absorption from the nose. The development and implementation of cost-effective pharmacokinetic models for intranasal drug delivery with good in vitro-in vivo correlation can accelerate pharmaceutical drug product development and improve economic and ecological aspects by reducing the time and costs spent on animal studies. Special considerations should be made with regard to the purpose of the in vitro/ex vivo study, namely, whether it is intended to predict systemic or brain delivery, source and site of tissue or cell sampling, viability window of selected model, and the experimental setup of diffusion chambers. The type of model implemented should suit the relevant needs and requirements of the project, researcher, and interlaboratory. This review aims to provide an overview of in vitro and ex vivo models that have been developed to study intranasal and direct nose-to-brain drug delivery.

Formulation of Chitosan Microparticles for Enhanced Intranasal Macromolecular Compound Delivery: Factors That Influence Particle Size during Ionic Gelation.

Therapeutic macromolecules (e.g., protein and peptide drugs) present bioavailability challenges via extravascular administration. The nasal route presents an alternative non-invasive route for these drugs, although low bioavailability remains challenging. Co-administration of permeation enhancers is a promising formulation approach to improve the delivery of poorly bioavailable drugs. The aim of this study was to prepare and characterize chitosan microparticulate formulations containing a macromolecular model compound (fluorescein isothiocyanate dextran 4400, FD-4) and a bioenhancer (piperine). Ionic gelation was used to produce chitosan microparticle delivery systems with two distinct microparticle sizes, differing one order of magnitude in size (±20 µm and ±200 µm). These two microparticle delivery systems were formulated into thermosensitive gels and their drug delivery performance was evaluated across ovine nasal epithelial tissues. Dissolution studies revealed a biphasic release pattern. Rheometry results demonstrated a sol-to-gel transition of the thermosensitive gel formulation at a temperature of 34 °C. The microparticles incorporating piperine showed a 1.2-fold increase in FD-4 delivery across the excised ovine nasal epithelial tissues as compared to microparticles without piperine. This study therefore contributed to advancements in ionic gelation methods for the formulation of particulate systems to enhance macromolecular nasal drug delivery.

Comparison of RPMI 2650 cell layers and excised sheep nasal epithelial tissues in terms of nasal drug delivery and immunocytochemistry properties.

Nasal drug administration has been identified as a potential alternative to oral drug administration, especially for systemic delivery of large molecular weight compounds. Major advantages of nasal drug delivery include high vascularity and permeability of the epithelial membranes as well as circumvention of first-pass metabolism. RPMI 2650 cell layers (in vitro cell model) and excised sheep nasal mucosal tissues (ex vivo sheep model) were evaluated with regard to epithelial thickness, selected tight junction protein expression (i.e. claudin-1, F-actin chains, zonula occludin-1), extent of p-glycoprotein (P-gp) related efflux of a model compound (Rhodamine-123, R123) and paracellular permeation of a large molecular weight model compound (FITC-dextran 4400, FD4). The cell model grown under liquid cover conditions (LCC) was thinner (24 ± 4 µm) than the epithelial layer of the sheep model (53 ± 4 µm), whereas the thickness of cell model grown under air liquid interface (ALI) conditions (53 ± 8 µm) compared well with that of the sheep model. Although the location and distribution of tight junction proteins and F-actin differed to some extent between the cell model grown under ALI conditions and the sheep model, the extent of paracellular permeation of FD4 was similar (Papp = 0.48 × 10-6 cm.s-1 and 0.46 × 10-6 cm.s-1, respectively). Furthermore, the bi-directional permeation of R123 yielded the same efflux ratio (ER = 2.33) in both models. The permeation results from this exploratory study indicated similarity in terms of compound permeation between the RPMI 2650 nasal epithelial cell line and the excised sheep nasal epithelial tissue model.

Molecular mechanisms and associated cell signalling pathways underlying the anticancer properties of phytochemical compounds from Aloe species (Review).

Naturally occurring components from various species of Aloe have been used as traditional folk medicine since the ancient times. Over the last few decades, the therapeutic effects of extracts and phytochemical compounds obtained from Aloe vera have been proven in preclinical and clinical studies. Recently, compounds from other Aloe species apart from Aloe vera have been investigated for the treatment of different diseases, with a particular focus on cancer. In the present review, the effects of phytochemical compounds obtained from different Aloe species are discussed, with a specific focus on the effects on cell signalling in cancer and normal cells, and their selectivity and efficacy. This information will be useful for the application of Aloe-derived compounds as therapeutic agents, either alone or in combination with other standard drugs for cancer treatment.

