Emergent Strategies for Natural Products Delivery.

Natural products have a wide range of applications with a profound impact in the medical and healthcare fields [...].

Formulation of a Phenol-Rich Extract from Unripe Olives (Olea europaea L.) in Microemulsion to Improve Its Solubility and Intestinal Permeability.

The beneficial properties of phenolic compounds from Olea europaea L. are well-known. An olive extract (OE) was prepared from unripe olives (Moraiolo cultivar). The study aimed to formulate OE into a microemulsion (ME) in oral dosage form. OE was extracted from olives with EtOH:H2O (80:20) and characterized by HPLC-DAD. ME composition was stated by a solubility and pseudo-ternary diagram. The ME was chemically and physically characterized, and its stability at 4 °C was analyzed for three months. The ability of the formulation to ameliorate the solubility and the intestinal permeability of OE was evaluated by a Parallel Artificial Membrane Permeability Assay (PAMPA) assay and Caco-2 cells. The total phenolic content of the extract was 39% w/w. The main constituent was oleuropein (31.0%), together with ligstroside (3.1%) and verbascoside (2.4%). The ME was prepared using Capryol 90 as the oily phase, and Cremophor EL and Transcutol (2:1) as surfactant and co-surfactant, respectively. ME droplet size was 14.03 ± 1.36 nm, PdI 0.20 ± 0.08, ζ-potential -1.16 ± 0.48. Stability of ME was confirmed for at least three months. The formulation was loaded with 35 mg/mL of OE, increasing the solubility of the extract by about four times. The enhanced permeability of OE was evaluated by PAMPA, as demonstrated by the Pe value (1.44 ± 0.83 × 10-6 cm/s for OE hydroalcoholic solution, 3.74 ± 0.34 × 10-6 cm/s for OE-ME). Caco-2 cell transport studies confirmed the same results: Papp was 16.14 ± 0.05 × 10-6 cm/s for OE solution and 26.99 ± 0.45 × 10-6 cm/s for OE-ME. ME proved to be a suitable formulation for oral delivery.

Enhanced dissolution, permeation and oral bioavailability of aripiprazole mixed micelles: In vitro and in vivo evaluation.

Aripiprazole (ARP) is an antipsychotic drug approved for the treatment of schizophrenia. It is poorly water-soluble and undergoes extensive hepatic metabolism and P-gp efflux, which lead to poor bioavailability and increased dose-related side effects. This study focuses on the preparation of mixed micelles (MM) to enhance the aqueous solubility, oral bioavailability, and blood-brain barrier permeation of ARP. For this purpose, Soluplus and D-α-tocopheryl polyethylene glycol 1000 succinate (TPGS) were selected for micelle preparation (ARP-MM). Micelles with borneol as penetration enhancer were also considered (ARP-B-MM). The optimized formulations have sizes of ca 50 nm, defined in distilled water, narrow size distribution (polydispersity index ≤0.1), and high encapsulation efficiency (greater than98%). Both formulations can be freeze-dried without losing their chemical-physical characteristics and are stable during storage for three months. The mixed micelles resulted stable in enzyme free-simulated gastric fluid (SGF, pH 1.2), simulated intestinal fluid (SIF, pH 6.8), and in serum. The in vitro ARP release was evaluated in the same biorelevant media, (SGF and SIF), and it disclosed that both micelles can give prolonged drug release. Furthermore, ARP solubility is greatly increased when loaded into mixed micelles. The absorption and efflux of ARP-loaded micelles were studied in vitro, employing two artificial membranes (Parallel Artificial Membrane Permeability Assay for the intestinal, PAMPA-GI, and the blood-brain barrier, PAMPA-BBB), to simulate the intestinal and brain epithelium, and the brain microvascular endothelial cell line hCMEC/D3. ARP-MM and ARP-B-MM increase the effective permeability of ARP by a factor of about three in the case of PAMPA-GI and about two for PAMPA-BBB. Furthermore, the P-gp mediated efflux was decreased by about six times in the case of ARP-MM and by about four times in the case of ARP-B-MM, compared to unformulated ARP. Finally, both ARP-loaded mixed micelles ameliorate the bioavailability of ARP, as demonstrated by the increase of the pharmacokinetic parameters, such as Cmax, AUC0-24h, and t1/2.

