NOVEL DRUG DELIVERY SYSTEM WITH SPECIAL REFERENCE TO PHYTOSOMES FOR HERBAL

FORMULATION

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Introduction

In the past few decades, considerable attention has been focused on

the development of novel drug delivery system (NDDS) for herbal drugs. The

novel carriers should ideally fulfill two prerequisites. Firstly, it should deliver

the drug at a rate directed by the needs of the body, over the period of

treatment. Secondly, it should channel the active entity of herbal drug to the

site of action. Conventional dosage forms including prolonged-release dosage

forms are unable to meet none of these. In phyto-formulation research,

developing nano dosage forms (polymeric nanoparticles and nanocapsules,

liposomes, solid lipid nanoparticles, phytosomes and nanoemulsion etc.) have

a number of advantages for herbal drugs, including enhancement of solubility

and bioavailability, protection from toxicity, enhancement of pharmacological

activity, enhancement of stability, improving tissue macrophages distribution,

sustained delivery, protection from physical and chemical degradation etc.

Thus the nano sized novel drug delivery systems of herbal drugs have a

potential future for enhancing the activity and overcoming problems

associated with plant medicines.

Liposomes, which are biodegradable and essentially non-toxic

vehicles, can encapsulate both hydrophilic and hydrophobic materials

(Medina et al., 2004). Liposome based drug delivery systems offer the

potential to enhance the therapeutic index of anti-cancer agents, either by

increasing the drug concentration in tumor cells and/or by decreasing the

exposure in normal tissues exploiting enhanced permeability and retention

effect phenomenon and by utilizing targeting strategies (Sharma et al., 2006).

The main advantages of using liposomes include: i) the high biocompatibility,

ii) the easiness of preparation, iii) the chemical versatility that allows the

loading of hydrophilic, amphiphilic, and lipophilic compounds, and iv) the

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simple modulation of their pharmacokinetic properties by changing the

chemical composition of the bilayer components (Terreno et al., 2008).

Delivery of agents to the reticuloendothelial system (RES) is easily achieved,

since most conventional liposomes are trapped by the RES (Medina et al.,

2004). The application of novel approaches can also improve the efficacy of

herbal cosmetic formulations on the human body (Chanchal and Swarnlata,

2008). Similarly the other vesicular systems like nanoemulsion, ethosomes

and transferosomes are highly useful assemblies and find various advantages

in the delivery of herbal medicines; some of them are summarized in present

article.

The phytosome process has also been applied to many popular herbal

extracts including Ginkgo biloba, grape seed,hawthorn, milk thistle (Barzaghi

et al., 1990), green tea, and ginseng. The flavonoid and terpenoid

components of these herbal extracts lend themselves quite well for the direct

binding to phosphatidylcholine. Phytosome is produced by binding individual

components of herbal extracts to phosphatidyl choline, resulting in a dosage

form that is better absorbed and thus, produces better results than the

conventional herbal extracts. The results indicate that the absorption of silybin

from silybin phytosome is approximately seven times greater compared to the

absorption of silybin from regular milk thistle extract. Drugs can be embedded

or dissolved in nanoparticles and can also be adsorbed or coupled on the

surface (Yuan and Yi, 2003). Encapsulating drugs within NPs can improve the

solubility and pharmacokinetics of drugs, and, in some cases, enable further

clinical development of new chemical entities that have stalled because of

poor pharmacokinetic properties (Alexis et al., 2008). The major carrier

materials of nanoparticles are synthetic biodegradable high molecular polymer

and natural polymer. The former usually includes poly-α-cyanoacrylate alkyl

esters, polyvinyl alcohol, polylactic acid, and polylacticcoglycolic acid, etc. The

latter is usually divided into two classes: proteins (albumin, gelatin and

vegetable protein) and polysaccharides (cellulose, starch and its derivatives,

alginate, chitin and chitosan, etc. Xiao and Li, 2002). In this seminar, an

attempt has been made to touch upon different aspects related to the

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development of novel herbal formulations, including method of preparation,

type of active ingredient, entrapment efficiency, and applications etc.

