Emerging Techniques for Herbosomes

Abstract

Herbosomes are a relatively new technology that involves encapsulating herbal extracts in liposomes, which are tiny spheres made of phospholipids. This allows for better absorption of the herbal compounds into the body. Herbosomes have a higher bioavailability compared to traditional herbal extracts, improved stability and can be designed to target specific areas of the body, as well as reduced side effects as they can be delivered in smaller doses. The production of herbosomes involves the use of various techniques including solvent injection, thin-film hydration, and sonication. The production of herbosomes involves the use of various techniques that aim to create stable and effective nanocarriers for herbal extracts. There is limited research available on the safety and toxicity of herbosomes specifically, but studies have been conducted on the safety of lipid-based nanoparticles in general. It is important to note that the safety and toxicity of herbosomes may vary depending on the specific herbal extract and lipid used in their formulation. Further research is needed to fully understand the potential risks and benefits of using herbosomes as a drug delivery system. In conclusion, herbosomes offer several advantages over traditional herbal extracts, making them a promising technology for the development of new herbal products.

1. Introduction

Extensive research and clinical trials have focused on natural products and secondary metabolites as potential treatments for various human diseases. Medicinal plants and their bioactive components have long been utilized in the food and pharmaceutical industries and for treating diverse ailments [1]. The increased use of herbal drugs can be attributed to several significant factors, including the limitations of modern medicine in addressing all human pathologies, concerns regarding the reliability and safety of synthetic drugs, and the demonstrated efficacy of many natural products in yielding superior outcomes compared to synthetic drugs, without accompanying adverse effects [2]. However, their utilization in pharmaceutical and food sectors is constrained by challenges such as poor water solubility and stability concerns. The inadequate absorption of active phytochemicals arises from factors such as the large, multi-ring structures of polyphenols hindering their passive absorption mechanisms and the limited solubility of active compounds in water or lipids impeding their passage across the outer membrane of gastrointestinal cells [1, 3]. Moreover, upon ingestion, they may undergo various reactions during the digestion process, potentially resulting in significant modifications to their molecular structure and consequently affecting their bioactive properties [4].

In response to these challenges, pharmaceutical research has focused on developing lipid-based drug delivery systems to improve bioavailability while preserving therapeutic efficacy [5]. One such approach involves integrating standardized herbal extracts into phospholipids, forming complexes known as “herbosomes” or “phytosomes.” These vesicular drug delivery systems, engineered to enhance the absorption and bioavailability of poorly soluble drugs, consist of phospholipids and natural active phytochemicals forming complexes through interactions with plant extracts in a solvent without protons [6]. Phospholipids, crucial for constructing cell membranes, serve as natural digestive aids and carriers for nutrients in both fat and water. They demonstrate compatibility with aqueous and lipid environments, allowing for effective oral absorption. The primary phospholipid utilized in phytosome formation is phosphatidylcholine, sourced from soybeans (Glycine max). Phospholipids, being lipophilic substances, are capable of forming complexes with polyphenolics, facilitating their absorption [7, 8].

Herbosomes, formed by loading phytoconstituents into phospholipids, exhibit improved physical stability due to the formation of hydrogen bonds between phospholipids and phytoconstituents. This enhances the absorption of hydrophilic polar phytoconstituents, leading to increased bioavailability and greater therapeutic benefits [9]. Herbosomes represent an innovative formula of botanicals and phytoconstituents, exhibiting enhanced absorption through both oral and transdermal routes when encapsulated with phosphatidylcholine. This technology serves as a bridge between conventional phytoconstituent delivery systems and emerging drug delivery methodologies [8, 10]. Demonstrating enhanced pharmacological and pharmacokinetic properties compared to conventional preparations, herbosomes have the lipid-soluble phosphatidyl component enveloping the hydrophilic phytoconstituent-choline complexes entirely [11]. Several methodologies, including solvent evaporation, rotatory evaporation, anti-solvent precipitation, freeze-drying, and solvent ether injection, are utilized for herbosome preparation. Evaluation of herbosomes involves techniques such as UV-spectra analysis, differential scanning calorimetry (DSC), assessment of drug entrapment and loading capacity, measurement of surface tension activity, and in vitro/in vivo studies [12]. Notable advantages include high drug encapsulation, stability attributed to chemical bonding, flexibility in administration routes, and increased bioavailability, including enhanced absorption, minimized side effects, controlled release, and targeted delivery, necessitating lower dosages of active constituents for biological effects, even for polar phytoconstituents [13, 14].

2. Structure of a phytosome

The four essential components required to produce herbosomes are phospholipids, phytoactive ingredients, solvents, and a specific ratio relationship involved in the creation of herbosome as shown in Figure 1.

1. Phospholipids: they mainly constitute egg yolk and plant seeds that are considered the most prevalent natural sources of phospholipids. Phospholipids produced in an industrial setting are available for commercial purposes. Phosphatidyl choline is the most used phospholipid in the formation of phospholipid complexes. Phosphatidylcholine has a moderate solubility in both aqueous and lipid environments. In addition to its amphipathic properties, it is considered as an essential component of cell membranes and exhibits high biocompatibility and low toxicity. Phospholipids are used as a vehicle-creating component in the creation of phytosomes [14].

2. Phytoactive constituents: the phytoactive constituents are typically selected based on significant in vitro pharmacological effects. Water-soluble flavonoids, such as quercetin, catechin, and silybin, are unable to cross biological membranes, unlike lipophilic curcumin and rutin, which are insoluble in aqueous gastrointestinal fluids. In the aqueous phase, phytosome complexes enhance the water solubility of lipophilic flavonoids and the membrane permeability of hydrophilic flavonoids. Furthermore, flavonoids can be shielded from external impacts, such as hydrolysis, photolysis, and oxidation, by forming complexes [13].

3. Solvents: different solvents are used for complexation, mainly aromatic hydrocarbons, halogen derivatives, methylene chloride, ethyl acetate, and cyclic ethers have all been employed in the past to form phytophospholipid complexes. Protonic solvents, such as ethanol and methanol, have recently been used successfully to form phospholipid complexes. Ethanol is a useful and popular solvent because it leaves behind fewer residues and causes minimal damage [13].

4. Stoichiometric ratio: phyto-phospholipid complexes are formed by reacting a synthetic or natural phospholipid with the active components in a molar ratio ranging from 0.5 to 2.0 in many cases. A stoichiometric ratio of 1:1 is considered the most efficient for creating phytosome complexes because it enhances interaction between the two components. The stoichiometric ratio of active components and phospholipids should be experimentally adjusted for various purposes, such as achieving maximal drug loading, in different types of pharmaceuticals [14].

5. pH maintenance: to maintain the consistency of the preparation’s pH, a buffering agent is used. Two commonly used buffering agents are 7% (v/v) saline phosphate buffer at pH 6.5 and ethanol tris buffer at pH 6.5. The purpose of using a buffer is to maintain the hydration of phytosomes [14].

Figure 1.

Principle of herbosome formation.

3. Preparation of phytosomes

The process of producing phytosomes involves the following steps: step 1: phospholipids and herbal compounds are present in aprotic media, such as dioxane and acetone; step 2: hydrogen bond formation; step 3: wrapping the non-polar tail around the polar complex.

Traditional methods: there are mainly three methods available for the preparation of phytosome; solvent evaporation method, rotary evaporation method, and anti-solvent precipitation method as summarized in Figure 2 [5].

Figure 2.

Traditional methods for the preparation of herbosome: (a) antisolvent precipitation technique; (b) rotary evaporation technique; (c) solvent evaporation technique.

Anti-solvent precipitation technique: involves combining a fixed amount and quantity of phospholipid with herbal extract in a suitable ratio in a 100 mL round bottom flask. The mixture is then refluxed with 20 mL of dichloromethane for 2 hours at a specific temperature.

Rotary evaporation: is a technique in which a specified amount of herb extract is mixed with phospholipids dissolved in 30 mL of tetrahydrofuran at a specific temperature.

