Abstract
This study optimized the formulation parameters of nanostructured lipid carriers (NLCs) loaded with Origanum vulgare L. essential oil (OEO), produced by hot emulsification followed by sonication. Phase separation with heating at 60–70 °C for 10min yielded nanoparticles with a mean diameter of 102.0 ± 3.8 nm, polydispersity index (PdI) of 0.16 ± 0.01, and zeta potential of −26.43 ± 0.09 mV. Sonication power (60–80%) and processing time (≤ 5 min/mL) were critical to maintaining particle size below 130 nm and PdI below 0.225; powers exceeding 80% induced particle re-aggregation. Laboratory-scale volume increased from 1 to 10 mL, and comparisons between different sonication probe types required fine-tuning of processing parameters. Fourier-transform infrared spectroscopy (FTIR) suggested successful incorporation of OEO into the lipid matrix, with no significant band shifts indicating the absence of strong covalent or ionic interactions between the oil and the lipid components, with an encapsulation efficiency of 99%. DSC/DTG analyses demonstrated thermal stability up to 200 °C and the preservation of a crystalline lipid matrix. Storage under refrigeration (4–7 °C) maintained the physicochemical integrity of the formulations over 60 days, whereas ambient temperature promoted aggregation. Interaction with collagen produced moderate shifts in PdI and zeta potential without significant aggregation. Overall, the formulation demonstrated favorable physicochemical characteristics that justify future biological evaluation.
1. Introduction
Natural products have historically been pillars in the discovery and development of pharmaceuticals, offering a vast array of bioactive compounds with diverse and promising pharmacological properties. (Gwiazdowska et al., 2024) This successful trajectory provides a solid foundation for the ongoing exploration of natural sources in the development of new therapies. However, the direct therapeutic application of natural products, especially essential oils such as Origanum vulgare L. (oregano) (OEO), is severely limited by intrinsic challenges, such as high volatility, instability under exposure to light, heat, and oxygen, and low solubility in aqueous media. (Miranda-Arizmendi et al., 2024a; H. M. Pires et al., 2024) Although essential oils such as OEO are susceptible to volatilization and oxidative degradation under specific storage and processing conditions, their thermal stability is context-dependent: recent evidence demonstrates that the major phenolic constituents (carvacrol and thymol) retain significant antioxidant efficacy even under high-temperature conditions, including industrial frying processes (López et al., 2024, 2023). Lipid nanoencapsulation is therefore targeted not solely at thermal protection, but primarily at improving aqueous dispersibility, controlled localization, and versatility of application. These characteristics compromise their stability, efficacy, and bioavailability, leading to rapid degradation of active compounds, reduced shelf life, and difficulties in formulation for biological systems. To overcome these technological barriers and maximize the biological potential of essential oils, the development of controlled delivery systems, particularly nanosystems, has been intensively investigated. (H. M. Pires et al., 2024)
In this context, Nanostructured Lipid Carriers (NLCs) have emerged as a highly promising strategy for the incorporation of essential oils. (Silva et al., 2025) These advanced colloidal systems, second-generation lipid nanoparticles composed of a mixture of solid and liquid lipids in their core, offer significant advantages, including enhanced stability of the active compound, protection against environmental degradation, and precise control over release profiles (Chura et al., 2023; Silva et al., 2025). The less crystalline matrix of NLCs, compared to other solid lipid nanoparticles (SLNs), allows for higher encapsulation capacity, with encapsulation efficiencies ranging from 85% to 95%. Moreover, NLCs demonstrate high biocompatibility and low cytotoxicity, positioning these nanosystems as a promising and versatile platform for pharmaceutical applications. (Viegas et al., 2023)
Despite the growing scientific interest in the incorporation of essential oils into NLCs, the literature still lacks robust methodological systematization and standardization. Available studies frequently diverge regarding the formulation variables employed, such as the composition and ratio of solid and liquid lipids, the type and concentration of surfactants, and the parameters of the production process, which hinders the comparison of results and the identification of optimal formulation conditions (Chura et al., 2023; Silva et al., 2025). Specifically, for Origanum vulgare L. essential oil, although its biological properties, including antimicrobial, antioxidant, and anti-inflammatory activities, are widely documented (Miranda-Arizmendi et al., 2024a), its incorporation into NLCs remains insufficiently explored from a technological and food perspective. The scarcity of studies that systematically and integrally evaluate the critical formulation and process parameters, correlating them with the final physicochemical properties of the nanoparticles, represents a significant gap that limits the advancement of this strategy towards real applications.