Liposome

The liposomes are spherical particles that encapsulate a fraction of the

solvent, in which they freely diffuse (float) into their interior. They can have

one, several or multiple concentric membranes. Liposomes are constructed of

polar lipids which are characterized by having a lipophilic and hydrophilic

group on the same molecules. Upon interaction with water, polar lipids self-

assemble and form self-organized colloidal particles. Simple examples are

detergents, components form micelles, while polar lipids with bulkier

hydrophobic parts cannot associate into micelles with high curvature radii but

form bilayers which can self-close into liposomes or lipid vesicles. A cross-

section of a liposome (Fig. 1) depicts the hydrophilic heads of the amphiphile

orienting towards the water compartment while the lipophilic tails orient away

from the water towards the center of the vesicle, thus forming a bilayer.

Consequently, water soluble compounds are entrapped in the water

compartment and lipid soluble compounds aggregate in the lipid section.

Uniquely, liposomes can encapsulate both hydrophilic and lipophilic materials.

Liposomes usually formed from phospholipids, have been used to change the

pharmacokinetics profile of, not only drugs, but herbs, vitamins and enzymes.

A variety of herbal liposomal formulations has been studied which are

summarized in Table 1. Because of their unique properties liposomes are able

to enhance the performance of products by increasing ingredient solubility,

improving ingredient bioavailability, enhanced intracellular uptake and altered

pharmacokinetics and bio distribution and in vitro and in vivo stability.

Liposomes as a drug delivery system can improve the therapeutic activity and

safety of drugs, mainly by delivering them to their site of action and by

maintaining therapeutic drug levels for prolonged periods of time (Barragan-

Montero et al., 2005). Milk thistle (Silybum marianum) is one of the few herbal

drugs whose excellent pharmacological profile readily lends itself to proof of

clinical efficacy (Weiss and Fintelmann, 2000). Meanwhile, silymarin is poorly

absorbed (20–50%) from the gastrointestinal tract (Blumenthal et al., 2000)

that causes the effects of silybin, one of the main active flavonoids commonly

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found in the dried fruits of silymarin, to be greater after parenteral than oral

administration (Carini et al., 1992).

Fig.1. Cross-section of a liposome (Chanchal and Swarnlata, 2008).

Incorporation of silymarin into liposomal dosage form administered

buccaly can improve its bioavailability. In this connection to improve the

bioavailability of silymarin through its incorporation in a stable liposomal

buccal dosage form, using commercially available soybean lecithin. El-

Samaligy et al., (2006) prepared silymarin encapsulated hybrid liposomes

which shows successful preparation with efficient encapsulation of silymarin.

Mixing silymarin loaded hybrid liposomes with unloaded ones in a (1:1)

proportion was useful in prevention of aggregates which threaten liposomal

stability. M50 proved stability regarding encapsulation efficiency, turbidity

measurement and particle size analysis after 3 months of storage at 4 °C or at

ambient temperature. Refrigeration is recommended to achieve better

stability. The introduced hybrid liposomal silymarin formula for buccal

administration have the advantages of exerting a mucoadhesive effect

(Takeuchi et al., 2003) besides its deformability due to the presence of Tween

20 as edge activator allowing the medicated liposomes to squeeze through

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buccal mucosal cells. It was also shown to be safe upon contacting the rat

buccal mucosa.

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Table 1: Liposomal herbal formulation. (Ajazuddin & Saraf, 2010)

Formulations Active Applications of liposome Biological activity Method of % Route ofingredients formulations preparation Entrapment administration

efficiency

Quercetin liposomes Quercetin Reduced dose, enhance Antioxidant Reverse 60% Intranasal

penetration in blood brain Anticancer evaporation

barrier technique

Liposomes Silymarin Improve bioavailability Hepatoprotective Reverse 69.22± Buccal

encapsulated silymarin evaporation 0.6%

technique

Liposoma artemisia Artemisia Targeting of essential oils to Antiviral Film method and 60–74% In vitro

arborescens arborescens cells, enhance penetration sonication

essential oil into, cytoplasmatic barrier

Ampelopsin liposome Ampelopsin Increase efficiency Anticancer Film-ultrasound 62.30% In vitro

method

Paclitaxel liposome Paclitaxel High entrapment efficiency Anticancer Thin film 94% In vitro

and PH sensitive hydration method

Curcumin liposome Curcumin Long-circulating with high Anticancer Ethanol injection 88.27± In vitro

entrapment efficiency method 2.16%

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Garlicin liposome Garlicin Increase efficiency Lungs Reverse-phase 90.77 % –

evaporation

method

Flavonoids liposomes Quercetin Binding of flavonoids with Hb Hemoglobin Solvent – In vitro

and rutin is enhanced evaporation

Usnea acid liposome with Usnea acid Incrase solubility and Antimycobacterial Hydration of a thin 99.5% In vitro