The solvent evaporation technique: involves mixing the specified quantity of herbal extract with phospholipids in a 100 mL round-bottom flask. The solution was refluxed with 20 mL of acetone for 2 hours at a temperature ranging from 50–60°C. The mixture was then concentrated to 5–10 mL, filtered to collect the precipitate, and the formed complex was finally stored in an amber-colored glass bottle at 25°C.

Ether-injection technique: dissolve the drug-lipid complex in an organic solvent. Inject this mixture slowly into a heated aqueous solution to form amphiphilic vesicles with different structures.

Sonication technique: place the appropriate amount of phospholipid and cholesterol in a flat-bottomed flask, dissolved in 10 mL of chloroform. Subsequently, sonicate the mixture in a bath sonicator using a rotating evaporator at 40°C, while reducing the pressure to remove organic solvents [14].

Traditional methods exhibit several drawbacks, including multistep processes, difficulty in extraction, and time consumption [13].

Non-traditional methods: supercritical fluid methods can be used to alter the size, shape, and morphology of materials of interest, in addition to other benefits such as high product purity, control of crystal polymorphism, the ability to process thermolabile substances in a single step, and eco-friendly technology (Figure 3).

Figure 3.

Preparation of herbosomes by non-traditional methods using supercritical fluid technique.

Supercritical fluids techniques: utilize a supercritical fluid, typically CO₂, as an anti-solvent to reduce the solute’s solubility in the solvent.

Gas anti-solvent technique (GAS): it is not necessary for the CO₂ gas used as an antisolvent to be in a supercritical state. The substance is injected into the solution within a closed chamber, ideally from the bottom, to ensure uniform mixing. As a result of CO₂ gas dissolving, the organic solvent’s ability to dissolve solutes is reduced, leading to the precipitation of solutes. The particles are washed with extra antisolvent to eliminate any remaining solvent. In comparison to the solvent-antisolvent technique, the GAS technique yields superior results when scaled up to industrial levels [13].

4. Evaluation, characterization, and stability

4.1 Introduction

A variety of nanoparticles, differing in both quantity and materials, are currently being developed. These materials exist in various chemical forms, such as micelles, metal oxides, or large biomolecules. This diversity underscores the need for the development of enhanced characterization methods and protocols that provide greater precision and increased credibility. However, each characterization technique has its own set of advantages and limitations (Figure 4). To overcome these constraints, it is advisable to use a combination of methods to effectively characterize individual nanoparticles. When choosing these methods of characterization, it is essential to ensure that they are suitable for the intended purpose [15, 16].

Figure 4.

Methods employed for characterization: (1) high performance liquid chromatography, (2) X-ray diffraction analysis, (3) scanning electron microscopy, (4) transmission electron microscopy, and (5) differential scan.

4.2 Characterization of herbosomes

1. Entrapment efficiency: to evaluate the efficiency of drug entrapment within planterosomes, we employ the ultracentrifugation method [1]. This method aids in the determination of the percentage of the drug present within the phospholipid mesh. In all phytosome formulations, approximately 100% of the drug is present [17]. The entrapment efficacy is calculated using the following formula:

$$\begin{array}{ll}\%\text{entrapment efficacy}=& (\text{amount of drug in sediment}\\ & /\text{total amount of drug added})\times 100\end{array}$$E1

1. Drug content: the quantity of drug in herbosomes is typically determined using a modified high-performance liquid chromatography method or by UV analysis. One way to measure the drug content is to dissolve a known quantity of phyto-phospholipid dispersion in 10 mL of methanol. The drug concentration of the phyto-phospholipid complex is then determined. After appropriate dilution, the absorbance is measured using spectroscopic techniques at a specific wavelength, and the drug content is calculated using the formula:

$$\begin{array}{ll}\%\text{drug content}=& (\text{actual drug content in phyto}-\text{phospholipid complex}\\ & /\text{theoretical yield})\times 100\end{array}$$E2

1. In vitro drug release studies: we utilize the Franz diffusion cell method or dialysis bag in combination with various kinetic models. These methods help to identify the mechanisms involved in the release of drug content. Furthermore, an in vitro dissolution test is conducted to understand the drug release process [14].

2. Visualization: the most commonly used visualization methods are transmission electron microscopy (TEM) and scanning electron microscopy (SEM). Additionally, when the SEM analysis of nanoparticles (NPs) does not yield clear results regarding the size and shape of the NPs due to its very high resolution, field emission scanning electron microscopy (FESEM) is used [18].

3. X-ray diffraction analysis (XRD): can provide valuable assistance in analyzing various particles. This method is utilized for identifying crystalline compounds [19] and for determining particle roughness, topography, surface area, and surface chemistry.

4. Transition temperature: a thermoanalytical method, such as differential scanning calorimetry (DSC), can be employed to assess the transition temperature of vesicular lipid systems. DSC plays a crucial role in elucidating changes in material properties in response to temperature variations. This tool is valuable for determining the crystal structure of the active pharmaceutical ingredient (API). Several phenomena are observed, including temperature transitions, the disappearance of endothermic peaks, alterations in relative peak areas, and the emergence of new peaks. These observations provide valuable insights into the melting and crystallization behavior of the sample being investigated [20].

5. Vesicle stability and zeta potential: the stability of vesicles can be assessed over an extended period through comprehensive measurements that include size, zeta potential, and structural characteristics. Zeta potential, which is the surface charge, is defined as the difference in electric potential (ΔV) between the dispersion medium and the stationary fluid layer on the surface of the dispersed phase [21]. A zeta potential of ±30 mV or ± 20 mV is preferred for high physical stability. For the determination of both size and zeta potential, dynamic light scattering (DLS) coupled with a computerized inspection system and photon correlation spectroscopy (PCS) proves to be a valuable approach [21]. Simultaneously, transmission electron microscopy (TEM) is used to observe structural changes, as mentioned earlier.

6. Spectroscopic techniques: to confirm the formation of a complex or investigate the interaction between the plant-based component and the phospholipids, scientists utilize spectroscopic techniques such as nuclear magnetic resonance (NMR), Fourier transform infrared spectroscopy (FTIR), and X-ray diffraction (XRD). This involves comparing the outcomes of the individual elements with those of the complexes [5].

5. Applications of herbosomes

The utilization of phytochemicals in pharmaceutical products is constrained by several factors, including solubility, bioavailability, and stability. The large molecular size of most phytochemicals, along with their high lipophilicity or hydrophilicity, affects their absorption and bioavailability and, consequently, their therapeutic effectiveness [1, 4]. Moreover, the stability of phytochemicals depends on temperature, pH, and enzymes. Degradation of phytoconstituents through hydrolysis, oxidation, or enzymatic activity during processing, storage, or after administration can limit their effectiveness. Furthermore, phytochemicals are recognized by the host immune system as foreign antigens, eliciting an immune response that leads to rapid clearance and reduced clinical effects. The use of phytochemicals in nanocarriers, such as herbosomes, presents an appealing strategy for overcoming their in vivo limitations and enhancing their therapeutic effects [22, 23]. Herbosomes are drug delivery systems that have several therapeutic effects, including anticancer, hepatoprotective, and wound healing properties. The details of their applications are discussed in the following section and summarized in Table 1.

