Researchers at the University of San Carlos and collaborators have prepared a paracetamol-β-cyclodextrin inclusion complex using a co-precipitation method, aiming to address structural and organoleptic challenges associated with high-dose orodispersible films. The study, published in AAPS Open, characterizes the solid-state properties of the complex and evaluates its antipyretic activity in a rat model. The primary finding is that while the complex successfully forms a new crystalline phase, its regulated drug release profile results in diminished pharmacologic activity at later stages compared to uncomplexed paracetamol, highlighting a trade-off between molecular shielding and therapeutic efficacy.
The motivation for this work stems from the widespread clinical use of paracetamol, also known as acetaminophen, for managing fever and mild to moderate pain. Despite its utility, conventional oral delivery faces significant biopharmaceutical and organoleptic challenges, particularly for pediatric and geriatric populations. Modern patient-centric dosage forms, such as orodispersible films, often struggle with paracetamol due to its high therapeutic dose and moderate aqueous solubility, which can exceed the loading capacity of thin-film matrices. This leads to recrystallization and mechanical failure, while the drug’s intrinsic bitterness necessitates taste-masking strategies to ensure compliance.
Cyclodextrins, water-soluble oligosaccharides with a hydrophobic inner cavity and hydrophilic outer surface, have emerged as promising excipients to overcome these limitations. Specifically, β-cyclodextrin offers an ideal spatial fit for the phenolic ring of paracetamol, favoring the formation of stable stoichiometric inclusion complexes. The co-precipitation method was selected for its simplicity and efficiency, involving the mixing of equimolar solutions of paracetamol in petroleum ether and β-cyclodextrin in distilled water. The mixture was maintained at 4 °C overnight, followed by vacuum filtration and drying, a process conducive to paracetamol, which is prone to thermal degradation.
Characterization of the co-precipitates revealed a flaky, white, non-hygroscopic powder with an encapsulation efficiency of 41.94% ± 4.41%. Differential scanning calorimetry measurements showed that the characteristic endothermic melting peak of pure paracetamol at 169 °C disappeared in the complex, indicating a transition from a native crystalline lattice to a molecularly dispersed state within the β-cyclodextrin cavity. Fourier transform infrared spectroscopy further supported this interaction, showing shifts in carbonyl peaks and broadening of hydroxyl peaks, suggesting that intermolecular hydrogen bonds within the paracetamol dimer dissociated upon complex formation.
Nuclear magnetic resonance spectroscopy provided evidence of a 1:1 stoichiometric molar ratio between paracetamol and β-cyclodextrin. The 1H-NMR spectra revealed distinct chemical shifts in the aromatic protons of paracetamol, indicating reduced conformational freedom and electronic shielding within the hydrophobic cavity. Two-dimensional NOESY measurements confirmed the guest-in-host geometry, with intense cross-peaks between the aromatic protons of paracetamol and the H-3 and H-5 protons of β-cyclodextrin, confirming that the phenolic ring was deeply sequestered while the acetamide group remained near the wider rim.
Powder X-ray diffraction analysis provided the first molecular evidence of a channel-type crystalline habit for the complex. While the physical mixture displayed a superimposition of individual components’ diffractograms, the co-precipitates revealed an entirely new diffraction pattern with peaks at 2θ = 11.9º, 15.3º, and 25.9º. This unique crystalline phase suggests that paracetamol is stabilized within the β-cyclodextrin channel, disrupting its native state to prevent recrystallization during drying, which is essential for the mechanical integrity of high-dose orodispersible films.
In vitro dissolution studies in phosphate buffer at pH 7.4 and 37 °C revealed a regulated, biphasic drug release profile. The complex achieved a cumulative drug release of 81.41% ± 11.31% after 60 minutes, compared to 107.11% ± 6.42% for uncomplexed paracetamol. A higher cumulative release from the complex was observed only at the first minute, indicating an initial burst effect. However, the dissolution efficiency at 30 minutes was significantly lower for the complex at 66.06% ± 6.92% compared to 90.36% ± 5.69% for uncomplexed paracetamol, with a p-value of 0.003.
The antipyretic activity was evaluated in Sprague-Dawley rats with brewer’s yeast-induced pyrexia. Pyrexia was induced by subcutaneous injection of a 20% aqueous suspension of brewer’s yeast, and rats with a rectal temperature increase of at least 0.5 °C after 18 hours were included in the study. Groups received oral doses of uncomplexed paracetamol at 25, 75, and 150 mg/kg, or equivalent doses of the co-precipitate at 213, 639, and 1278 mg/kg. Rectal temperatures were measured at one-hour intervals up to five hours to calculate the percent reduction of pyrexia.
Results showed that all groups exhibited a reduction in temperature within the first hour, consistent with the initial burst release observed in vitro. However, the temperature reduction and antipyretic activity observed with the co-precipitate across all doses and time points were not significant when compared to the negative control. There was also no significant difference between equivalent doses of uncomplexed paracetamol and the complex across all time points, indicating that the regulated release from the complex resulted in subtherapeutic levels of paracetamol at later stages of the assay.
