Abstract by Ziwei Yu

Nanoemulsions (NE) for intestinal drug delivery are predominantly oil-in-water dispersions in which an oil phase is distributed as nanosized droplets in an aqueous phase. They can be prepared through high- or low-energy approaches and have attracted considerable interest as delivery systems for improving the absorption of poorly water-soluble drugs across different intestinal segments. In NE development, polyethylene glycol (PEG)-ylated surfactants are commonly used to stabilize the droplet surface and modify their physicochemical properties. Such PEGylation may improve mucus permeation and protect droplets against destabilization, while potentially limiting drug release or interactions with the epithelium. Moreover, changing the content of PEGylated surfactants commonly alters other NE properties simultaneously, including droplet size, surface composition, and drug-loading capacity. These complex effects of PEGylated surfactants make their role in the intestinal absorption of drug from NE remain insufficiently understood. This thesis aimed to investigate the rational design of NE with a PEGylated surfactant for intestinal absorption of celecoxib (CCX).

In this study, NE were developed by dispersing the preconcentrates of self-nanoemulsifying drug delivery systems (SNEDDS) containing the model PEGylated surfactant polyoxyl-40 hydrogenated castor oil (PEG40HCO), together with the co-surfactant glyceryl mono- and dicaprylocaprate (GMD) and the oil medium-chain triglycerides (MCT). Surface PEGylation was introduced as an operational parameter describing the relative contribution of PEG40HCO to the PEG40HCO/GMD composition at the droplet surface. First, twelve blank NE (NE-blank) were developed using design of experiments (DoE). They were classified into four predefined size ranges: small (S, ~30 nm), medium-small (MS, ~50 nm), medium-large (ML, ~100 nm), and large (L, ~130 nm). Within each size category, three NE (1, 2 and 3) were prepared with different PEG40HCO contents. Developed NE preconcentrates contained 12-58% PEG40HCO, corresponding to surface PEGylation of 40-84%. Two sets of NE, NE-5 or NE-70, were prepared by loading 5 or 70 mg/g CCX to preconcentrates to consider the drug-loading influence on NE properties. Secondly, another three NE were developed with a particular focus on surface PEGylation. By keeping the total PEG40HCO content constant at 60%, F67, F80 and F100 were prepared with surface PEGylation levels of 67%, 80% and 100%, respectively. These three NE showed comparable droplet size, CCX load and saturation level. All the developed NE showed acceptable short-term stability in relevant media.

NE-5 and NE-70 were first studied under the relatively simple and low-enzyme colonic conditions to identify the PEG40HCO-related property most relevant to absorption. CCX release and mucus permeation were evaluated using rapid equilibrium dialysis (RED) and Transwell-based permeation model, respectively. Pearson correlation analysis was applied to assess the relationships between NE properties and their biopharmaceutical performance. CCX release from NE was found to consistently increase with lower surface PEGylation. In contrast, a higher PEG40HCO content contributed more significantly to enhanced CCX permeation flux through porcine colonic mucus. Although CCX loading altered droplet sizes and drug saturation, NE-5 and NE-70 exhibited generally similar trends in in vitro performance. However, in vivo pharmacokinetics (PK) studies after colonic administration of selected NE-70 showed that both C_max and AUC were significantly negatively correlated with surface PEGylation. Principal component analysis (PCA) further supported a positive association between PK performance and drug release, and a weak association with mucus permeation. These findings indicate that drug release played a greater role in CCX absorption than mucus permeation. Surface PEGylation is an important NE property, and higher surface PEGylation may limit CCX release and reduce CCX colonic absorption.

F67, F80 and F100 were subsequently studied under lipolysis-relevant conditions to specifically investigate how surface PEGylation influences the small intestinal absorption of CCX. Using a dynamic digestion model, the behavior of CCX and NE under lipolysis and the subsequent mucus permeation were evaluated, with the colloidal transformation visualized by cryo-TEM. Increasing surface PEGylation was observed to progressively suppress lipolysis, with F67 undergoing the fastest digestion, F80 showing a delayed onset of lipolysis, and F100 exhibiting the most inhibited lipolysis. This lipolysis resistance helped maintain CCX solubilization and preserve droplet integrity. Samples of F100 collected as lipolysis proceeded showed progressively enhanced permeation through porcine small intestinal mucus, whereas F67 and F80 showed fluctuating or reduced permeation during lipolysis. However, these in vitro advantages of higher surface PEGylation did not translate into improved oral absorption in rats. Instead, F67 achieved significantly higher C_max and AUC_0-1.5h than F100, while F80 showed intermediate performance. These results suggest that intestinal absorption was more closely associated with an early onset of lipolysis than with maintained solubilization or enhanced mucus permeation. Co-distribution of GMD on the droplet surface may therefore play an important role, whereas excessive surface PEGylation may be unfavorable for small intestinal absorption of CCX from NE.

Given the limited association between improved mucus permeation and absorption indicated in the above studies, the role of small intestinal mucus barrier was further evaluated using confocal imaging on ex vivo mouse small intestine tissue and mouse model with different mucus integrity. Nile Red (NR)-labelled F67, F80 and F100 showed comparable permeation across the mucus layer and redistribution toward the intestinal villi within 30 min. After oral administration of the CCX suspension or F80, CCX absorption was generally higher in wild-type male mice than in female mice. Plasma CCX concentrations at 1 h post-dosing showed no significant differences among mice with normal (MUC2^+/+), partially absent (MUC2^+/-) and absent (MUC2^-/-) mucus layers. These findings were unlikely to be confounded by inflammation related to mucus deficiency. The results further indicated that the small intestinal mucus layer was not the dominant barrier controlling CCX absorption for the investigated NE systems.

Overall, this thesis demonstrates that PEG40HCO, particularly its contribution to surface PEGylation, plays an important role in adjusting intestinal CCX absorption from NE. Rational design of oral NE should balance the benefits of PEGylated surfactants with their potential limitations.

so is the conclusion that releasse is more important the rate of mucus permeation?

what does this mean?