Formulation of Medicated Chewing Gum Containing Sceletium tortuosum and Process Optimization Utilizing the SeDeM Diagram Expert System.

Sceletium tortuosum is one of the most promising medicinal plant species for treating anxiety and depression. Traditionally, aerial parts are chewed (masticatory herbal medicine) providing fast relief and rendering the masticatory route for delivery, ideal. This study intended formulating novel medicated chewing gum containing S. tortuosum to alleviate depression and anxiety. S. tortuosum extract was formulated into directly compressed medicated chewing gum (MCG) containing different Health-in-Gum® (HIG) bases through process optimization with the SeDeM Diagram Expert System. Physical properties of MCGs were characterized, and specialized drug release studies performed. According to the manufacturer, only HIG-03 was specifically developed for direct compression; however, the SeDeM System was successfully applied to all HIG-bases investigated. HIG-01 and HIG-04 are also considered useful in direct compression as no considerable differences in these MCG formulations' physical properties were recognized. Inclusion of a lubricant, however, is deemed essential, and MCG comprising HIG-01, most suited for direct compression. Dissolution experiments found only two alkaloids used as markers, mesembrine and mesembrenone, were released in quantifiable concentrations regardless formulation constituents. Novel directly compressed MCG-containing S. tortuosum extract was successfully formulated by which the biologically active phytochemicals of S. tortuosum can be scientifically delivered through the traditionally applied mastication method.

Physicochemical Stability of Enriched Phenolic Fractions of Cyclopia genistoides and ex vivo Bi-directional Permeability of Major Xanthones and Benzophenones.

Fractions of an ultrafiltered Cyclopia genistoides extract, respectively enriched in xanthones and benzophenones, were previously shown to inhibit mammalian α-glucosidase in vitro. The present study investigated ex vivo intestinal transport of these fractions, using excised porcine jejunal tissue, to determine whether the gut could be a predominant in vivo site of action. The major bioactive compounds, the xanthones (mangiferin, isomangiferin) and benzophenones (3-ß-D-glucopyranosyliriflophenone, 3-ß-D-glucopyranosyl-4-O-ß-D-glucopyranosyliriflophenone) exhibited poor permeation in the absorptive direction with a relatively high efflux ratio (efflux ratio > 1). The efflux ratio of 3-ß-D-glucopyranosyl-4-O-ß-D-glucopyranosyliriflophenone (3.05) was similar to rhodamine 123 (2.99), a known substrate of intestinal P-glycoprotein 1 efflux transporters. Low epithelial membrane transport rates, coupled with efflux mechanisms, would effectively concentrate these bioactive compounds at the target site (gut lumen). Storage stability testing and moisture sorption assays of the xanthone-enriched fraction, benzophenone-enriched fraction, and ultrafiltered Cyclopia genistoides extract were performed to determine their susceptibility to physical and chemical degradation during storage. Hygroscopicity of the powders, indicated by moisture uptake, decreased in the order: benzophenone-enriched fraction (22.7%) > ultrafiltered Cyclopia genistoides extract (14.0%) > xanthone-enriched fraction (10.7%). 3-ß-D-Glucopyranosylmaclurin, a minor benzophenone, was the least stable of the compounds, degrading faster in the benzophenone-enriched fraction than in ultrafiltered Cyclopia genistoides extract, suggesting that the ultrafiltered extract matrix may provide a degree of protection against chemical degradation. Compound degradation during 12 wk of storage at 40 °C in moisture-impermeable containers was best explained by first order reaction kinetics.

Characterization of an Alginate Encapsulated LS180 Spheroid Model for Anti-colorectal Cancer Compound Screening.

