Bioactive Delivery Research Articles

Preparation of β-lactoglobulin/gum arabic complex nanoparticles for encapsulation and controlled release of EGCG in simulated gastrointestinal digestion model

In this work, the β-lactoglobulin/gum arabic (β-Lg-GA) complexes were prepared to encapsulate epigallocatechin gallate (EGCG), forming β-Lg-GA-EGCG complex nanoparticles with an average particle size of 133 nm. The β-Lg-GA complexes exhibited excellent encapsulation efficiency (84.5%), and the antioxidant performance of EGCG in vitro was improved after encapsulation. It was recorded that 86% of EGCG could be released in simulated intestinal fluid after 3 h of digestion, much faster than that in simulated gastric fluid, indicating that the β-Lg-GA complexes were effective in enhancing EGCG stability, which was confirmed using SDS-PAGE and SEM. Further spectrum results demonstrated that various intramolecular interactions including electrostatic, hydrophobic and hydrogen bonding interactions contribute to the formation of β-Lg-GA-EGCG complex nanoparticles. Also, XRDexperiments indicated that EGCG was successfully encapsulated by β-Lg-GA complexes. Therefore, the β-Lg-GA complexes hold great potentials in the protective delivery of sensitive bioactives.

Advancements in liposome technology: Preparation techniques and applications in food, functional foods, and bioactive delivery: A review.

Liposomes play a significant role in encapsulation of various bioactive compounds (BACs), including functional food ingredients to improve the stability of core. This technology can be used for promoting an effective application in functional food and nutraceuticals. Incorporation of traditional and emerging methods for the developments of liposome for loading BACs resulted in viable and stable liposome formulations for industrial applications. Thus, the advance technologies such as supercritical fluidic methods, microfluidization, ultrasonication with traditional methods are revisited. Liposomes loaded with plant and animal BACs have been introduced for functional food and nutraceutical applications. In general, application of liposome systems improves stability, delivery, and bioavailability of BACs in functional food systems and nutraceuticals. This review covers the current techniques and methodologies developed and practiced in liposomal preparation and application in functional foods.

How alginate properties influence in situ internal gelation in crosslinked alginate microcapsules (CLAMs) formed by spray drying.

Alginates gel rapidly under ambient conditions and have widely documented potential to form protective matrices for sensitive bioactive cargo. Most commonly, alginate gelation occurs via calcium mediated electrostatic crosslinks between the linear polyuronic acid polymers. A recent breakthrough to form crosslinked alginate microcapsules (CLAMs) by in situ gelation during spray drying ("CLAMs process") has demonstrated applications in protection and controlled delivery of bioactives in food, cosmetics, and agriculture. The extent of crosslinking of alginates in CLAMs impacts the effectiveness of its barrier properties. For example, higher crosslinking extents can improve oxidative stability and limit diffusion of the encapsulated cargo. Crosslinking in CLAMs can be controlled by varying the calcium to alginate ratio; however, the choice of alginates used in the process also influences the ultimate extent of crosslinking. To understand how to select alginates to target crosslinking in CLAMs, we examined the roles of alginate molecular properties. A surprise finding was the formation of alginic acid gelling in the CLAMs that is a consequence of simultaneous and rapid pH reduction and moisture removal that occurs during spray drying. Thus, spray dried CLAMs gelation is due to calcium crosslinking and alginic acid formation, and unlike external gelation methods, is insensitive to the molecular composition of the alginates. The 'extent of gelation' of spray dried CLAMs is influenced by the molecular weights of the alginates at saturating calcium concentrations. Alginate viscosity correlates with molecular weight; thus, viscosity is a convenient criterion for selecting commercial alginates to target gelation extent in CLAMs.