Study	Phospholipid	Herbal component	Application	Therapeutic outcomes
Murugesan et al. [24]	Milk phospholipids	Aloe vera	Breast cancer	Cytotoxicity on the MCF-7 cell line
Wanjiru et al. [25]	Soy phosphatidylcholine	Moringa oleifera		Cytotoxicity on the 4 T1 breast cancer cell line Induction of apoptosis and cell proliferation
Alhakamy et al. [26]	Soybean phosphatidylcholine	Quercetin		Cytotoxicity against the MCF-7 cell line Induction of apoptosis and necrotic cell death
Sabzichi et al. [27]	Phosphatidylcholin, phosphatidyletanolamine, and phosphatidylserin	Luteolin		Cytotoxicity against MDA-MB231 Enhance sensitivity of doxorubicin
Talaat et al. [28]	Soy phosphatidylcholine	Fisetin		Cytotoxicity against MDA-MB231 Induction of apoptosis Inhibition of the activity of TGF-β and MMP-9 Enhance E-cadherin expression levels
Komeil et al. [29]	Soy phosphatidylcholine	Genistein		Significant reduction in tumor size Significant reduction in levels of CEA and CA15.3
Kumar et al. [30]	Soy lecithin	Taxifolin		Cytotoxicity against the MCF-7 cell line Radical scavenging activity (H2O2, NO, and DHHP)
Hashemzehi et al. [31]	N/A	Curcumin		Cytotoxicity against MCF-7 cell lines Reduction in cell invasion Increased expression of E-cadherin and MMP-9 Reduced levels of MDA and thiol Inhibition of mTOR and Wnt/β-catenin signaling
El-Far et al. [32]	Soy phosphatidylcholine	Monascin, ankaflavin, and resveratrol		Cytotoxicity on the MCF-7 cell line Decrease tumor weight and volume Reduce levels of VEGF, NF-κB, and CD1 Induce apoptosis and necrosis
Xu et al. [33]	Lecithin	Diosgenin	Lung cancer	Cytotoxicity against A549 and PC9 cell lines Induction of apoptosis
Al-Rabia et al. [34]	Soy phosphatidylcholine	Curcumin	Prostate cancer	Cytotoxicity against the PC3 cell line Induction of apoptosis and necrosis Disrupt the mitochondrial membrane potential
Pastorelli et al. [35]	Phosphatidylcholine	Curcumin		Increase in disease control rate and overall survival rate Reduced levels of IL-6, sCD40L, and CRP
Li et al. [36]	Soy phosphatidylcholine	Mitomycin C	Cervical cancer	Cytotoxicity on HeLa cell lines Reduction in tumor weight and volume
Komeil et al. [37]	Phosphatidylcholine solubilized in medium-chain and long-chain TGs	Genistein	Liver cancer	Cytotoxicity on HepG2 cells Induction of apoptosis Reduced the levels of VEGF and MMP-9
Teng et al. [38]	Phosphatidylcholine	Curcumin		Cytotoxicity against the Huh-7 cell line Reduced tumor volume Decreased lipid and leukocyte accumulation
Mazumder et al. [39]	Soy phosphatidylcholine	Sinigrin	Skin cancer	Cytotoxicity against the A375 cell line
Mukherjee et al. [40, 41]	Phosphatidylcholine	Curcumin	Glioblastoma	Improved survival of mice Reduced the number of CD68 high GBM tumor cells Increased the level of iNOS Reduced the level of ARG1 Induced polarization of M2-TAMs into the M1 phenotype Induced the expression of monocyte chemotactic protein-1
Singh et al. [42]	Phosphatidylcholine	Silibinin	Colorectal cancer	Enhanced antitumor activity Inhibition of angiogenesis Reduced the expression of VEGF, COX, iNOS, and HIF-1α
Marjaneh et al. [43]	N/A	Curcumin		Cytotoxicity on the CT26 cell line Induced cell death Reduced cell invasion Reduced the levels of cyclin-D1 Increased the level of E-cadherin and beclin Increase in CAT, and SOD activity Reduced the level of MDA
Karekar et al. [44]	Soy phosphatidylcholine	Andrographis paniculata	Hepatoprotective effect	Reduced the levels of hepatic enzymes Antioxidant activity
Chi et al. [45]	Soy phosphatidylcholine	Silybin		Reduced the levels of hepatic enzymes Reduced hepatocyte denaturation, inflammation, and fibrosis
Naik and Panda [46]	Soy phospholipids	Gingko biloba		Reduced the levels of hepatic enzymes Antioxidant activity
Shriram et al. [47]	Soy phosphatidylcholine	Silymarin		Reduced the levels of hepatic enzymes Antioxidant activity
El-Gazayerly et al. [48]	Soy phosphatidylcholine Egg yolk phosphatidylcholine	Silymarin		Reduced the levels of hepatic enzymes Antioxidant activity
Mahmoudabad et al. [49]	Soy phosphatidylcholine	Silymarin		Reduced liver enzymes and lipid peroxidation Antioxidant activity
Al-Kahtani et al. [50]	Soy phosphatidylcholine	Curcumin		Reduced liver enzymes and lipid peroxidation Antioxidant activity Downregulated caspase-3 expression Upregulated anti-apoptotic protein Bcl-2
Bui et al. [51]	Phosphatidylcholine	Curcumin		Reduced liver enzymes and lipid peroxidation Antioxidant activity
Jain et al. [52]	Soy phosphatidylcholine	Mangiferin		Antioxidant and hepatoprotective activity
Sharma et al. [53]	Soy phosphatidylcholine	Abutilon indicum and Piper longum		Reduced the levels of hepatic enzymes Antioxidant activity
Telange et al. [54]	Hydrogenated soy phosphatidylcholine	Apigenin		Reduced liver enzymes and lipid peroxidation Antioxidant activity
Mangrulkar et al. [55]	Hydrogenated soy phosphatidylcholine	Caffeic acid		Antihyperlipidemic and hepatoprotective activity Reduced liver enzyme levels and fat deposition
Mazumder et al. [39]	Hydrogenated soy phosphatidylcholine	Sinigrin	Wound healing	Wound healing properties on the HaCaT cell line
Varadkar and Gadgoli [56]	Phosphatidylcholine	Corcetin		Improved wound healing in rats Improved % of contraction and the breaking strength of the wounds Reduction in granulation tissue formation
Tafish et al. [57]	Soybean phosphatidylcholine Hydrogenated soy phosphatidylcholine	Carvacrol		Enhanced skin permeation Increase % of wound closure Reduced wound area Increased collagen deposition, tissue remodeling, and wound healing capacity
Lim et al. [58]	Soybean lecithin	Moringa oleifera		Improved wound closure
Jeeja et al. [59]	Soybean lecithin	Onosma echioides		Improved breaking and tensile strength Increased wound inhibition Reduced lipid peroxidation
Refai et al. [60]	L α-phosphatidylcholine	Spirulina platensis		Improved contraction rate Complete wound closure Improved skin appearance Reduced the expression of HMGB1, TLR-4, and NF-κB Increased NRF-2 and HO-1 levels Autophagy and anti-apoptotic properties Increased VEGF and collagen deposition
Rajamma et al. [61]	Soy phosphatidylcholine	Geophila repens	Alzheimer’s disease	Inhibition of cholinesterase
Habbu et al. [62]	L α-phosphatidylcholine	Bacopa monnieri		Reduced transfer latency Reduced the escape latency time and TSTQ Improved the mice’s response to shock Reduced the activity of acetylcholinesterase
Naik et al. [63]	N/A	Ginkgo biloba		Increased the levels of SOD and CAT Enhanced the activity of glutathione peroxidase and glutathione reductase
Naik et al. [64]	Soy phospholipids	Ginkgo biloba	Cognitive Impairment and Neuronal Damage	Reduced the phenobarbital-induced sleeping time Increased spontaneous motor activity Reduced the recovery time from convulsions Reduction in transfer latency Antiamnesic properties Reduced mobility time
Mancini et al. [65]	Phosphatidylcholine	Annona muricata L.		Enhanced translocation into hCMEC/D3 cells Inhibited MAO activity (antidepressant activity) Hydrogen peroxide scavenging activity
Ullah et al. [66]	Soy phosphatidylcholine	Curcumin		Reduced neuroinflammation
Sbrini et al. [67, 68]	N/A	Centella asiatica L.		Increased mRNA levels of Bdnf and its receptor TRKB Increased local proteins peEF2 Thr56 and OPHN-1 Improved performance in rats
Ahmad et al. [69]	Egg phosphatidylcholine	Rutin	Cerebral ischemia	Neuroprotective activity Increased level of GSH Decreased level of MDA Reduced the infarction area
Ahmad et al. [70]	Hydrogenated soy phosphatidylcholine	NMITLI118RT+		Increased GSH levels Decreased MDA levels Decreased neurological deficit score Reduced the infarction area
Hatamipour et al. [71]	Phosphatidylcholine Phosphatidylserine	Curcumin	Atherosclerosis	Significant reduction in atherosclerotic plaque area
Panda et al. [72]	Soy phospholipids	Ginkgo biloba	Myocardial infarction	Reduced the serum levels of AST, LDH, and CPK Antioxidant effect
Riva et al. [73]	N/A	Artichoke; Bergamot	Hypercholesterolemia	Reduced the levels of total cholesterol and LDL Increased level of HDL Combination reduced glycated hemoglobin levels
Singh et al. [74]	Soy lecithin	Lawson	Skin conditions (antifungal and anti-inflammatory)	Enhanced antifungal activity Enhanced skin permeation Reduction of edema
Djekic et al. [75]	N/A	Escin β-sitosterol	Skin conditions (anti-hyperalgesic)	Potent anti-hyperalgesic as compared to ibuprofen gel
Kalita et al. [76]	Phosphatidylcholine	Resveratrol	Skin conditions (anti-inflammatory)	Enhanced skin permeation Reduced edema
Baradaran et al. [77]	Soy phospholipids	Curcumin		Increased activity of CAT and SOD Reduced latency times
Maramaldi et al. [78]	Phosphatidylcholine	Quercetin	Skin conditions (soothing and anti-itch effect)	Reduced erythema Photoprotective effect Decrease in wheal diameter and itching Increased skin hydration levels Reduced TEWL values
Antiga et al. [79]	Phosphatidylcholine	Curcumin	Skin conditions (psoriasis)	Reduction in PASI values Decrease in the level of IL-22
Yu et al. [80]	Dipalmitoyl phosphatidylcholine	Naringenin	Respiratory tract diseases	Reduced edema and fluid exudation Reduced the levels of total proteins in BALF Increased the levels of SOD Inhibited the expression of COX-1 and ICAM-1
Singh et al. [81]	Soya Lecithin	Gingerol		Antioxidant and scavenging properties Antimicrobial activity against S. aureus and E. coli Reduced RBC membrane lysis and albumin denaturation
Abd El-Fattah et al. [82]	Phosphatidylcholine	Quercetin	Metabolic syndrome	Decrease in the levels of TNF-α and MDA Increased levels of GSH Reduced the body weight of rats Reduced the levels of bone biomarkers (ACP and ALP) Increased the levels of calcium and phosphorus in bones Decreased levels of TG, TC, LDL-C, and VLDL-C Increased the level of HDL-C Decreased blood glucose level
Kim et al. [83]	Phosphatidylcholine Egg phospholipid	Chrysin		Enhanced glucose uptake effect in C2C12 cell lines Upregulation of PPAR γ and GLUT4
Poruba et al. [84]	N/A	Silymarin		Reduced the levels of TG and (TC) Increased the level of HDL-C Increased levels of ABCG5 and ABCG8 Increased levels of CYP7A1 and CYP4A
Palachai et al. [85]	Phosphatidylcholine	Mulberry and ginger		Reduced body weight Decreased levels of TG, TC, and LDL-C Increased the level of HDL-C Reduce insulin resistance ACE gene expression Reduction in MDA levels Increased levels of SOD, CAT, and GSH Reduced levels of HDAC3 Increased level of PPAR γ Reduction in IL-6 and TNF-α
Yu et al. [86]	Soybean phosphatidylcholine	Berberin		Reduction in fasting blood glucose levels Reduced the level of TGs