Colorectal cancer is one of the leading causes of cancer-related deaths. A main problem for its treatment is resistance to chemotherapy, requiring the development of new drugs. The success rate of new candidate cancer drugs in clinical trials remains dismal. Three-dimensional (3D) cell culture models have been proposed to bridge the current gap between in vitro chemotherapeutic studies and the human in vivo, due to shortcomings in the physiological relevance of the commonly used two-dimensional cell culture models. In this study, LS180 colorectal cancer cells were cultured as 3D sodium alginate encapsulated spheroids in clinostat bioreactors. Growth and viability were evaluated for 20 days to determine the ideal experimental window. The 3- (4,5- dimethylthiazol- 2- yl)-2,5-diphenyltetrazolium bromide assay was then used to establish half maximal inhibitory concentrations for the standard chemotherapeutic drug, paclitaxel. This concentration was used to further evaluate the established 3D model. During model characterization and evaluation soluble protein content, intracellular adenosine triphosphate levels, extracellular adenylate kinase, glucose consumption, and P-glycoprotein (P-gp) gene expression were measured. Use of the model for chemotherapeutic treatment screening was evaluated using two concentrations of paclitaxel, and treatment continued for 96 h. Paclitaxel caused a decrease in cell growth, viability, and glucose consumption in the model. Furthermore, relative expression of P-gp increased compared to the untreated control group. This is a typical resistance-producing change, seen in vivo and known to be a result of paclitaxel treatment. It was concluded that the LS180 sodium alginate encapsulated spheroid model could be used for testing new chemotherapeutic compounds for colorectal cancer.

The Role of Functional Excipients in Solid Oral Dosage Forms to Overcome Poor Drug Dissolution and Bioavailability.

Many active pharmaceutical ingredients (APIs) exhibit poor solubility and low dissolution rates in aqueous environments such as the luminal fluids of the gastrointestinal tract. The oral bioavailability of these compounds is usually very low as a result of their poor solubility properties. In order to improve the bioavailability of these poorly soluble drugs, formulation strategies have been applied as a means to improve their aqueous solubility and dissolution rates. With respect to formulation approaches, excipients can be incorporated in the formulation to assist in the dissolution process of the drug, or specialized dosage forms can be formulated that improve dissolution rate through various mechanisms. This paper provides an overview of selected excipients (e.g., alkalinizing agents, surfactants and sugars) that can be used in formulations to increase the dissolution rate as well as specialized dosage forms such as self-emulsifying delivery systems and formulation techniques such as inclusion complexes and solid dispersions. These formulation approaches are discussed with available examples with specific reference to positive outcomes in terms of drug solubility and bioavailability enhancement.

Artificial membranes in combination with selected natural oils for in vitro drug passive diffusion screening in Ussing type chamber apparatus applied to gastro-retentive systems.

This study aimed at developing an effective in vitro technique for the screening of drug passive diffusion utilising artificial membranes in combination with three selected oils (i.e. cognac, emu, and olive oil). Artificial membranes of varying chemical composition and characteristics have been investigated individually and in combination with the selected oils in terms of the passive diffusion of a fluorescent probe (i.e. Rhodamine 6G or R6G), in a diffusion apparatus as compared to excised pig intestinal tissues. In general, the permeation results showed that the rate and extent of R6G permeation were dependent on the membrane composition as well as the type of oil used. The apparent permeability coefficient (Papp) value for R6G across the cellulose nitrate membrane (0.197 × 10-7 ± 0.069 cm/s) was the closest to the Papp of R6G across the excised pig intestinal tissue (0.210 × 10-7 ± 0.080 cm/s). The cellulose acetate-nitrate mixture membrane impregnated with emu oil also produced a Papp value (0.191 × 10-7 ± 0.010 cm/s) that was relatively close to that of R6G across the excised pig intestinal tissue. The delivery of R6G from gastro-retentive matrix type tablets correlated with the release of R6G from the gastro-retentive tablets.

Models used to screen for the treatment of multidrug resistant cancer facilitated by transporter-based efflux.

PURPOSE: Efflux transporters of the adenosine triphosphate-binding cassette (ABC)-superfamily play an important role in the development of multidrug resistance (multidrug resistant; MDR) in cancer. The overexpression of these transporters can directly contribute to the failure of chemotherapeutic drugs. Several in vitro and in vivo models exist to screen for the efficacy of chemotherapeutic drugs against MDR cancer, specifically facilitated by efflux transporters. RESULTS: This article reviews a range of efflux transporter-based MDR models used to test the efficacy of compounds to overcome MDR in cancer. These models are classified as either in vitro or in vivo and are further categorised as the most basic, conventional models or more complex and advanced systems. Each model's origin, advantages and limitations, as well as specific efflux transporter-based MDR applications are discussed. Accordingly, future modifications to existing models or new research approaches are suggested to develop prototypes that closely resemble the true nature of multidrug resistant cancer in the human body. CONCLUSIONS: It is evident from this review that a combination of both in vitro and in vivo preclinical models can provide a better understanding of cancer itself, than using a single model only. However, there is still a clear lack of progression of these models from basic research to high-throughput clinical practice.