In Vitro Permeability of Saponins and Sapogenins from Seed Extracts by the Parallel Artificial Membrane Permeability Assay: Effect of in Vitro Gastrointestinal Digestion.

The permeability of saponins and sapogenins from fenugreek and quinoa extracts, as well as dioscin and diosgenin, was evaluated by the parallel artificial membrane permeability assay (PAMPA). The effect of the digestion process on permeability was determined, with previous development of a gastrointestinal process coupled to PAMPA. Saponins from both seeds displayed a moderate-to-poor permeability (>1 × 10-6 cm/s), although the digestion enhanced their permeability values in the order of 10-5 cm/s (p < 0.001). Sapogenins exhibited a similar permeability to that of saponins, although the digestion enhanced the permeability of sapogenins from quinoa (1.14 ± 0.47 × 10-5 cm/s) but not from fenugreek (2.33 ± 0.99 × 10-6 cm/s). An overall positive impact of coexisting lipids on the permeability was evidenced. PAMPA is shown as a useful, rapid, and easy tool for assessing the permeability of bioactive compounds from complex matrices, with the previous gastrointestinal process being a relevant step.

Niosomal Formulation of a Lipoyl-Carnosine Derivative Targeting TRPA1 Channels in Brain.

The transient receptor potential akyrin type-1 (TRPA1) is a non-selective cation channel playing a pivotal role in pain sensation and neurogenic inflammation. TRPA1 channels expressed in the central nervous system (CNS) have a critical role in the modulation of cortical spreading depression (CSD), which is a key pathophysiological basis of migraine pain. ADM_09 is a recently developed lipoic acid-based TRPA1 antagonist that is able to revert oxaliplatin-induced neuropathic pain and inflammatory trigeminal allodynia. In this context, aiming at developing drugs that are able to target TRPA1 channels in the CNS and promote an antioxidant effect, permeability across the blood-brain barrier (BBB) represents a central issue. Niosomes are nanovesicles that can be functionalized with specific ligands selectively recognized by transporters expressed on the BBB. In this work, the activity of ADM_09 on neocortex cultures was studied, and an efficient formulation to cross the BBB was developed with the aim of increasing the concentration of ADM_09 into the brain and selectively delivering it to the CNS rapidly after parenteral administration.

Comparison of Chitosan Nanoparticles and Soluplus Micelles to Optimize the Bioactivity of Posidonia oceanica Extract on Human Neuroblastoma Cell Migration.

Posidonia oceanica (L.) Delile is a marine plant endemic of Mediterranean Sea endowed with interesting bioactivities. The hydroalcholic extract of P. oceanica leaves (POE), rich in polyphenols and carbohydrates, has been shown to inhibit human cancer cell migration. Neuroblastoma is a common childhood extracranial solid tumor with high rate of invasiveness. Novel therapeutics loaded into nanocarriers may be used to target the migratory and metastatic ability of neuroblastoma. Our goal was to improve both the aqueous solubility of POE and its inhibitory effect on cancer cell migration. METHODS: Chitosan nanoparticles (NP) and Soluplus polymeric micelles (PM) loaded with POE have been developed. Nanoformulations were chemically and physically defined and characterized. In vitro release studies were also performed. Finally, the inhibitory effect of both nanoformulations was tested on SH-SY5Y cell migration by wound healing assay and compared to that of unformulated POE. RESULTS: Both nanoformulations showed excellent physical and chemical stability during storage, and enhanced the solubility of POE. PM-POE improved the inhibitory effect of POE on cell migration probably due to the high encapsulation efficiency and the prolonged release of the extract. CONCLUSIONS: For the first time, a phytocomplex of marine origin, i.e., P. oceanica extract, has enhanced in terms of acqueous solubility and bioactivity once encapsulated inside nanomicelles.

Nanostructured lipid carriers for oral delivery of silymarin: Improving its absorption and in vivo efficacy in type 2 diabetes and metabolic syndrome model.