In this context, the present work distinguishes itself from the existing literature by adopting a systematic and multifactorial experimental approach for the optimization of OEO-loaded NLCs, integrally investigating the influence of formulation and processing variables on the physicochemical characteristics of the nanoparticles, including particle size, polydispersity index (PdI), zeta potential, and encapsulation efficiency. By providing a clearer mechanistic understanding of the relationships between formulation parameters and nanosystem performance, this study directly contributes to bridging the gaps identified in the literature and establishes a solid scientific foundation for the rational development of nanotechnological formulations based on essential oils. The findings obtained have the potential to positively impact the field of nanotechnology, paving the way for a more efficient and safe biological use of OEO, while also serving as a replicable methodological framework for other essential oils of clinical relevance.
Therefore, this research highlights that the essential oil of Origanum vulgare L. is widely recognized as a source of high-value bioactive compounds, most notably carvacrol and thymol, who’s antimicrobial and antioxidant properties have been extensively explored for food preservation, natural additive development, active packaging systems, and crop protection strategies. (Miranda-Arizmendi et al., 2024b) The technological limitations that constrain OEO’s direct application in food and agricultural matrices, namely its volatility, susceptibility to oxidative degradation, and poor aqueous dispersibility, are precisely the challenges that lipid-based nanoencapsulation is positioned to resolve. By establishing a systematic and reproducible framework for the production of physiochemically stable OEO-loaded NLCs, this study provides the molecular and materials foundation for rationally designing food-grade and agrochemical formulations of enhanced efficacy. In this way, the present study was designed to systematically investigate the key formulation and processing parameters influencing the physicochemical properties of Origanum vulgare L. essential oil (OEO)–loaded NLCs, to optimize their performance, stability, and overall pharmaceutical applicability. Furthermore, this research aims to develop a robust and efficient delivery system capable of enhancing the technological properties of the essential oil, such as stability and solubility, thereby justifying future biological evaluation.
2. Experimental section
Precirol® ATO 5 was obtained from Gattefossé (Paramus, NJ, USA). Origanum vulgare L. essential oil (OEO) was supplied by Ferquima Indústria e Comércio (São Paulo, Brazil). Tween® 80, Span® 60, collagen, and phosphate-buffered saline (PBS) pH 7.0 were purchased from Sigma-Aldrich® (St. Louis, MO, USA). All materials were used as received.
2.1. NLCs production
The lipid carriers were produced using the methodology of Uchôa et al. (2025) with some modifications. Thus, the formulation components were weighed in two different phases, where the oily phase contained Precirol® ATO 5 (1%), Span® 60 (1%), OEO (2%), and Tween® 80 (3%), and the aqueous phase was formed by ultra-purified water final volume of 1 mL. Subsequently, both phases were heated in a water bath at 60 °C (±5 °C) for 180 s under constant stirring at 1100 rpm (SPLabor, n° SP-10,206/A, São Paulo, Brazil). Then, the aqueous phase was poured over the oily phase under continuous stirring at 1100 rpm for 8 min. The formulation was subjected to an ultrasonicator (Ultronique 550 w, 20 kHz, Indaiatuba, Brazil) at 60% power for 90 s. Subsequently, the colloidal dispersions (OEO
NLCs) were kept at room temperature for 5 min for complete system formation and then characterized for their physicochemical parameters: hydrodynamic particle size (nm), PdI, and Zeta potential (ζ) (mV).
NLCs) were kept at room temperature for 5 min for complete system formation and then characterized for their physicochemical parameters: hydrodynamic particle size (nm), PdI, and Zeta potential (ζ) (mV).Read the full article here.
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Source:
Pedro H.O. Souza, Ana F.C. Uchôa, Allessya L.D. Formiga, Lucas M.M. Carvalho, Graziela M.A. Pereira, Luís F.A. Silva, Marcele F. Passos, José M. Barbosa-Filho, Marcelo S. Silva, Francisco H. Xavier-Júnior,
Process optimization and stability enhancement of Origanum vulgare L. essential oil-loaded nanostructured lipid carriers,
Applied Food Research, Volume 6, Issue 2, 2026, 102339, ISSN 2772-5022, https://doi.org/10.1016/j.afres.2026.102339.