Table 1.

Summary of herbosome-based studies included.

5.1 Anticancer effects

Current cancer therapy strategies, such as chemotherapy, radiotherapy, immunotherapy, and surgery, have numerous limitations and can cause systemic adverse effects. Furthermore, complete remission and full recovery are not always attained in cancer patients. Various plant-based compounds have inherent anticancer activity through various mechanisms, including antioxidant effects, interference with signaling pathways, and inhibition of chemoresistance. Several studies have demonstrated the efficacy and safety of herbosomes as a form of anticancer therapy [87, 88].

5.1.1 Breast cancer

Murugesan and team [24] prepared an Aloe vera-based herbosome gel as an anticancer nanosystem against breast cancer. The herbosome gel showed enhanced concentration-dependent cytotoxicity on the MCF-7 cell line. The mechanism by which the system induces the anticancer effect is through Aloe vera’s antioxidant properties. The cytotoxic effects of polyphenolic compounds from Moringa oleifera leaves were explored against breast cancer [25]. Moringa oleifera herbosomes (Mop) showed enhanced dose-dependent cytotoxicity on the 4 T1 breast cancer cell line as compared to doxorubicin. Mop was found to induce apoptosis in 4 T1 cell lines and thus have an antiproliferative effect on cancer cells. A novel scorpion venom-decorated phytosomes encapsulating quercetin (QRT-PHM-SV) were developed by Alhakamy et al. [26] QRT-PHM-SV exhibited significantly high cytotoxicity against the MCF-7 cell line as compared to the free drug. The nanosystem could induce apoptosis and necrotic cell death more significantly than the free drug as demonstrated by increased levels of caspase-9, Bax, and p53, while levels of Bcl2 decreased. Furthermore, the level of TNF-α was significantly increased after treatment with QRT-PHM-SV, while the level of NF-κB decreased confirming the induction of apoptosis. Sabzichi and colleagues [27] investigated the combination of luteolin-loaded phytosomes (Nano-lut) with doxorubicin in the MDA-MB231 cell line. Nano-lut in combination with doxorubicin had enhanced cytotoxicity against MDA-MB231 as compared to doxorubicin alone. Nano-lut was shown to inhibit the Nrf2 signaling pathway and its downstream genes HO1 and MDR1 resulting in enhanced sensitivity of MDA-MB231 to doxorubicin. The anticancer effect of self-assembled fisetin phytosomes (FIS-PHY) against the MDA-MB-231 cell line was studied by Talaat et al. [28]. The FIS-PHY formulation exhibited increased cytotoxicity with a lower IC50 value than the free drug. The phytosome could induce apoptosis and necrosis in a time-dependent manner. FIS-PHY was shown to inhibit the activity of TGF-β and MMP-9 by interfering with NF-κB and ERK1/2 signaling pathways. Furthermore, the phytosome system may enhance E-cadherin expression levels, leading to the inhibition of tumor progression. A pegylated, hyaluronic acid-CD44 targeting genistein phytosome (G-PHA) was formulated by Komeil et al. [29]. G-PHA demonstrated enhanced deposition in mammary glands and a significant reduction in tumor size compared to the free drug. Levels of CEA and CA15.3 biomarkers significantly decreased after treatment with G-PHA. Taxifolin phytosomes (PC3) were developed, and their antioxidant and cytotoxic effects on the MCF-7 cell line were studied [30]. PC3 exhibited an antioxidant effect that was dependent on its concentration, as demonstrated by its ability to scavenge radicals such as H₂O₂, NO, and DHHP. Cell viability studies showed a concentration-dependent cytotoxicity of PC3 against the MCF-7 cell line. Hashemzehi and team [31] investigated the antitumor effects of curcumin-loaded phytosomes (CUR-PHY) both independently and in conjunction with fluorouracil (FU). CUR-PHY exhibited concentration-dependent cytotoxicity against MCF-7 cell lines, as well as a reduction in cell invasion. The antitumor activity of CUR-PHY was found to be associated with increased expression of E-cadherin and MMP-9, which was further enhanced after combination with FU. The combination showed increased cytotoxicity in vivo compared to each treatment alone. Moreover, the antioxidant activity of CUR-PHY + FU was superior to that of either treatment alone, as demonstrated by the reduced levels of MDA and thiol, while catalase activity was significantly enhanced. The antitumor activity of curcumin is attributed to the activation of the AMPK signaling pathway, which in turn inhibits mTOR and Wnt/β-catenin signaling, resulting in cell cycle arrest through the inhibition of cyclin D1 (CD1). A multi-reservoir nanosystem comprising casein micelles incorporated into resveratrol phytosomes (PC-CAS MCs) was developed by El-Far et al. [32] PC-CAS micelles demonstrated significant cytotoxicity on the MCF-7 cell line compared to casein/resveratrol micelles and free drugs. Furthermore, the phagosomal-nano micelle system was able to decrease tumor weight and volume in vivo. The levels of tumor biomarkers, including aromatase, VEGF, NF-κB, and CD1, showed significant reductions following treatment with PC-CAS MCs. The phytosomal nanomicelles were found to induce apoptosis and necrosis, as evidenced by elevated levels of caspase-3 and % necrosis in histopathological analysis.