Silymarin (SLM) is a mixture of flavonolignans extracted from the fruit of Silybum marianum L. Gaertn. which has been used for decades as a hepatoprotector. Silymarin has recently been proposed to be beneficial in type 2 diabetic patients. Constituents of SLM are poorly water-soluble and low permeable compounds, with consequently limited oral bioavailability. This study aimed to investigate the possibility of delivery of SLM via nanostructured lipid carriers (NLCs) to overcome these issues and for preparation of an oral dosage form. NLCs were prepared through an emulsion/evaporation/solidifying method. Cetyl palmitate:Lauroglycol 90 was selected as the lipid mixture and Brij S20 as surfactant. NLCs were chemically and physically characterized. Encapsulation efficiency was more than 92%. The storage stability of the NLC suspension was also investigated and the freeze-drying process was taken into consideration. After assessing the stability of the formulation in a simulated gastrointestinal environment, the release of SLM was monitored in different pH conditions. In vitro experiments with artificial membranes (PAMPA) and Caco-2 cells revealed that the NLCs enhanced the permeation of SLM. Active processes are involved in the internalization of NLCs, as evidenced by cellular uptake studies. After preliminary toxicological studies, the formulation was studied in vivo in a streptozotocin (STZ)-induced diabetic mouse model in the presence of metabolic syndrome. The formulation was also compared to an NLC containing stearic acid:Capryol 90, to evaluate the effect of the lipid matrix on the in vivo performance of nanocarriers. Finally, hepatic histopathological analyses were also conducted. Both SLM-loaded NLCs exhibited in vivo a significant down-regulation of blood glucose and triglyceride levels better than free SLM, with a liver-protective effect. Furthermore, both formulations showed a significant anti-hyperalgesic effect on STZ-induced neuropathy.

Successful Brain Delivery of Andrographolide Loaded in Human Albumin Nanoparticles to TgCRND8 Mice, an Alzheimer's Disease Mouse Model.

Andrographolide (AG) was encapsulated in human albumin nanoparticles (AG NPs), and their crossing properties of the blood-brain barrier (BBB), brain distribution, and effects in TgCRND8 mice were evaluated. The development of appropriate NP formulations is mandatory because of the scarce BBB permeability properties of AG. Developed NPs had proper size (mean size: 159.2 ± 4.5 nm), size distribution (PDI nearby 0.12 ± 0.01), and ζ potential (-24.8 ± 1.2 mV), which were not affected by sodium fluorescein (NAF) loading. When AG was loaded to NPs, it slightly affected their size (210.4 ± 3.2 nm) and ζ potential (-20.3 ± 1.5) but not the PDI. Both NAF and AG had a remarkable encapsulation efficiency (more than 99%). The in vitro release of AG from the NPs reached the highest percentage (48%) after 24 h, and the Higuchi's equation was found to be the best fitting model (R2 = 0.9635). Both AG and AG NPs did not alter the viability of N2a murine neuroblastoma cells when compared with the untreated control cells. In the step-down inhibitory avoidance test, AG NPs administered to TgCRND8 mice significantly improved their performance (P < 0.0001), reaching levels comparable to those displayed by wild-type mice. In the object recognition test, treated and untreated animals showed no deficiencies in exploratory activity, directional movement toward objects, and locomotor activity. No cognitive impairments (discrimination score) were detected in TgCRND8 mice (P < 0.0001) treated with AG NPs. After acute intravenous administration (200 µl), NPs loaded with the probe NAF were detected in the brain parenchyma of TgCRND8 mice. Immunofluorescent analyses evidenced the presence of NPs both in the pE3-Aß plaque surroundings and inside the pE3-Aß plaque, indicative of the ability of these NPs to cross the BBB and to penetrate in both undamaged and damaged brain tissues. Furthermore, the immunohistochemical analysis of GFAP-positive astrocytes in the hippocampus of Tg mice evidenced the anti-inflammatory activity of AG when AG NPs were intraperitoneally administered. AG was not effective in counteracting amyloid Aß aggregation and the resulting toxicity but significantly decreased the oxidative stress levels. In conclusion, AG NPs have extraordinary versatility, nontoxicity, nonimmunogenicity, strong biocompatibility, high biodegradability, and astonishing loading capacity of drug.

