Exploring the use of modified in vitro digestion assays for the evaluation of ritonavir loaded solid lipid-based formulations.

Solid lipid-based formulations (sLBFs) have the potential to increase the oral bioavailability of drugs with poor solubility in water, while counteracting some of the disadvantages of liquid LBFs. The most common experimental set-up to study the performance of LBFs in vitro is the lipolysis assay, during which the LBFs are digested by lipases in an environment mimicking the human small intestine. However, this assay has failed in many cases to correctly predict the performance of LBFs in vivo, highlighting the need for new and improved in vitro assays to evaluate LBFs at the preclinical stage. In this study, the suitability of three different in vitro digestion assays for the evaluation of sLBFs was assessed; the classic one-step intestinal digestion assay, a two-step gastrointestinal digestion assay and a bicompartmental assay permitting the simultaneous monitoring of digestion and permeation of the active pharmaceutical ingredient (API) across an artificial membrane (Lecithin in Dodecane – LiDo). Three sLBFs (M1-M3) with varied composition and ritonavir as model drug were prepared and examined. When comparing the ability of these formulations to keep the drug solubilized in the aqueous phase, all three assays show that M1 performs better, while M3 presents poor performance. However, the classic in vitro intestinal digestion assay fails to provide a clear ranking of the three formulations, something that is more evident when using the two modified and more physiologically relevant assays. Also, the two modified assays provide additional information about the performance of the formulations including the performance in the gastric environment and intestinal flux of the drug. These modified in vitro digestion assays are valuable tools for the development and evaluation of sLBFs to make better informed decisions of which formulations to pursue for in vivo studies.

2.1. Materials

Acetonitrile (ACN, ≥ 99.9%), dimethyl sulfoxide (DMSO, ≥ 99.9%), methanol (≥ 99.9%), pancreatin from porcine pancreas (8 x USP specifications), lipase from Rhizopus oryzae (≥ 30 U/mg), 4-bromophenylboronic acid (4-BBBA, ≥ 95.0%), D-α-tocopherol polyethylene glycol 1000 succinate (TPGS), Trizma® maleate (TRIS maleate salt), sodium hydroxide (NaOH) pellets, sodium chloride (NaCl), potassium chloride (KCl), calcium chloride (CaCl₂) granules, sodium phosphate monobasic dihydrate (NaH₂PO₄-2H₂O) and potassium phosphate monobasic (KH₂PO₄) were purchased from Merck (Darmstadt, Germany). WITEPSOL® E 85 pastilles, IMWITOR® 491 powder and DYNASAN® 116 flakes were kindly provided from IOI Oleo GmbH (Hamburg, Germany). Gelucire® 48/16 and Gelucire® 50/13 pellets were kindly donated by Gattefossé (Saint-Priest, France). Ethanol (99.5%, denatured with 100 g isopropyl alcohol) and n-dodecane (≥ 99%) were purchased from Solveco (Rosersberg, Sweden) and Alfa Aesar (Lancashire, UK), respectively. Lecithin (L-α-phosphatidylcholine) Soy PC extract (20%) was obtained from Avanti Polar Lipids (Alabaster, AL, USA). Ritonavir (> 99%) was purchased from Shanghai Desano Pharmaceuticals Co., Ltd. (Shanghai, China) and Lucifer yellow (LY) CH dilithium salt was purchased from Biotium (Fremont, CA, USA). FaSSIF/FeSSIF/FaSSGF powder was obtained from biorelevant.com (London, UK). Ultrapure Milli-Q® water (grade I) from a direct water purification system (Merck, Darmstadt, Germany) was used for all the experiments.

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Ioannis I. Andreadis, Arne Schulzen, Julian Quodbach, Christel A.S. Bergström, Exploring the use of modified in vitro digestion assays for the evaluation of ritonavir loaded solid lipid-based formulations., European Journal of Pharmaceutical Sciences, 2023, 106524, ISSN 0928-0987,
https://doi.org/10.1016/j.ejps.2023.106524.