5.1.2 Lung cancer

Xu et al. [33] developed a diosgenin derivative-based herbosome (P2P) as an anticancer nanosystem for treating lung cancer. P2P exhibited cytotoxicity against A549 and PC9 cell lines that was both time- and dose-dependent, in comparison to the free drug. P2P demonstrated anticancer activity by inducing apoptosis through G0/G1 cell cycle arrest.

5.1.3 Prostate cancer

A new nanoplatform, CUR-PL-SV was formulated by conjugating scorpion venom with curcumin phytosomes by to combat prostate cancer effectively Al-Rabia et al. [34]. CUR-PL-SV exhibited increased cytotoxicity against the PC3 cell line compared to curcumin and scorpion venom-conjugated phytosomes. CUR-PL-SV cytotoxicity was demonstrated through the induction of apoptosis and necrosis, as evidenced by increased levels of Bax, p53, and caspase-3, while levels of Bcl-2, NF-κB, and TNF-α were significantly reduced. Furthermore, the nanophytosome disrupted the mitochondrial membrane potential, leading to further induction of apoptosis. A Phase II clinical trial conducted by Pastorelli and colleagues [35] investigated the antitumor activity of curcumin phytosomes as a combination therapy with gemcitabine in prostate cancer. Complementary administration of curcumin phytosomes increased the efficacy of gemcitabine, as evidenced by an increase in disease control rate (DCR) and overall survival rate (OS). Furthermore, there was a decrease in hematological and neurotoxic adverse events associated with the gemcitabine-curcumin phytosome combination. Analysis of tumor markers, such as IL-6, sCD40L, and CRP, showed reduced levels after treatment with complementary therapy, highlighting the role of curcumin in reducing inflammation and inhibiting tumor progression and metastasis.

5.1.4 Cervical cancer

Li and team [36] developed folic acid-decorated pegylated mitomycin C phytosomes (FA-PEG-MMC) as a targeted anticancer nanoplatform for cervical cancer. The specific nanosystem exhibited enhanced cytotoxicity on HeLa cell lines in a concentration and time-dependent manner compared to non-targeted NPs. In vivo studies demonstrated preferential accumulation in tumor tissue and enhanced antitumor activity, as evidenced by a reduction in tumor weight and volume.

5.1.5 Liver cancer

The antitumor activity of oral medium-chain (GP) and long-chain (GPL) phosphatidylcholine, genistein-loaded phytosomes was investigated by Komeil et al. [37] against hepatocellular carcinoma (HCC). GP demonstrated a time-dependent increase in cellular uptake compared to the genistein solution and GPL. The cytotoxicity in HepG2 cells demonstrated increased toxicity of genistein-loaded phytosomes compared to free genistein. The in vivo antitumor activity of GP and GPL was found to be enhanced through the induction of apoptosis, as indicated by increased levels of AIF, caspase-3, and caspase-8. Furthermore, genistein-loaded phytosomes reduced the levels of VEGF and MMP-9, which are crucial factors in cancer advancement and metastasis. In another study, the antitumor impact of curcumin phytosomes on hepatitis B virus-induced HCC was investigated [38]. Phytosomal curcumin NPs showed enhanced time-dependent cytotoxicity against the Huh-7 cell line. The cytotoxic effect was achieved through the activation of PPARγ and the inhibition of mTOR and NF-κB signaling pathways. The in vivo study demonstrated that phytosomal curcumin significantly reduced tumor volume and also decreased lipid and leukocyte accumulation compared to free curcumin.

5.1.6 Skin cancer

Sinigrin phytosomes (sin-phy) were developed by Mazumder et al. [39] to study their antitumor activity against skin cancer. Sin-phy exhibited concentration-dependent cytotoxicity against the A375 cell line but not the HaCat cell line.

5.1.7 Brain tumors

Mukherjee and colleagues [40, 41] investigated the anticancer effects of phytosomal curcumin (CCP) in glioblastoma. CCP significantly improved the survival of mice compared to the control and reduced the number of CD68 high GBM tumor cells. CCP significantly increased the level of iNOS and reduced the level of ARG1, as demonstrated by histopathology and flow cytometry analysis. In addition, CCP enhanced the activation of NF-κB to p65 NF-kB and the activation of STAT1 to its phosphorylated form (P-STAT1). CCP induces polarization of M2-TAMs into the M1 phenotype, as evidenced by a reduction in the level of IL-10 and an increase in the level of IL-12. Finally, curcumin phytosomes induced the expression of monocyte chemotactic protein-1 (MCP-1), leading to the recruitment of M1-TAMs and activated NK cells.

5.1.8 Colorectal cancer

Silibinin phytosomes’ (SP) anticancer activity against colorectal cancer was investigated in an HT29 xenograft model [42]. In comparison to free silibinin, SP exhibited enhanced antitumor activity, as demonstrated by a decrease in tumor weight and volume. The mechanism by which silibinin produced its antitumor activity was found to be through its antiproliferative and pro-apoptotic effect demonstrated by a reduction in PCNA+ and CD-1+ cells. Furthermore, silibinin inhibited ERK1/2 and Akt signaling which play a role in tumor progression. Furthermore, it inhibited angiogenesis by reducing the expression of VEGF, COX, iNOS, and HIF-1α in tumor cells. Marjaneh et al. [43] investigated the antitumor activity of curcumin phytosomes against colitis-induced colorectal cancer. Phytosomal curcumin showed a concentration and time-dependent cytotoxicity on the CT26 cell line. The tumor spheroids model showed the ability of curcumin phytosomes to induce cell death as compared to control. The preparation also reduced cell invasion and reduced the levels of cyclin-D1 while increasing the level of E-cadherin and beclin. Curcumin phytosomes induced cell cycle arrest as demonstrated by increased cell population in the G0/G1 phase as compared to control. In vivo studies showed that the nanophytosomes significantly reduced tumor numbers, area, and disease activity. The antioxidant effects of curcumin phytosomes were demonstrated through an increase in CAT and SOD activity, while the level of MDA was reduced significantly. Furthermore, the nanophytosome inhibited the Wnt/β-catenin signaling pathway as shown by reduced CD-1 and p-Gsk-3a/b expression levels.

5.2 Hepatoprotective effects

Herbal remedies have been long used in the management of liver diseases such as hepatitis, fatty liver, and acute liver injury. Medicinal plants contain a plethora of phytoconstituents that possess inherent hepatoprotective activities [89]. Bioactive molecules are classified into phenols, flavonoids, monoterpenes, coumarins, alkaloids, and glycosides. There are several mechanisms by which phytoconstituents produce their hepatoprotective effects. Phytochemicals regulate gastrointestinal and liver functions, boost the immune system, scavenge free radicals, and reduce lipid peroxidation. Moreover, they suppress the activity of cytochrome P450 enzymes which in some cases are responsible for the conversion of drugs/compounds into toxic products. Furthermore, phytochemicals shield the structure of the mitochondrial membrane and augment the activity of ATPase enzymes. Incorporation of phytochemicals into phospholipids to form phytosomes enhanced their hepatoprotective effects as phosphatidylcholine which is the most commonly used phospholipid and has inherent hepatoprotective properties. Phosphatidylcholine enhances the activity of collagenase enzyme which prevents liver fibrosis. The most commonly used phytochemicals are silymarin, eugenol, silybin, piperidine, and caffeine, among others.