Exploiting the robust network structure of zein/low-acyl gellan gum nanocomplexes to create Pickering emulsion gels with favorable properties

Zein/low-acyl gellan gum (GG) composite particles (ZGPs) were fabricated to stabilize Pickering emulsions (termed “ZGPEs”). The wettability of ZGPs was manipulated simply by adjusting the concentration of GG. The effects of GG concentration, oil fraction and pH on ZGPEs were systematically evaluated by confocal laser scanning microscopy (CLSM), cryo-scanning electron microscopy (cryo-SEM), dynamic light scattering technique, stimulated emission depletion (STED) nanoscopy and rheology. The results showed that ZGPEs exhibited robust colloidal properties and distinct advantage over other previously reported zein-polysaccharide-based Pickering emulsions. CLSM, STED and cryo-SEM analyses revealed that the network structures formed by GG and ZGPs at the continuous phase and oil–water interface were the main contributors to the emulsion’s characteristics. This study provides insights into the fabrication of food-grade Pickering emulsions with distinct characteristics that impart favorable properties to various foods and bioactive delivery systems.

Soy Protein Pressed Gels: Gelation Mechanism Affects the In Vitro Proteolysis and Bioaccessibility of Added Phenolic Acids.

In this study, a model system of firm tofu (pressed gel) was prepared to study how the coagulation mechanism—acidification with glucono δ-lactone (GDL) or coagulation with magnesium sulphate (MgSO4)—affected the physical properties of the gels along with their in vitro proteolysis (or extent of proteolysis). The two types of gels were also fortified with 3.5 mM protocatechuic (PCA) and coumaric acid (CMA) to test whether they can be used as bioactive delivery systems. Texture analysis showed that all MgSO4-induced gels (fortified and control) had a higher hydration capacity and a weaker texture than the GDL-induced gels (p < 0.05). MgSO4 gels had almost double proteolysis percentages throughout the in vitro digestion and showed a significantly higher amino acid bioaccessibility than the GDL gels (essential amino acid bioaccessibility of 56% versus 31%; p < 0.05). Lastly, both gel matrices showed a similar phenolic acid release profile, on a percentage basis (80% for PCA and 100% for CMA). However, GDL gels delivered significantly higher masses of bioactives under simulated intestinal conditions because they could retain more of the bioactives in the gel after pressing. It was concluded that the coagulation mechanism affects both the macro- and microstructure of the soy protein pressed gels and as a result their protein digestibility. Both pressed gel matrices are promising delivery systems for bioactive phenolic acids.

Mechanisms of absorption of vitamin D3 delivered in protein nanoparticles in the absence and presence of fat.

Novel protein-based nanovehicles offer alternatives to fat for delivery of lipophilic bioactives (nutraceuticals and drugs), yet they raise important questions regarding the bioavailability and absorption mechanism of the bioactive without fat. To provide answers, we chose vitamin D3 (VD3) as a model lipophilic-nutraceutical, re-assembled casein-micelles (rCM) as model protein-based nanovehicles, and non-fat yoghurt as a model food. We prepared three yoghurt formulations: 3% fat with VD3 dissolved in milk-fat, non-fat and 3% fat, both latter enriched with VD3 within rCM. Following in vitro digestion, VD3 retention and bioaccessibility were high (∼90% and ∼70%, respectively) in all formulations. VD3 uptake by Caco-2 cells was three-fold higher (p < 0.005) in the non-fat yoghurt enriched with VD3 in rCM compared with enriched fat-containing yoghurts. SR-BI, CD36 and NPC1L1 transporters were involved in VD3 absorption irrespective of the composition. Thus, our findings demonstrate that protein nanovehicles may improve VD3 bioavailability, without altering its absorption mechanism compared to that from fat.

In-vitro stress stability, digestibility and bioaccessibility of curcumin-loaded polymeric nanocapsules