The discussion highlights that the channel-type habit creates a substantial kinetic barrier, as paracetamol molecules sequestered deep within the hydrophobic channels are shielded from rapid solvent access. This structural arrangement reduces the fraction of free, non-complexed drug available for passive diffusion, which is advantageous for preventing dose-dumping and maintaining paracetamol levels below the bitterness threshold in the oral cavity. However, the limited drug reservoir and reduced free fraction likely resulted in systemic availability falling below the minimum effective concentration required for robust fever suppression in this model.
The study acknowledges that the encapsulation efficiency indicates a significant portion of paracetamol remained uncomplexed and was lost during manufacturing, contributing to the diminished pharmacologic activity. The relationship between encapsulation efficiency and pharmacologic properties remains unclear due to conflicting reports in the literature. The authors note that the study was not primarily aimed at enhancing the intrinsic solubility of paracetamol, which already has an acceptable aqueous solubility profile, but rather at addressing formulation challenges in high-dose orodispersible films.
These findings provide a foundational blueprint for stabilizing biopharmaceutics classification system class III drugs in space-constrained platforms. The partial inclusion mechanism offers a strategy for taste masking without sacrificing aqueous interaction, as the bitter-tasting aromatic core is molecularly shielded while polar groups remain accessible. However, the regulated release acted as a kinetic barrier to peak clinical efficacy in this instance, underscoring the necessity of optimizing critical process parameters to improve encapsulation efficiency and calibrate the drug-to-cyclodextrin molar ratio.
Future research should focus on pharmacokinetic profiling of the paracetamol-β-cyclodextrin system to better understand the dissociation kinetics in a biological environment. The study validates the feasibility of β-cyclodextrin-based stabilization for paracetamol orodispersible films while highlighting the need to balance structural stability with therapeutic performance. The identification of a critical trade-off between molecular shielding and kinetic barriers to optimal clinical efficacy provides a baseline for the rational design of stabilized, patient-centric formulations.
The study was conducted under the oversight of the University of San Carlos Institutional Animal Care and Use Committee, which approved the protocol for the animal experiments. The research team utilized Sprague-Dawley rats that were acclimatized for twelve hours under standard laboratory conditions prior to the induction of pyrexia. The antipyretic assay followed a method described by Muhammad et al., with minor modifications tailored to the specific co-precipitate formulation.
Statistical analysis of the data was performed using Design-Expert v23.1 software, with results expressed as mean values plus or minus the standard deviation. To compare differences among the characterization results of the co-precipitates and pure paracetamol across experimental groups, the researchers employed Tukey’s HSD test following a one-way analysis of variance. A p-value of less than 0.05 was established as the threshold for statistical significance.
Paracetamol was obtained from Xi’an Biohorlden Industry & Trade Co., Ltd, while β-cyclodextrin was sourced from TCI Tokyo Chemical Industry. All other necessary materials for the preparation process were provided by the Department of Pharmacy at the University of San Carlos. The co-precipitation procedure itself was adapted from the work of El-Kemary et al., involving the slow addition of a paracetamol solution in petroleum ether into a β-cyclodextrin solution in distilled water. This mixture was agitated with a magnetic stirrer and maintained at 4 °C overnight to facilitate crystallization and precipitation.
Following the overnight incubation, the co-precipitates were isolated through vacuum filtration and subsequently dried in a desiccator before storage. The resulting product appeared as a flaky, white, and light non-hygroscopic powder. The drying process was kept relatively brief compared to other techniques, a consideration that is particularly conducive to paracetamol, which is prone to thermal degradation. This methodological choice helps preserve the integrity of the drug substance during the formation of the new solid crystalline phase, which is critical for maintaining the physicochemical stability of the encapsulated paracetamol.
The in vitro dissolution tests were conducted using a DR-2 Thermonik paddle-type dissolution tester. An equivalent amount of the inclusion complex containing 10 mg of paracetamol was placed in a vessel with 900 mL of phosphate buffer at pH 7.4 and a temperature of 37 ± 0.50 °C. Aliquots of 5 mL were collected at specific time points and filtered through an 8 µm membrane filter. Quantification of the released drug was performed using a UV–Vis spectrophotometer at 257 nm. The dissolution profiles were evaluated based on cumulative percent release and the percent dissolution efficiency at 30 minutes, providing a detailed kinetic profile of the drug release from the complex.
Subject of Research: Preparation and characterization of paracetamol-β-cyclodextrin inclusion complex and its antipyretic activity in rats
Article Title: Preparation, characterization, and antipyretic activity of paracetamol and β-cyclodextrin inclusion complex
Article References: Lescano, J. I., Sani, J., Tabornal, M. A., Arce, F. J., See, G. L., Kodama, N., & Inoue, Y. (2026). Preparation, characterization, and antipyretic activity of paracetamol and β-cyclodextrin inclusion complex. AAPS Open, 12(1), Article 33. https://doi.org/10.1186/s41120-026-00170-6
Image Credits: AI Generated
DOI: 10.1186/s41120-026-00170-6
Keywords: paracetamol, β-cyclodextrin, inclusion complex, antipyretic activity, orodispersible films, drug delivery, crystalline habit, dissolution profile, pharmaceutical formulation, rat model, Preparation, characterization
News Source: Ophelia Keating. (November 13, 2026). Paracetamol-Cyclodextrin Complex Shows Regulated Release and Limited Antipyretic Effect. Scienmag.