Karekar and team [44] developed a nanophytosome incorporating Andrographis paniculata extract (APP) and studied its hepatoprotective activity. APP successfully reduced the levels of hepatic enzymes (ALT, AST, ALP, and total bilirubin). Serum biochemistry showed that APP could reduce the levels of gamma-glutamyl transferase (GGT) and lactate dehydrogenase (LDH) which are biomarkers of liver damage. APP could significantly enhance the levels of superoxide dismutase (SOD) and glutathione (GSH), while decreasing the level of malondialdehyde (MDA) highlighting the antioxidant activity of the nanophytosome. Silybin-phospholipid complex loaded nanosuspension (SPC-NPs) was formulated by Chi et al. [45] as a hepatoprotective nanosystem. SPC-NPs could significantly reduce the levels of ALT, AST, and AKP as compared to free silybin. Histopathological analysis showed the ability of SPC-NPs to reduce hepatocyte denaturation, inflammation, and fibrosis. Gingko biloba phytosomes (GBP) were investigated by Naik and Panda [46] as hepatoprotective nanosystems in carbon tetrachloride-induced (CCl4) liver injury. GBP was shown to reduce hepatic enzyme levels (ALT, AST, and ALP) while increasing the levels of albumin and total protein. Analysis of the antioxidant effect of GBP which is a measure of its hepatoprotective activity showed the ability of the nanosystem to enhance the activity of catalase (CAT), SOD, and GSH in addition to reducing lipid peroxidation demonstrated by reduced levels of TBARS. Shriram et al. [47] and El-Gazayerly et al. [48] demonstrated the hepatoprotective ability of silymarin phytosomes in CCl4-induced liver damage. Silymarin phytosomes reduced the levels of hepatic enzymes and enhanced the levels of antioxidant enzymes. Furthermore, the nanophytosomes could reduce lipid peroxidation evidenced by reduced levels of MDA. Another study by Mahmoudabad and colleagues [49] also investigated silymarin phytosomes hepatoprotective activity in ethanol-induced hepatotoxicity which demonstrated an enhanced ability to reduce liver enzymes and lipid peroxidation while enhancing the activity of antioxidant enzymes. Curcumin phytosomes’ (CP) ability to reverse aluminum chloride (AlCl3)-induced hepatotoxicity was investigated by Al-Kahtani et al. [50]. CP successfully reversed hepatotoxicity shown by reduced levels of hepatic and antioxidant enzymes. The ability of CP to reduce oxidative stress was demonstrated by decreased levels of nitric oxide (NO) and lipid peroxidase (LPO) expression. On the other hand, CP enhanced the activity of SOD and GSH. Immunohistochemistry assessment showed that treatment with CP downregulated caspase-3 expression while upregulated the anti-apoptotic protein Bcl-2. A similar approach has been developed by Tung et al. [51] proving the hepatoprotective ability of curcumin phytosomes in paracetamol-induced liver toxicity. Mangiferin phytosomal preparation was developed by Jain et al. [52] and demonstrated antioxidant and hepatoprotection in ethanol-induced liver damage. A combination of ethanolic extracts of Abutilon indicum and Piper longum was incorporated in a nanophytosome [53]. The combination nanophytosome significantly reduced liver enzymes (ALT, AST, and APL) and bilirubin levels (total and direct) as compared to combined ethanolic extracts. The potential of apigenin phytosome to produce antioxidant effects and thus reverse liver damage was studied by Telange et al. [54]. The nanophytosome enhanced antioxidant enzymes levels (SOD, CAT, and GSH) and reduced lipid peroxidation (reduced MDA levels). Caffeic acid phytosome (CA-PC) formulated by Mangrulkar and team [55] showed its potential use as an antihyperlipidemic and hepatoprotective nanosystem in non-alcoholic fatty liver disease (NAFLD). CA-PC reduced the levels of total cholesterol, low-density lipoprotein (LPL), triglycerides, very low-density lipoprotein (VLDL), and enhanced levels of high-density lipoprotein (HDL). Furthermore, the nanophytosome reduced liver enzyme levels and fat deposition demonstrated by histopathological analysis.

5.3 Wound healing properties

Wound healing is a complex and dynamic process that involves multiple sequential steps, including homeostasis, inflammation, proliferation, and remodeling [90]. In acute wounds, the healing process progresses smoothly until the wound is completely resolved. On the other hand, chronic wounds result from an impaired wound healing process, leading to complications such as fibrosis and non-healing ulcers. The use of herbal medicine in wound healing has emerged as a promising strategy because of the pleiotropic effects of phytochemicals. Phytochemicals possess antioxidant, anti-inflammatory, angiogenic, and cell synthesis-modulating properties. Several studies provide evidence of the wound-healing properties of herbal medicine. Incorporating phytochemicals into phytosomes has been shown to enhance their wound-healing properties by improving absorption and bioavailability at the wound site.

Sinigrin phytosomes demonstrated enhanced wound healing properties on the HaCaT cell line compared to free sinigrin [39]. Varadkar and Gadgoli [56] investigated the wound healing properties of corcetin phytosomes (F2) gel preparation. F2 showed improved wound healing in rats demonstrated by an increase of the level of hydroxyproline. Furthermore, excision and incision wounds showed that F2 improved both the percentage of contraction and the breaking strength of the wounds. Histopathological analysis revealed a significant reduction in granulation tissue formation, confirming the wound healing properties of the nanophytosome. Carvacrol-loaded phytosomes (CAR-PHY) were developed by Tafish and team [57] to investigate their potential for wound healing activity. CAR-PHY demonstrated enhanced permeation across the skin with sustained release, indicating that the skin can act as a reservoir for the phytosomal preparation. In vivo wound healing showed the ability of CAR-PHY hydrogel to increase the percentage of wound closure and reduce wound area. Histopathological examination revealed increased collagen deposition, tissue remodeling, and wound healing capacity. Moringa oleifera extract phytosomes (MOPCT) were developed as a potential platform for wound dressing [58]. MOPCT showed rapid and improved wound closure in the NHDF cell line compared to the control and free MO extract. Phytosomal gel loaded with Onosma echioides extract was evaluated for its wound healing effects by Jeeja et al. [59]. The phytosomal gel exhibited improved breaking and tensile strength compared to the control group. Moreover, the percentage of wound inhibition was significantly higher in the group treated with the phytosomal gel compared to control. The analysis of hydroxyproline and collagen deposition demonstrated the enhanced effect of the gel on wound healing. Furthermore, the gel reduced lipid peroxidation, as demonstrated by an increased level of catalase, while the level of MDA was reduced. Refai and colleagues [60] investigated an intriguing nanophytosomal gel containing Spirulina platensis extract (SPNP-gel). The SPNP-gel markedly improved contraction rate compared to the control and standard treatment demonstrated by complete wound closure and improved skin appearance. The analysis of inflammatory markers revealed that SPNP-gel exhibited anti-inflammatory properties by reducing the expression of HMGB1, TLR-4, and NF-κB. Furthermore, SPNP-gel increased NRF-2 and HO-1 levels, demonstrating its antioxidant capability. Moreover, histopathological analysis of TNF-α demonstrated enhanced reduction after treatment with the gel, further confirming its anti-inflammatory effects. SPNP-gel possess autophagy and anti-apoptotic properties, as evidenced by increased levels of LC3BII/I and Beclin-1 and reduced levels of caspase-3 and AIF. Furthermore, the levels of VEGF and collagen deposition markedly increased after treatment with the phytosomal gel, demonstrating its ability to promote wound healing through several mechanisms.

5.4 Nervous system conditions

5.4.1 Alzheimer’s disease

Alzheimer’s disease (AD) is a progressive neurodegenerative disease that accounts for 60% of all cases of dementia worldwide in people aged 65 years and older [91]. Alzheimer’s disease (AD) is associated with memory loss and cognitive deficits, including aphasia, agnosia, and apraxia. The exact cause of the disease is still unclear. However, the deposition of β-amyloid and τ-proteins, loss of synapses, and cholinergic neuron apoptosis appear to be the most common factors. Pharmacological treatment of Alzheimer’s disease mainly includes acetylcholinesterase inhibitors (AChE) and N-methyl-D-aspartate (NMDA) receptor antagonists. However, pharmacological treatment is not entirely effective and is often accompanied by several adverse effects. The efficacy of phytoconstituents like huperzine-A, ginkgolides, and asiaticosides in treating AD has been investigated in a number of studies proving their efficacy. Yet, their effectiveness is hindered by their low solubility and bioavailability, as well as their ability to pass through the blood-brain barrier (BBB). Therefore, incorporating phytochemicals into phytosomes offers a way to enhance their therapeutic effects.