Nanoemulsion-based delivery systems have a considerable potential for the encapsulation, protection and delivery of lipophilic bioactives. In the current study, in-vitro stability of curcumin-loaded single (NEI), double (NEII), and triple-layered nanostructures was investigated against varying environmental stress (temperature, pH, ionic strength) and storage conditions. In addition, their functional performance, as influenced by the number of polymer layers, was also assessed by comparing in-vitro lipid digestibility and curcumin bioaccessibility. Initially, NEI was prepared through ultrasonic homogenisation technique, and stabilised with modified starch. Next, sequential deposition of chitosan and carboxymethyl cellulose (CMC) on the NEI droplets resulted into the formation of NEII and nanocapsules, respectively. Nanocapsules showed good stability against aggregation over a wide range of pH (3.0–7.0), salt concentration (50–300 mM NaCl) and temperature (30–90 °C for 30 min). Digestion studies suggested that polymeric coatings interfered with lipolysis as the digestibility (% free fatty acid released) of oil droplets was considerably reduced with increasing the number of layers surrounding the oil droplets. However, curcumin bioaccessibility increased with adding the number of coating layers. Overall, results indicate that the developed nano-system could be helpful in improving the oral delivery of curcumin besides incorporating it into functional foods and beverages.

Transethosomes: Novel Technology for Skin Delivery of Drugs

The human body’s contact with the outside world is the skin, which has a total surface area of roughly 1.8 m2. Because it removes many of the difficulties associated with the oral route, the transdermal mode of medicine administration has aroused a lot of interest in pharmaceutical research. Although the skin, particularly the stratum corneum, is a barrier to most medicine absorption, it does provide a large (1.2 m2) and easily accessible surface area for drug diffusion. Several approaches for improving bioactive delivery through transdermal distribution have recently been investigated. Some of the most frequent examples are iontophoresis, electrophoresis, sonophoresis, chemical permeation enhancers, magnetophoresis, microneedles, and vesicular systems (niosomes, liposomes, elastic liposomes such as transfersomes, ethosomes, and transethosomes). Transethosomes seem to be the most promising of the group, since they include both lipophilic and hydrophilic sections and can take therapeutic compounds with a wide range of solubility. Transethosomes may bend and pass through constrictions that are 5 to 10 times smaller in diameter than their own. Intact vesicles may be pierced more readily due to their high deformability. These vesicles may transport analgesics, anesthetics, corticosteroids, sex hormones, anticancer drugs, insulin, and other pharmaceuticals transdermally.

Study on the possibility of developing food-grade hydrophobic bio-aerogels by using an oleogel template approach

The feasibility of producing food-grade hydrophobic bio-aerogels by supercritical-carbon dioxide (SC–CO2) extraction of oil from oleogels was investigated for the first time. Medium chain triglycerides (MCT) oil was gelled using ethylcellulose (EC) at increasing concentration (10, 15, 20% w/w) and grade (EC20, 45, 100), eventually in combination with fillers. Different SC-CO2 oil extraction procedures were tested. The acquired results show that both oleogel formulation and extraction conditions can steer the EC scaffold structure. The increase in EC concentration and grade resulted in oleogels more structurally stable to SC-CO2 extraction. The application of a pulsed extraction procedure allowed obtaining a low-density (0.39 ​g/cm3) EC scaffold presenting 60% oil. Addition of freeze dried lettuce powder improved macrostructure homogeneity. The obtained results lay the foundations for developing food-grade hydrophobic bio-aerogels, which are expected to present unique oil absorption and bioactive delivery features.

Nano-Biomimetic Drug Delivery Vehicles: Potential Approaches for COVID-19 Treatment.

The current COVID-19 pandemic has tested the resolve of the global community with more than 35 million infections worldwide and numbers increasing with no cure or vaccine available to date. Nanomedicines have an advantage of providing enhanced permeability and retention and have been extensively studied as targeted drug delivery strategies for the treatment of different disease. The role of monocytes, erythrocytes, thrombocytes, and macrophages in diseases, including infectious and inflammatory diseases, cancer, and atherosclerosis, are better understood and have resulted in improved strategies for targeting and in some instances mimicking these cell types to improve therapeutic outcomes. Consequently, these primary cell types can be exploited for the purposes of serving as a “Trojan horse” for targeted delivery to identified organs and sites of inflammation. State of the art and potential utilization of nanocarriers such as nanospheres/nanocapsules, nanocrystals, liposomes, solid lipid nanoparticles/nano-structured lipid carriers, dendrimers, and nanosponges for biomimicry and/or targeted delivery of bioactives to cells are reported herein and their potential use in the treatment of COVID-19 infections discussed. Physicochemical properties, viz., hydrophilicity, particle shape, surface charge, composition, concentration, the use of different target-specific ligands on the surface of carriers, and the impact on carrier efficacy and specificity are also discussed.