Formulation of Nanomicelles to Improve the Solubility and the Oral Absorption of Silymarin.

Two novel nanomicellar formulations were developed to improve the poor aqueous solubility and the oral absorption of silymarin. Polymeric nanomicelles made of Soluplus and mixed nanomicelles combining Soluplus with d-α-tocopherol polyethylene glycol 1000 succinate (vitamin E TPGS) were prepared using the thin film method. Physicochemical parameters were investigated, in particular the average diameter, the homogeneity (expressed as polydispersity index), the zeta potential, the morphology, the encapsulation efficiency, the drug loading, the critical micellar concentration and the cloud point. The sizes of ~60 nm, the narrow size distribution (polydispersity index ≤0.1) and the encapsulation efficiency >92% indicated the high affinity between silymarin and the core of the nanomicelles. Solubility studies demonstrated that the solubility of silymarin increased by ~6-fold when loaded into nanomicelles. Furthermore, the physical and chemical parameters of SLM-loaded formulations stored at room temperature and in refrigerated conditions (4 °C) were monitored over three months. In vitro stability and release studies in media miming the physiological conditions were also performed. In addition, both formulations did not alter the antioxidant properties of silymarin as evidenced by the 1,1-Diphenyl-2-picrylhydrazyl radical (DPPH) assay. The potential of the nanomicelles to increase the intestinal absorption of silymarin was firstly investigated by the parallel artificial membrane permeability assay. Subsequently, transport studies employing Caco-2 cell line demonstrated that mixed nanomicelles statistically enhanced the permeability of silymarin compared to polymeric nanomicelles and unformulated extract. Finally, the uptake studies indicated that both nanomicellar formulations entered into Caco-2 cells via energy-dependent mechanisms.

Chitosan coated human serum albumin nanoparticles: A promising strategy for nose-to-brain drug delivery.

The aim of the present study was the development of human serum albumin nanoparticles (HSA NPs) as nose-to-brain carrier. To strengthen, the efficacy of nanoparticles as drug delivery system, the influence of chitosan (CS) coating on the performance of HSA NPs was investigated for nasal application. HSA NPs were prepared by desolvation technique. CS coating was obtained adding the CS solution to HSA NPs. The mean particle sizes was 241 ±â€¯18 nm and 261 ±â€¯8 nm and the ζ-potential was -47 ±â€¯3 mV and + 45 ±â€¯1 mV for HSA NPs and CS-HSA NPs, respectively. The optimized formulations showed excellent stability upon storage both as suspension and as freeze-dried product after 3 months. The mucoadhesion properties were assessed by turbidimetric and indirect method. NPs were loaded with sulforhodamine B sodium salt as model drug and the effect of CS coating was investigated performing release studies, permeation and uptake experiments using Caco-2 and hCMEC/D3 cells as model of the nasal epithelium and blood-brain barrier, respectively. Furthermore, ex vivo diffusion experiments have been carried out using rabbit nasal mucosa. Finally, the ability of the formulations to reversibly open tight and gap junctions was explored by western blotting and RT-PCR analyzing in both Caco-2 and hCMEC/D3 cells.

Stable, Monodisperse, and Highly Cell-Permeating Nanocochleates from Natural Soy Lecithin Liposomes.