Capsaicin and Piperine as Functional Excipients for Improved Drug Delivery across Nasal Epithelial Models.

The fruit from various pepper plants has been employed for the seasoning of food, as perfuming agents, and also as traditional medicines. Phytochemicals isolated from different pepper species have been found to modulate the pharmacokinetics of orally administered drugs. This study investigated the possibility to apply capsaicin and piperine (extracted alkaloids) as modulators for drug delivery across the nasal epithelium. Both a nasal epithelial cell line (RPMI 2650) and excised sheep nasal tissue were used as models to investigate the effects of the selected pepper compounds on drug permeation. FITC-dextran 4400 (MW 4400 Da) was used as a large molecular weight marker compound for paracellular transport, while rhodamine 123 was used as a marker compound that is a substrate for P-glycoprotein-mediated efflux. From the permeation results, it was clear that capsaicin inhibited P-glycoprotein efflux to a larger extent, while piperine showed drug permeation enhancement via other mechanisms. The cell cytotoxicity studies indicated that capsaicin was noncytotoxic up to a concentration of 200 µM and piperine up to a concentration of 500 µM as indicated by cell viability above 80%. The histological analysis of the excised nasal tissue and cultured RPMI 2650 cell layers indicated that some damage occurred after treatment with 200 µM capsaicin, but no changes were observed for piperine up to a concentration of 50 µM.

Treatment of Skin Disorders with Aloe Materials.

The skin is the largest organ and functions as a barrier to protect the underlying tissues against the elements and pathogens, while also fulfilling many physiological roles and biochemical functions such as preventing excessive water loss. Skin disorders vary greatly in terms of origin, severity, symptoms and affect persons of all ages. Many plants have been used for medicinal purposes since ancient times including the treatment of skin disorders and diseases. Aloe represents one of the earliest medicinal plant species mentioned in antique scriptures and even in rock art dating back thousands of years. Different Aloe species and materials have been used in the prevention and treatment of skin related disorders. Aloe vera is the most commonly used Aloe species for medicinal purposes. Some of the most prominent skin related applications and disorders that Aloe materials have been investigated for are discussed in this paper, which include cosmetic, radiation, cancer, wound and antimicrobial applications. Both in vitro and in vivo studies are included in the discussions of this paper and comprehensive summaries of all these studies are given in tables in each section. Although some contradictory results were obtained among studies, certain Aloe materials have shown excellent efficacy and exhibited potential for the treatment of skin related disorders and cosmetic applications.

A sub-chronic Xysmalobium undulatum hepatotoxicity investigation in HepG2/C3A spheroid cultures compared to an in vivo model.

ETHNOPHARMACOLOGY RELEVANCE: Traditional herbal medicines are utilized by 27 million South Africans. Xysmalobium undulatum (Uzara) is one of the most widely used traditional medicinal plants in Southern Africa. A false belief in the safety of herbal medicine may result in liver injury. Herb-induced liver injury (HILI) range from asymptomatic elevation of liver enzymes, to cirrhosis and in certain instances even acute liver failure. Various in vitro and in vivo models are available for the pre-clinical assessment of drug and herbal hepatotoxicity. However, more reliable and readily available in vitro models are needed, which are capable of bridging the gap between existing models and real human exposure. Three-dimensional (3D) spheroid cultures offer higher physiological relevance, overcoming many of the shortcomings of traditional two-dimensional cell cultures. AIMS OF THIS STUDY: This study investigated the hepatotoxic and anti-prolific effects of the crude X. undulatum aqueous extract during a sub-chronic study (21 days), in both a 3D HepG2/C3A spheroid model and the Sprague Dawley rat model. METHODS: HepG2/C3A spheroids were treated with a known hepatotoxin, valproic acid, and crude X. undulatum aqueous extract for 21 days with continuous evaluation of cell viability and proliferation. This was done by evaluating cell spheroid growth, intracellular adenosine triphosphate (ATP) levels and extracellular adenylate kinase (AK). Sprague Dawley rats were treated with the same compounds over 21 days, with evaluation of in vivo toxicity effects on serum chemistry. RESULTS: The results from the in vitro study clearly indicated hepatotoxic effects and possible liver damage following treatment with valproic acid, with associated growth inhibition, loss of cell viability and increased cytotoxicity as indicated by reduced intracellular ATP levels and increased AK levels. These results were supported by the increased in vivo levels of AST, ALT and LDH following treatment of the Sprague Dawley rats with valproic acid, indicative of hepatic cellular damage that may result in hepatotoxicity. The in vitro 3D spheroid model was also able to predict the potential concentration dependant hepatotoxicity of the crude X. undulatum aqueous extract. Similarly, the results obtained from the in vivo Sprague Dawley model indicated moderate hepatotoxic potential. CONCLUSION: The data from both the 3D spheroid model and the Sprague Dawley model were able to indicate the potential concentration dependant hepatotoxicity of the crude X. undulatum aqueous extract. The results obtained from this study also confirmed the ability of the 3D spheroid model to effectively and reliably predict the long-term outcomes of possible hepatotoxicity.