Targeted Delivery of Natural Bioactives and Lipid-nanocargos against Signaling Pathways Involved in Skin Cancer.

At present, skin cancer is considered a widespread malignancy in human beings. Among diverse population types, Caucasian populations are much more prone to this malignancy in comparison to darker skin populations due to the lack of skin pigmentation. Skin cancer is divided into malignant and non-melanoma skin cancer, which is further categorized as basal and squamous cell carcinoma. Exposure to ultraviolet radiation, chemical carcinogen (polycyclic aromatic hydrocarbons, arsenic, tar, etc.), and viruses (herpes virus, human papillomavirus, and human T-cell leukemia virus type-1) are major contributing factors to skin cancer. There are distinct pathways available through which skin cancer develops, such as the JAK-STAT pathway, Akt pathway, MAPKs signaling pathway, Wnt signaling pathway, to name a few. Currently, several targeted treatments are available, such as monoclonal antibodies, which have dramatically changed the line of treatment of this disease but possess major therapeutic limitations. Thus, many phytochemicals have been evaluated either alone or in combination with the existing synthetic drugs to overcome their limitations and have been found to play a promising role in the prevention and treatment. In this review, a complete overview of skin cancer, starting from the signaling pathways involved, newer developed drugs with their targets and limitations, along with the emerging role of natural products alone or in combination as potent anticancer agents and their molecular mechanism involved has been discussed. Apart from this, various nano-cargos have also been mentioned here, which can play a significant role in the management and treatment of different types of skin cancer.

Xylo-oligosaccharide microparticles with synbiotic potential obtained from enzymatic hydrolysis of sugarcane straw

Synbiotic formulations and microencapsulation techniques have been explored in food industries to guarantee the viability of probiotic organisms; playing an important role in microbiota balance. Microparticles of alginate, gelatin and xylo-oligosaccharides (XOS) were produced by external gelation with the purpose of enhancing the survival rate of the probiotic L. acidophilus. XOS was obtained through enzymatic hydrolysis of xylan extracted from sugarcane straw, achieving more than 70% conversion and used for microparticle preparation. Microparticles containing 3% XOS provided greater cell protection during exposure to the gastrointestinal tract and during refrigerated storage; keeping 97.86 ± 0.44% of viability during 28 days of storage and enabling 87.50 ± 0.02% survival after digestive simulation. However, particles without XOS showed 84.49 ± 0.59% of viability after storage and 68.45 ± 0.03% after digestion assay. These results lead to promising applications in synbiotic and functional food formulations comprised of components requiring extended shelf-life, protection from gastrointestinal conditions and gradual bioactive delivery.

Evaluating the structural properties of bioactive-loaded nanocarriers with modern analytical tools.

With the emergence of nanocarriers for offering smart transformers improving the bioavailability and functionality of food bioactive compounds, a critical issue is experimental and analytical evaluation of the bioactive-loaded nanocarriers; so, further technical advancements are necessary for the precise structural characterization of these nanodelivery systems and application of sophisticated analytical techniques. Here, we have portrayed the current progress in the structural characterization approaches including X-ray diffraction, differential scanning calorimetry, thermogravimetric analysis, fluorescence spectroscopy, circular dichroism, Fourier-transform infrared spectroscopy, nuclear magnetic resonance, X-ray photoelectron spectroscopy, and small-angle X-ray scattering/small-angle neutron scattering as powerful and informative tools to have a better control on the design, development, and appraisal of nanoengineered bioactive delivery systems. Different examples and case studies have been tabulated along with illustrative and schematic representation of relevant results. An emerging view concerning the challenges for future structural characterization of these systems is also discussed in detail.