(1) Background: Andrographolide (AN), the main diterpenoid constituent of Andrographis paniculata, has a wide spectrum of biological activities. The aim of this study was the development of nanocochleates (NCs) loaded with AN and based on phosphatidylserine (PS) or phosphatidylcholine (PC), cholesterol and calcium ions in order to overcome AN low water solubility, its instability under alkaline conditions and its rapid metabolism in the intestine. (2) Methods: The AN-loaded NCs (AN⁻NCs) were physically and chemically characterised. The in vitro gastrointestinal stability and biocompatibility of AN⁻NCs in J77A.1 macrophage and 3T3 fibroblasts cell lines were also investigated. Finally, the uptake of nanocarriers in macrophage cells was studied. (3) Results: AN⁻NCs obtained from PC nanoliposomes were suitable nanocarriers in terms of size and homogeneity. They had an extraordinary stability after lyophilisation without the use of lyoprotectants and after storage at room temperature. The encapsulation efficiency was 71%, while approximately 95% of AN was released in PBS after 24 h, with kinetics according to the Hixson⁻Crowell model. The in vitro gastrointestinal stability and safety of NCs, both in macrophages and 3T3 fibroblasts, were also assessed. Additionally, NCs had extraordinary uptake properties in macrophages. (4) Conclusions: NCs developed in this study could be suitable for both AN oral and parental administration, amplifying its therapeutic value.

Solid Lipid Nanoparticles and Chitosan-coated Solid Lipid Nanoparticles as Promising Tool for Silybin Delivery: Formulation, Characterization, and In vitro Evaluation.

BACKGROUND: Silybin (Sb) is the major flavolignan of the extract of Silybum marianum. It is used for the treatment of various acute and chronic liver toxicities, inflammation, fibrosis and oxidative stress. Many studies indicate that Sb is also active against different carcinomas and it has been very recently proposed to be beneficial in type 2 diabetes patients. However, Sb is a low water soluble and low permeable compound. OBJECTIVE: In this study, Solid Lipid Nanoparticles (SLNs) were proposed to enhance the solubility and the intestinal absorption of Sb. METHODS: SLNs were made of stearic acid and Brij 78 and subsequently coated with chitosan. Formulations were physically and chemically characterized. Stability studies were also assessed. Sb in vitro release was evaluated in different pH media. In vitro permeability test with artificial membranes and Caco-2 cells were performed. Cellular uptake and mucoadhesion studies were conducted. RESULTS: Both nanoparticles were found to be stable. In vitro release indicated that SLNs may prevent burst release and gastric degradation of Sb. Higher extent of Sb permeation was observed for both nanoparticles in PAMPA and Caco-2 cell monolayer models. The results of the cellular uptake study suggested the involvement of active endocytic processes. Chitosan significantly improves mucoadhesion properties of nanoparticles. CONCLUSIONS: Together with the excellent stability, strong mucoadhesive property, and slow release, chitosan coated SLNs demonstrated promising potential to enhance absorption of hydrophobic Sb after oral administration.

Nanocarriers: A Successful Tool to Increase Solubility, Stability and Optimise Bioefficacy of Natural Constituents.

Natural products are fascinating molecules in drug discovery for their exciting structure variability and also for their interaction with various targets. Drugs multi-targeting effect represents a more realistic approach to develop successful medications for many diseases. However, besides a large number of successful in vitro and in vivo studies, most of the clinical trials fail. This is generally related to the scarce water solubility, low lipophilicity and inappropriate molecular size of natural compounds, which undergo structural instability in biological milieu, rapid clearance and high metabolic rate. Additionally, some molecules are destroyed in gastric juice or suffer to a massive pre-systemic metabolism in the liver, when administered orally, limiting their clinical use. A reduced bioavailability can also be linked to drug distribution/accumulation in non-targeted tissues and organs that increase the side effects lowering the therapeutic efficacy and patient compliance. Nanomedicine represents a favourable tool to increase bioavailability and activities of natural products. Generally, nanovectors provide a large surface area and can overcome anatomic barriers. Each nanovector has its own advantages, disadvantages, and characteristics. In this review, different nanocarriers made of compounds which are Generally Recognized As Safe (GRAS) for the delivery of natural products, marketed as food supplements and medicines are reported.

Stealth and Cationic Nanoliposomes as Drug Delivery Systems to Increase Andrographolide BBB Permeability.