Rajamma et al. [61] studied the therapeutic effects of Geophila repens phytosomal gel (MEGR-PG) for intranasal delivery in Alzheimer’s disease (AD). MEGR-PG demonstrated good intranasal permeation, which was further enhanced by the addition of 1% transcutol as a permeation enhancer. The in vitro cholinesterase inhibition assay demonstrated that MEGR-PG has the ability to inhibit cholinesterase compared to the control. Bacopa monnieri-loaded phytosomes (BPC) were studied for their antiamnesic effects in AD [62]. BPC could reduce transfer latency in the elevated plus maze test compared to control. In addition, in the Morris water maze test, BPC significantly reduced the escape latency time (ELT) and TSTQ as compared to the control groups. BPC also improved the mice’s response to shock (step-through latency time) and significantly reduced the activity of acetylcholinesterase. Ginkgo biloba phytosomes were formulated and assessed for their antioxidant effects to enhance cognitive function in AD [63]. The nanophytosomes significantly increased the levels of superoxide dismutase (SOD) and (CAT) enzymes in rat brains compared to the free Ginkgo biloba and control groups. Furthermore, the phytosomal system enhanced the activity of both glutathione peroxidase and glutathione reductase, confirming the antioxidant properties of Ginkgo biloba phytosomes.

5.4.2 Cognitive impairment and neuronal damage

A study by Naik and team [64] assessed the central nervous system (CNS) activity of Ginkgo biloba (GB) phytosomes in Wistar rats. The phytosomal preparation reduced the phenobarbital-induced sleeping time in a concentration-dependent manner compared to the control. In addition, the phytosomes increased spontaneous motor activity (SMA) for up to 60 minutes at concentrations of 50 and 100 mg/kg. GB phytosomes did not alter the duration of convulsions, but they reduced the recovery time compared to control. There was a significant reduction in transfer latency after treatment with GB phytosomes in the elevated plus maze. On the other hand, in the scopolamine-induced amnesia test, the phytosomes prolonged the transfer latency time, demonstrating the preparation’s antiamnesic properties. Finally, the phytosomes reduced the rats’ mobility time and prolonged their swimming time in the forced swimming test. The antidepressant activity of mApoE-decorated phytosomes loaded with Annona muricata L. extract (mApoE-P-AE) was investigated by Mancini et al. [65]. mApoE-P-AE demonstrated enhanced translocation into hCMEC/D3 cells, indicating the system’s ability to cross the BBB. The phytosomes significantly inhibited MAO activity compared to the free extract, demonstrating its antidepressant activity. Moreover, mApoE-P-AE exhibited strong hydrogen peroxide scavenging activity compared to the free extract. The anti-inflammatory effect of curcumin phytosomes (MC) on chronic glial activation was studied by Ullah and colleagues [66]. MC demonstrated the ability to reduce neuroinflammation, as evidenced by a decrease in Iba-1+ microglia, TSPO+ microglial cells/macrophages, and GFAP+ astrocytes in the hippocampus and cerebellum of GFAP-IL6 mice. Two studies by Sbrini et al. [67, 68] investigated the potential of Centella asiatica L. phytosomes, with or without Curcuma longa L., to enhance cognitive function by promoting Bdnf expression. Phytosomes increased the mRNA levels of Bdnf and its receptor TRKB. Moreover, the levels of local proteins peEF2 Thr56 and OPHN-1 increased significantly. The novel object recognition test demonstrated improved performance in rats following chronic treatment with phytosomes.

5.4.3 Cerebral ischemia

Ahmad and team [69] investigated the neuroprotective effect of rutin phytosomes (RU-PLC) in ischemic stroke. RU-PLC exhibited high and preferential accumulation in the brain compared to free rutin. The neuroprotective activity of the phytosomes was demonstrated by an increased level of GSH, while the level of MDA was significantly decreased. Furthermore, RU-PLC reduced the infarction area at a dose less than half of free rutin, demonstrating its neuroprotective role in ischemic stroke. Another study by Ahmad et al. [70] evaluated the neuroprotective activity of NMITLI118RT+ loaded phytosomes (NIMPLC). NIMPLC significantly increased GSH levels and decreased MDA levels. The neurological deficit score significantly decreased after treatment with NIMPLC. Moreover, the infarction area was also reduced compared to the free extract at 1 and 6 hours post-injury.

5.5 Cardiovascular disease

Cardiovascular disease (CVD) is a widespread health issue that causes the death of one-third of the global population. CVD encompasses a plethora of conditions, including atherosclerosis, coronary artery disease, diabetes mellitus, and cerebrovascular diseases. Although pharmacological treatment has advanced, it has improved symptoms and survival but does not cure the disease. Herbal medicine has been utilized in the treatment of various cardiovascular diseases, with many therapies derived from plant sources. For example, digoxin is derived from Digitalis purpurea, aspirin from Salix alba, and lovastatin from Monascus purpureus, among others. Currently, researchers are exploring the use of traditional herbs such as ginseng, Ginkgo biloba, and Gynostemma pentaphyllum incorporated in phytosomes.

5.5.1 Atherosclerosis

Hatamipour et al. [71] investigated the anti-atherosclerotic effects of curcumin phytosomes by formulating curcumin-phosphatidylcholine (curcumin-PC) and curcumin-phosphatidylserine phytosomes (curcumin-PS). Curcumin-PC/PS had no significant effect on lipid profile (TC, TG, LDL, VLDL, and HDL) or CRP levels as compared to the control. However, curcumin-PS (100 mg/kg) demonstrated a significant reduction in atherosclerotic plaque area compared to curcumin-PC and the control group.

5.5.2 Myocardial infarction

The study by Panda and colleagues [72] studied the cardioprotective effects of Ginkgo biloba phytosomes (GBP) in rats with isoproterenol-induced myocardial necrosis. GBP could reduce the serum levels of AST, LDH, and CPK in a concentration-dependent manner compared to the control. The antioxidant effect of GBP was demonstrated by a decrease in the level of MDA, yet an increase in GSH, CAT, GPx, and GR levels.

5.5.3 Hypercholesterolemia

An RCT investigating the efficacy of a combination of Artichoke Phytosomes and Bergamot Phytosomes in patients with mild hypercholesterolemia was conducted by Riva et al. [73]. Supplementation with both phytosomal preparations reduced the levels of total cholesterol and LDL, while the level of HDL was significantly increased. Furthermore, the combination reduced glycated hemoglobin levels compared to the control group, but there was no significant change in glycemia, insulin resistance, or triglyceride levels.

5.6 Inflammatory skin conditions

Singh et al. [74] evaluated the antifungal and anti-inflammatory properties of Lawson phytosomes. The phytosomes exhibited enhanced antifungal activity compared to the plant drug and standardized ketoconazole. The phytosomes exhibited enhanced skin permeation abilities compared to the free drug. The nanophytosome’s anti-inflammatory properties were demonstrated by the significant reduction of edema in rat paws compared to the free plant drug. An escin β-sitosterol phytosome (ES) investigated the anti-hyperalgesic effects of topical hydrogel preparation [75]. ES exerted a more potent anti-hyperalgesic effect in vivo compared to the control and ibuprofen gel. Resveratrol phytosomes embedded in a polymeric patch (RSVP) were developed, and their anti-inflammatory properties were investigated [76]. RSVP polymeric patch showed enhanced skin permeation compared to the free drug. The in vivo anti-inflammatory effect of RSVP was investigated using carrageenan-induced paw swelling, and the study showed that the phytosomes significantly reduced edema compared to the control group. A study by Baradaran et al. [77] examined the anti-inflammatory impact of curcumin phytosomes on carrageenan-induced inflammation. Curcumin phytosomes exhibited antioxidant properties, as evidenced by the increased activity of CAT and SOD enzymes compared to the control. Behavioral responses of the mice were evaluated using the tail pinch and hot plate tests, which revealed that mice treated with curcumin phytosomes exhibited reduced latency times compared to the control group. A single-blind study was conducted by Maramaldi and team [78] to evaluate the soothing and anti-itch effects of phytosomal quercetin on healthy volunteers. Quercetin phytosomes reduced erythema caused by UV irradiation, demonstrating a photoprotective effect. Additionally, they led to a significant decrease in wheal diameter and itching after a histamine prick test. Furthermore, quercetin phytosomes increased skin hydration levels and reduced TEWL values compared to the control. Antiga et al. [79] conducted a phase III, double-blind, placebo-controlled randomized controlled trial (RCT) to investigate the effectiveness of oral curcumin as a supplementary treatment to topical methylprednisolone aceponate 0.1% ointment in psoriasis. Curcumin phytosomes, when combined with topical methylprednisolone, resulted in a reduction in PASI values compared to methylprednisolone alone. The combination therapy resulted in a decrease in the level of IL-22 compared to the placebo, but it had no effect on IL-17.