Development of a genetic framework to improve the efficiency of bioactive delivery from blueberry

In the present study, we applied a novel high-throughput in vitro gastrointestinal digestion model to phenotype bioaccessibility of phenolics in a diverse germplasm collection representing cultivated highbush blueberries. Results revealed significant (P < 0.05) differences between accessions, years, and accession by year interaction for relative and absolute bioaccessibility of flavonoids and phenolic acids. Broad sense heritability estimates revealed low to moderate inheritances of relative and absolute bioaccessibility, suggesting that besides environmental variables, genetics factors could control bioaccessibility of phenolics. Acylated anthocyanins had significantly higher relative bioaccessibility than non-acylated anthocyanins. Correlation analysis indicated that relative bioaccessibility did not show significant association with fruit quality or raw concentration of metabolites. The study also identified accessions that have high relative and absolute bioaccessibility values. Overall, combining the bioaccessibility of phenolics with genetic and genomic approaches will enable the identification of genotypes and genetic factors influencing these traits in blueberry.

Intervertebral Disc Regeneration Using Stem Cell/Growth Factor-Loaded Porous Particles with a Leaf-Stacked Structure.

Although biological therapies based on growth factors and transplanted cells have demonstrated some positive outcomes for intervertebral disc (IVD) regeneration, repeated injection of growth factors and cell leakage from the injection site remain considerable challenges for human therapeutic use. Herein, we prepare human bone marrow-derived mesenchymal stem cells (hBMSCs) and transforming growth factor-β3 (TGF-β3)-loaded porous particles with a unique leaf-stack structural morphology (LSS particles) as a combination bioactive delivery matrix for degenerated IVD. The LSS particles are fabricated with clinically acceptable biomaterials (polycaprolactone and tetraglycol) and procedures (simple heating and cooling). The LSS particles allow sustained release of TGF-β3 for 18 days and stable cell adhesiveness without additional modifications of the particles. On the basis of in vitro and in vivo studies, it was observed that the hBMSCs/TGF-β3-loaded LSS particles can provide a suitable milieu for chondrogenic differentiation of hBMSCs and effectively induce IVD regeneration in a beagle dog model. Thus, therapeutically loaded LSS particles offer the promise of an effective bioactive delivery system for regeneration of various tissues including IVD.

Physicochemical and biological properties of nanochitin—abamectin conjugate for Noctuidae insect pest control

Abamectin (Am) is an excellent biopesticide for insect pests control but environmental sensitivity and a short duration in conventional formulations limit its application in agriculture. In order to promote the dispersity, stability, and biological activity of Am, well-monodisperse nanochitin (NCh) and Am-loaded nanoparticles (NCh-Am) were prepared and examined in this study. NCh was prepared by ultrasonication combining with low concentration acidic hydrolysis and NCh-Am nanoparticles were combined by conjugation in dispersion naturally. The effective particle size of NCh was 110.6 ± 18.6 nm with cationic nature, while those of NCh-Am conjugates ranged from 160 to 700 nm based on the mixing ratio. The binding efficiency of Am to NCh in the conjugates reached 53.7 ± 7.6% in DI water at pH 6.2. The thermal stability and water solubility of Am were greatly promoted by NCh. The photodegradation of Am in the conjugates decreased 48% when exposed to UV light for 56 h. The conjugate containing half of Am showed significant control efficiency in larva growth and enhanced mortality against 2 instar larvae of noctuid. The results suggested that NCh significantly promoted Am water dispersity, thermal stability, and photostability, and effectively reduced pesticides dosage in insect pest control. This finding indicates that NCh-Am can be used as a new bioactive delivery system in future pest management.