(1) Background: Andrographolide (AG) is a natural compound effective for the treatment of inflammation-mediated neurodegenerative disorders. The aim of this investigation was the preparation of liposomes to enhance the penetration into the brain of AG, by modifying the surface of the liposomes by adding Tween 80 (LPs-AG) alone or in combination with Didecyldimethylammonium bromide (DDAB) (CLPs-AG). (2) Methods: LPs-AG and CLPs-AG were physically and chemically characterized. The ability of liposomes to increase the permeability of AG was evaluated by artificial membranes (PAMPA) and hCMEC/D3 cells. (3) Results: Based on obtained results in terms of size, homogeneity, ζ-potential and EE%. both liposomes are suitable for parenteral administration. The systems showed excellent stability during a month of storage as suspensions or freeze-dried products. Glucose resulted the best cryoprotectant agent. PAMPA and hCMEC/D3 transport studies revealed that LPs-AG and CLPs-AG increased the permeability of AG, about an order of magnitude, compared to free AG without alterations in cell viability. The caveolae-mediated endocytosis resulted the main mechanism of up-take for both formulations. The presence of positive charge increased the cellular internalization of nanoparticles. (4) Conclusions: This study shows that developed liposomes might be ideal candidates for brain delivery of AG.

Enhanced Solubility and Permeability of Salicis cortex Extract by Formulating as a Microemulsion.

A microemulsion system was developed and investigated as a novel oral formulation to increase the solubility and absorption of Salicis cortex extract. This extract possesses many pharmacological activities, in particular, it is beneficial for back pain and osteoarthritic and rheumatic complaints. In this work, after qualitative and quantitative characterization of the extract and the validation of an HPLC/diode array detector analytical method, solubility studies were performed to choose the best components for microemulsion formulation. The optimized microemulsion consisted of 2.5 g of triacetin, as the oil phase, 2.5 g of Tween 20 as the surfactant, 2.5 g of labrasol as the cosurfactant, and 5 g of water. The microemulsion was visually checked, characterized by light scattering techniques and morphological observations. The developed formulation appeared transparent, the droplet size was around 40 nm, and the ζ-potential result was negative. The maximum loading content of Salicis cortex extract resulted in 40 mg/mL. Furthermore, storage stability studies and an in vitro digestion assay were performed. The advantages offered by microemulsion were evaluated in vitro using artificial membranes and cells, i.e., parallel artificial membrane permeability assay and a Caco-2 model. Both studies proved that the microemulsion was successful in enhancing the permeation of extract compounds, so it could be useful to ameliorate the bioefficacy of Salicis cortex.

Lipid Nanocarriers for Oral Delivery of Serenoa repens CO2 Extract: A Study of Microemulsion and Self-Microemulsifying Drug Delivery Systems.

The aim of this study was the development and characterization of lipid nanocarriers using food grade components for oral delivery of Serenoa repens CO2 extract, namely microemulsions (MEs) and self-microemulsifying drug delivery systems (SMEDDSs) to improve the oral absorption. A commercial blend (CB) containing 320 of S. repens CO2 extract plus the aqueous soluble extracts of nettle root and pineapple stem was formulated in two MEs and two SMEDDSs. The optimized ME loaded with the CB (CBM2) had a very low content of water (only 17.3%). The drug delivery systems were characterized by dynamic light scattering, transmission electron microscopy, and high-performance liquid chromatography (HPLC) with a diode-array detector analyses in order to evaluate the size, the homogeneity, the morphology, and the encapsulation efficiency. ß-carotene was selected as marker for the quantitative HPLC analysis. Additionally, physical and chemical stabilities were acceptable during 3 wk at 4 °C. Stability of these nanocarriers in simulated stomach and intestinal conditions was proved. Finally, the improvement of oral absorption of S. repens was studied in vitro using parallel artificial membrane permeability assay. An enhancement of oral permeation was found in both CBM2 and CBS2 nanoformulations comparing with the CB and S. repens CO2 extract. The best performance was obtained by the CBM2 nanoformulation (~ 17%) predicting a 30 - 70% passive oral human absorption in vivo.

Topical formulations of delta-aminolevulinic acid for the treatment of actinic keratosis: Characterization and efficacy evaluation.