5.7 Respiratory tract diseases

5.7.1 Anti-inflammatory

Yu and team [80] investigated the use of naringenin-loaded phytosomes as a dry powder inhalation (NPDPI) treatment for acute lung injury. NPDPI significantly reduced edema and fluid exudation resulting from acute lung injury compared to the control group. The mechanism by which the preparation exerts its effect is through the inhibition of MAPK phosphorylation, leading to p38 MAPK. NPDPI reduced the levels of total proteins in bronchoalveolar lavage fluid (BALF) compared to the positive control. In addition, the phytosomal preparation significantly increased the levels of SOD compared to the positive control. RT-PCR analysis showed that NPDPI inhibited the expression of COX-1 and ICAM-1 compared to the positive control, demonstrating its anti-inflammatory effect.

5.7.2 Anti-microbial

Gingerol-loaded phytosomes complexed with chitosan (GLPC4) were investigated for their antimicrobial and anti-inflammatory properties in lung infections [81]. GLPC4 exhibited antioxidant and scavenging properties by inhibiting free radicals (DPPH and H₂O₂). The study demonstrated that GLPC4 exhibited greater antimicrobial activity against S. aureus and E. coli compared to free gingerol, effectively inhibiting bacterial growth. Furthermore, GLPC4 demonstrated anti-inflammatory properties by reducing RBC membrane lysis and albumin denaturation compared to free gingerol.

5.8 Metabolic syndrome

Abd El-Fattah et al. [82] investigated the antiestrogenic activity of quercetin-loaded phytosomes (QP) in ovariectomized rats (Ovx). The QP compound exhibited antioxidant properties, as evidenced by a decrease in the levels of TNF-α and MDA, while the levels of GSH increased compared to the Ovx group. The phytosomes also significantly reduced the body weight of rats in a concentration-dependent manner. QP reduced the levels of bone biomarkers (ACP and ALP) compared to the Ovx group and consequently increased the levels of calcium and phosphorus in bones. Furthermore, QP had a significant impact on the lipid profile by decreasing levels of TG, TC, LDL-C, and VLDL-C, while increasing the level of HDL-C. Furthermore, the phytosomal preparation significantly decreased blood glucose levels compared to the Ovx group. Chrysin-loaded phytosomes using soy phosphatidylcholine (CSP) and egg phospholipid (CEP) were developed to investigate their glucose uptake-promoting activity [83]. CEP demonstrated an enhanced glucose uptake effect in C2C12 cell lines compared to the control in a dose-dependent manner. The mechanism by which CEP produces its activity has been shown to be through the upregulation of peroxisome proliferator-activated receptor γ (PPAR γ) and glucose transporter type 4 (GLUT4). The study by Poruba and team [84] investigated the anti-hyperlipidemic activity of silymarin phytosomes (PS) in metabolic syndrome. PS reduced the levels of triglycerides (TG) and total cholesterol (TC) compared to the control, while the level of high-density lipoprotein (HDL) was significantly increased. The levels of ABCG5 and ABCG8 transporters involved in cholesterol metabolism were enhanced in the group treated with PS. In addition, the levels of CYP7A1 and CYP4A were significantly increased in the PS group compared to control. Another study investigated the effectiveness of phytosomes containing a combination of mulberry and ginger extracts (PMG) in treating metabolic syndrome [85]. PMG reduced body weight and levels of triglycerides (TG), total cholesterol (TC), and low-density lipoprotein cholesterol (LDL-c) compared to the positive control, while increasing the level of high-density lipoprotein cholesterol (HDL-c). In addition, the homeostasis model assessment of insulin resistance (HOMA-IR) indicated that PMG could reduce insulin resistance in a dose-dependent manner. The level of ACE gene expression was also reduced in the group treated with PMG compared to the positive control. Furthermore, PMG exhibited antioxidant effects as demonstrated by the reduction in MDA levels, while the levels of SOD, CAT, and GSH were significantly increased in a dose-dependent manner. The expression levels of HDAC3 were significantly reduced, while PPARγ was significantly increased in the group treated with PMG compared to the control group. Furthermore, the PMG group showed a dose-dependent reduction in the expression of inflammatory cytokines (IL-6 and TNF-α). Berberine phytosomes encapsulated within microparticles (microparticles@P-BER) were developed, and their anti-diabetic properties were studied [86]. Microparticles containing berberine (P-BER) exhibited anti-diabetic activity, as evidenced by a reduction in fasting blood glucose levels compared to free berberine and the positive control (db/db mice). However, there was no difference in insulin levels. Moreover, Microparticles@P-BER reduced the level of TGs in the liver significantly as compared to free berberine and positive control.

6. Future perspectives

Phytosomes have been demonstrated to be an effective drug delivery system for phytochemicals. However, there are still several limitations that hinder their clinical application and commercialization in the market. While phytosomes are complexes of naturally occurring phospholipids and herbal constituents, safety concerns still arise because of their nanoscale size. Some parameters, such as biocompatibility, bioaccumulation, metabolism, and excretion, should be carefully assessed before their release to the market [92, 93]. Another limitation is their biological effectiveness and safety, which is related to their ability to cross biological barriers and reach target cells. The complete pharmacokinetic and pharmacodynamic profile of phytosomes should be assessed in vivo and in clinical trials. The selection of the dosage form is a crucial aspect that should be taken into consideration during the manufacturing and scaling-up process. Post-marketing surveillance and quality control of commercial phytosomes are necessary to ensure their safety and efficacy. Currently, herbal medicine has gained tremendous popularity worldwide, and many people are shifting toward natural remedies instead of relying on chemical drugs. Several phytosomes are already on the market, including Siliphos^®, Ginkgoselect^®, Centevita^®, and Soyselect^®. Further optimization of phytosomal preparations will facilitate their seamless transition from the laboratory to clinical use.

7. Conclusion

Phytochemicals have recently garnered significant interest in the prevention and management of various disease conditions due to people’s increasing inclination toward natural products. Phytochemicals comprise a range of compounds, including flavonoids, tannins, glycosides, carotenoids, phytosterols, and saponins, among others. They exhibit various biological activities, including antioxidant, antimicrobial, anticancer, hepatoprotective, cardioprotective, anti-hyperlipidemic, anti-inflammatory, and neuroprotective actions. However, they suffer from poor solubility in water, low permeability, biotransformation, and the formation of insoluble complexes in the gastrointestinal tract (GIT), which results in low bioavailability and reduced efficacy. As a result, integrating these phytochemicals into delivery systems has become a critical factor in enhancing their therapeutic activity. Phytosomes are lipid-based nanoparticles formed by complexing phospholipids with phytochemicals. Phytosomes address the limitations of phytochemicals by enhancing their solubility, improving bioavailability, providing sustained and controlled release, enabling targeted delivery, reducing toxic effects, and enhancing stability. Numerous studies have investigated the biological activities of phytosomes and have demonstrated their superiority to free phytochemicals. Phytosomes have been studied in various ailments such as cancer, CNS diseases (including neurodegenerative diseases), liver diseases, skin conditions and wound healing, respiratory tract conditions, and metabolic syndromes. Several phytosomes are currently available in the market. However, further optimization of phytosomes is necessary to ensure safety and efficacy and to facilitate their clinical translation on a larger scale.