Abstract 2868: A novel fusogenic peptide for delivery of bioactive siRNAs targeting CSNK2A1 in ovarian cancer cells in vitro

Abstract Introduction: Ovarian cancer recurrence and survival rates have remained stagnant over time with current therapeutics displaying low efficacy and often rendered ineffective by degradation. This work presents novel fusogenic peptides that will electrostatically complex with siRNAs and cause a pH-responsive insertion into the endosomal membrane, resulting in destabilization and breakdown allowing for release of siRNAs into the cytoplasm of ovarian cancer cells. Delivery of bioactive siRNAs targeting the CSNK2A1 gene in ovarian cancer cells is hypothesized to reduce cell viability, migration, and invasion. Experimental Procedures: Peptide formulations were complexed with non-targeting siRNAs (siNT) and CSNK2A1 siRNAs (siCSNK2A1) at various N:P ratios for all procedures. Analysis of complex formation was completed through gel-shift assays and dynamic light scattering. After characterization of the complexes, each formulation without siRNA was delivered to OVCAR-3 and CAOV-3 cells for cytotoxicity analysis measured via MTS assay. Observation of cellular uptake and endosomal escape was completed through fluorescent imaging after delivering each peptide formulation complexed with Cy3-labeled siNT (Cy3-siNT) and performing early endosome antigen-1 staining. To confirm bioactivity of targeted siRNAs, complexes with siCSNK2A1 were delivered and gene silencing was analyzed via western blotting. Results: Complexation of siRNAs with each formulation was achieved at N:P ratios of 40:1 and greater. Each of the three peptide formulations were able to form uniform nanocomplexes, at 195 nm, 111 nm, and 59 nm with 8.25 mV, -14.56 mV, and -18.42 mV surface charges respectively, and did not display vehicle cytotoxicity at an 80:1 ratio. Each peptide also mediated siRNA uptake into ovarian cancer cells, with fluorescent siRNA signal increasing with increasing N:P ratios. Of note, we observed that Cy3-siRNA appeared separate from the endosome, indicative of endosomal escape, which was seen in several peptide formulations with both CAOV-3 and OVCAR-3 cell lines. Confirmation of bioactive delivery of siCSNK2A1 is shown through western blot imaging, demonstrating knockdown of CSNK2A1 in comparison to cells treated with siNT alone or left untreated. Conclusion: Our novel fusogenic peptides have shown the ability to complex with and deliver siRNAs into ovarian cancer cells. Each peptide mediates endosomal escape activity, with varying degrees of effectiveness, and silencing of CSNK2A1 as measured by western blot. Further study into downstream effects of CSNK2A1 is ongoing. Future work includes optimization of a single fusogenic sequence for optimal delivery of siRNAs and development of a novel targeting peptide to increase cancer targeting and uptake efficiency. Acknowledgements: This research was supported in part by the National Science Foundation EPSCoR Program under NSF Award # OIA-1655740. Citation Format: Timothy Samec, Jessica Boulos, Serena Gilmore, Angela Alexander-Bryant. A novel fusogenic peptide for delivery of bioactive siRNAs targeting CSNK2A1 in ovarian cancer cells in vitro [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2868.

Cannabis extract nanoemulsions produced by high-intensity ultrasound: Formulation development and scale-up

Over the past several decades, it has been demonstrated that cannabinoids offer a wide range of therapeutic benefits. Their oral administration, however, while arguably the most convenient and discrete, has been associated with low bioavailability, delayed onset of action and poor reproducibility resulting from the extracts' strongly lipophilic character. To overcome these obstacles, cannabinoids can be incorporated into oil-in-water nanoemulsions: a process known to enhance the delivery of lipophilic bio-actives by making them behave like water-soluble (hydrophilic) compounds. In this manuscript, formulation development and production scale-up procedures for a cannabis extract (CBDX, 55% cannabidiol)-containing nanoemulsion are described. Nanoemulsion samples were prepared by high-intensity ultrasonic liquid processing, and the formulation was optimized for carrier oil and surfactant(s) contents as well as for the hydrophilic-lipophilic balance (HLB) of the surfactant mixture. Translucent CBDX-containing nanoemulsions with median droplet sizes well below 100 nm were possible to form with synthetic surfactants (Tween 80/Span 80 mixture), but not with a natural surfactant (Q-naturale®). By utilizing Barbell Horn® Ultrasonic Technology (BHUT), the nano-emulsification process was successfully scaled up, achieving a commercial-level processing rate equivalent to one million nanoemulsified 10 mg CBDX doses made per month with a single bench-scale ultrasonic liquid processor (BSP-1200).