Drug Bioavailability Enhancing Agents of Natural Origin (Bioenhancers) that Modulate Drug Membrane Permeation and Pre-Systemic Metabolism.

Many new chemical entities are discovered with high therapeutic potential, however, many of these compounds exhibit unfavorable pharmacokinetic properties due to poor solubility and/or poor membrane permeation characteristics. The latter is mainly due to the lipid-like barrier imposed by epithelial mucosal layers, which have to be crossed by drug molecules in order to exert a therapeutic effect. Another barrier is the pre-systemic metabolic degradation of drug molecules, mainly by cytochrome P450 enzymes located in the intestinal enterocytes and liver hepatocytes. Although the nasal, buccal and pulmonary routes of administration avoid the first-pass effect, they are still dependent on absorption of drug molecules across the mucosal surfaces to achieve systemic drug delivery. Bioenhancers (drug absorption enhancers of natural origin) have been identified that can increase the quantity of unchanged drug that appears in the systemic blood circulation by means of modulating membrane permeation and/or pre-systemic metabolism. The aim of this paper is to provide an overview of natural bioenhancers and their main mechanisms of action for the nasal, buccal, pulmonary and oral routes of drug administration. Poorly bioavailable drugs such as large, hydrophilic therapeutics are often administered by injections. Bioenhancers may potentially be used to benefit patients by making systemic delivery of these poorly bioavailable drugs possible via alternative routes of administration (i.e., oral, nasal, buccal or pulmonary routes of administration) and may also reduce dosages of small molecular drugs and thereby reduce treatment costs.

Intestinal Drug Absorption Enhancement by Aloe vera Gel and Whole Leaf Extract: In Vitro Investigations into the Mechanisms of Action.

The co-administration of absorption enhancing agents with macromolecular drugs (e.g., protein and peptide drugs) has been identified as a means to improve the oral bioavailability of these drugs. Absorption-enhancing agents of natural origins have received a great deal of attention due to their sustainable production, in support of green chemistry. In previous studies, certain parts of the Aloe vera leaf (e.g., gel and whole leaf extract) have shown a potential to enhance drug permeation across the intestinal epithelial barrier. The mechanism of the drug-absorption-enhancement action and the capacity for absorption-enhancement of the A. vera gel and whole leaf, were investigated in this study. A clear decrease in transepithelial electrical resistance (TEER) of Caco-2 cell monolayers exposed to A. vera gel and wholeleaf extract, in various concentrations, indicated the opening of tight junctions between the epithelial cells. The transport of Fluorescein isothiocyanate (FITC)-dextran, with a molecular weight of 4 kDa (FD-4), could be enhanced across the Caco-2 cell monolayers, by the A. vera gel and whole-leaf extract, but not the FITC-dextran with larger molecular weights (i.e., 10, 20, and 40 kDa), which indicated a limited drug absorption enhancement capacity, in terms of the molecular size. Accumulation of FD-4 between the Caco-2 cells (and not within the cells), after treatment with the A. vera gel and whole-leaf extract was shown with a confocal laser scanning microscopy (CLSM) imaging, indicating that the paracellular transport of FD-4 occurred after the interaction of the A. vera gel and whole-leaf extract, with the epithelial cell monolayers. Furthermore, changes in the F-actin distribution in the cytoskeleton of the Caco-2 cell monolayers was observed by means of a fluorescence staining, which confirmed tight junction modulation as the mechanism of action for the absorption enhancement effect of the A. vera gel and whole-leaf extract.