Actinic keratosis (AK) is a pre-cancerous disease, with worldwide increasing incidence, which consists in squamous cutaneous lesion caused by excessive exposure to ultraviolet radiation. An established treatment option is photodynamic therapy (PDT), based on light, oxygen and a photosensitizer. The most widely used is 5-aminolevulinic acid (ALA) which however, being a hydrophilic molecule, has difficultly penetrating the skin to achieve the desired therapeutic effect. To solve this limit, the present study provides for the development of three galenic gel formulations (Natrosol, Sepigel and Carbopol) containing 10% w/w of ALA for the treatment of AK with PDT and their comparison with a lipophilic cream used in the Hospital. The aim of this study is to offer an appealing topical treatment that improves patients' observance and compliance. Formulations were characterized in terms of chemical, physical and microbiological stability, viscosity and pH. An HPLC-DAD analytical method was also developed and validated. Sepigel gel resulted the best gel formulation in terms of technological characteristics and stability. A comparative study between this gel and the lipophilic cream was assessed, by evaluating the therapeutic efficacy and the compliance of the patients.

Solid lipid nanoparticles for delivery of andrographolide across the blood-brain barrier: in vitro and in vivo evaluation.

Andrographolide is a major diterpenoid of Andrographis paniculata and possesses several biological activities, including protection against oxidative stress mediated neurotoxicity, inflammation-mediated neurodegeneration, and cerebral ischemia. However, this molecule shows low bioavailability, poor water solubility, and high chemical and metabolic instability. The present study reports preparation of solid lipid nanoparticles (SLN) to deliver andrographolide (AG) into the brain. SLN were prepared using Compritol 888 ATO as solid lipid and Brij 78 as surfactant and applying emulsion/evaporation/solidifying method as preparative procedure. Nanoparticles have a spherical shape, small dimensions, and narrow size distribution. Encapsulation efficiency of AG-loaded SLN was found to be 92%. Nanoparticles showed excellent physical and chemical stability during storage at 4°C for one month. The lyophilized product was also stable at 25°C during the same period. SLN remained unchanged also in the presence of human serum albumin and plasma. In vitro release at pH 7.4 was also studied. The release of AG was prolonged and sustained when the compound was entrapped in SLN. The ability of SLN to cross the blood-brain barrier (BBB) was evaluated first in vitro by applying a permeation test with artificial membrane (parallel artificial membrane permeability assay, PAMPA) to predict passive and transcellular permeability through the BBB, and then by using hCMEC/D3 cells, a well-established in vitro BBB model. In vitro results proved that nanoparticles improved permeability of AG compared to free AG. Fluorescent nanoparticles were then prepared for in vivo tests in healthy rats. After intravenous administration, fluorescent SLN were detected in brain parenchyma outside the vascular bed, confirming their ability to overcome the BBB.

Andrographolide-loaded nanoparticles for brain delivery: Formulation, characterisation and in vitro permeability using hCMEC/D3 cell line.

Andrographolide (AG) is a major diterpenoid of the Asian medicinal plant Andrographis paniculata which has shown exciting pharmacological potential for the treatment of inflammation-related pathologies including neurodegenerative disorders. Conversely, the low bioavailability of AG still represents a limiting factor for its use. To overcome these limitations, AG was loaded into human serum albumin based nanoparticles (HSA NPs) and poly ethylcyanoacrylate nanoparticles (PECA NPs). HSA NPs were prepared by thermal (HSAT AG NPs) and chemical cross-linking (HSAC AG NPs), while PECA AG NPs were produced by emulsion-polymerization. NPs were characterized in terms of size, zeta (ζ)-potential, polydispersity, and release studies of AG. In addition, the ability of free AG and AG-loaded in PECA and HSAT NPs to cross the blood-brain barrier (BBB) was assessed using an in vitro BBB model based on human cerebral microvascular endothelial cell line (hCMEC/D3). For BBB drug permeability assays, a quantitative UPLC-MS/MS method for AG in Ringer HEPES buffer was developed and validated according to international regulatory guidelines for industry. Free AG did not permeate the BBB model, as also predicted by in silico studies. HSAT NPs improved by two-fold the permeation of AG while maintaining the integrity of the cell layer, while PECA NPs temporarily disrupted BBB integrity.