Nanotechnology for bioactives delivery systems

The properties of functional foods depend largely on their micro- and/or nano-scale structure. Conventionally, the preparation of functional foods is based on large-scale processing. As a consequence, the structure of the products cannot be precisely controlled. Recently, micro-/nano-technology have been developing and increasingly applied to the formulation of functional foods. This paper summarizes some applications of micro-/nano-processing of food materials into dispersions, foreseeing their application as bioactives delivery systems. Nanodispersions containing antioxidant food materials were formulated and evaluated using various top-down techniques or a bottom-up method, particularly the performance of various polyglycerol esters (PGE) of fatty acid in the preparation and characterization of β-carotene nanodispersions during in vitro gastric digestion. MicroChannel (MC) and nanochannel (NC) emulsification for producing monodisperse emulsions were investigated. The development of NC emulsification devices for monodisperse fine emulsions is also presented.

Impact of liquid lipid on development and stability of trimyristin nanostructured lipid carriers for oral delivery of resveratrol

Nanostructured lipid carriers (NLCs) have emerged as versatile carriers to improve oral bioavailability of poorly water-soluble drugs as well as to protect labile drugs from degradation and metabolism. Prepared by blending solid and liquid lipids, the choice of liquid lipid can have a great influence on their physicochemical characteristics and stability. The present work investigated the impact of six different liquid lipids with diverse chemical structures and hydrophilic and lipophilic balance (HLBs) on the critical quality attributes (CQAs) and storage stability of NLCs with trimyristin as solid lipid. Resveratrol (RES) was used as model drug as its low water solubility, poor bioavailability, rapid metabolism and clearance from systemic circulation restricts its clinical use despite its wide spectrum of biological activities. Liquid lipids investigated included, two triglycerides, one medium chain (C8) glycerol tricaprylate (GTC) and second, long chain (C18) glyceryl trioleate (GTO); two propylene glycol fatty acid esters, propylene glycol monocaprylate (PGMC) and propylene glycol monolaurate (PGML); fatty acid ester decyl oleate (DO) and a PEGylated lipid polyethylene glycol-8 caprylic/capric glycerides (PCG). Box–Behnken experimental design was employed to ascertain the effect of four independent factors viz. type of liquid lipid, amount of liquid lipid, amount of drug and surfactant concentration and interactions between these factors on the CQAs of NLCs as response variables viz. particle size (PS), polydispersity index (PDI), zeta potential (ZP), drug encapsulation efficiency (EE), and drug loading (DL). The relationship between various factors and responses was established by response surface methodology (RSM). The oils with higher lipophilicity C18 triglycerides (GTO) and C18 fatty acid ester DO yielded NLCs with lower PS as compared to the oils with lower lipophilicity (PGC, PGMC and PGML). Although increasing the concentration of liquid lipids had an upward trend on the PS of NLCs, its PDI was more predominantly influenced by the nature of liquid lipid. The characteristic of the liquid lipid influenced the DL remarkably which varied from 2.94 to 7.56%. The ZP of nanoparticles varied from −21.3 to −39.9 mV with liquid lipids with free hydroxyl groups and higher HLB playing a more prominent role contributing to the increase in the negative surface charge. The characteristics of liquid lipid influenced the depression of melting point of RES with maximum distortion of the crystal lattice was caused by PGMC and least by GTO. The two, long chain oleates, DO and GTO exhibited a shift of lipid peak in NLCs to higher melting points (116 and 111 °C) than the less lipophilic liquid lipids (103–104 °C). The attributes of liquid lipid also discriminate whether the particle growth during storage followed Oswald's ripening or coalescence. NLC containing GTO exhibited the highest stability in terms of maintenance of the PS and particle size distribution at 20 °C. This study provides vital insight on impact of liquid lipids and future strategy for rational design of stable NLC systems for delivery of various bio-actives for drug delivery applications.