β-CD localization with prolonged- lipid film method

release profile with sonication

Wogonin liposome Wogonin Sustained release effect Anticancer Film dispersion 81.20± In vivo

method 4.20%

Colchicine Liposome Colchicine Enhance skin accumulation, Antigout Rotary 66.3±2.2% Topical

prolong drug release and evaporation

improve site specificity sonication method

Catechins liposomes Catechins Increased permeation Antioxidant and Rotary 93.0±0.1 Transdermal

through skin chemopreventive evaporation

sonication method

Breviscapine liposomes Breviscapin Sustained delivery of Cardiovascular Double 87.9±3.1% Intramuscular

breviscapine diseases emulsification

process

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Nanoparticles

In recent year, the nanonization of herbal medicines has attracted

much attention;(Zhinan et al., 2003). Some of them are illustrated in Table 2.

Nanoparticles and nanoemulsions (Fig. 2) are colloidal systems with particles

varying in size from 10 nm to 1000 nm (Ratnam et al., 2006). Nanoparticle

systems with mean particle size well above the 100 nm standard have also

been reported in literature, including nanonized curcuminoids (Tiyaboonchai

et al. , 2007), paclitaxel ( Arica et al., 2006) and praziquantel (Mainardes et

al., 2005) which have a mean particle size of 450, 147.7, and even higher

than 200 nm, respectively. In addition, nanoparticles could also be defined as

being submicronic (b1 lm) colloidal systems (Brigger et al., 2002). The

nanospheres have a matrix type structure in which the active ingredient is

dispersed throughout (the particles), whereas the nanocapsules have a

polymeric membrane and an active ingredient core. Nanonization possesses

many advantages, such as increasing compound solubility, reducing

medicinal doses, and improving the absorbency of herbal medicines

compared with the respective crude drugs preparations.

Fig. 2. Cross-section of (a) nanoemulsion and (b) biopolymeric

nanoparticle (Chanchal and Swarnlata, 2008).

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Table 2: Nano structured herbal formulations. (Ajazuddin & Saraf, 2010)

Formulations Active Applications of Biological activityMethod of preparation

% Entrapment Route of

ingredients nanostructured formulations efficiency administration

Triptolide TriptolideEnhance the penetration of drugs through the Anti-inflammatory

Emulsification-ultrasound – Topical (skin)

nanoparticlestratum corneum by increased hydration

Nanoparticles of Cuscuta Flavonoids Improve water solubility, Hepatoprotective andNanosuspension method 90% Oral

chinensis and lignans antioxidant effects

Triptolide-loaded Triptolide Decreasing the toxicity Anti-inflammatoryEmulsification-ultrasound – Oral

solid lipid nanoparticle

Artemisinin nanocapsules Artemisinin Sustained drug release Anticancer

Self-assembly procedure 90–93% In vitro

Radix salvia R. salvia Improve the bioavailabilityCoronary heart diseases, angina

Spray-drying technique Upto 96.68% In vitro

miltiorrhiza nanoparticles miltiorrhiza

pectoris and myocardial infarction

Taxel-loaded nanoparticles Taxel Enhance the bioavailability and Anticancer

Emulsion solvent evaporation 99.44% –

sustained drug release method

Berberine-loaded Berberine Sustained drug release Anticancer Ionic gelation method 65.40±0.70% In vitro

nanoparticles

Silibini-loaded nanoparticles Silibini

High entrapment efficiency and stability Hepatoprotective

High pressure homogenization 95.64% –

Tetrandrine-loaded Tetrandrine Sustained drug release LungSelf-emulsification and solvent 84% In vitro

nanoparticles evaporating

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Glycyrrhizic acid-loaded Glycyrrhizic Improve the bioavailability Anti-inflammatory, Rotary-evaporated 91.76% –

nanoparticles acid antihypertensivefilmultrasonication method

Quercetin-loaded QuercetinIncrease antioxidant activity and release of the Antioxidant

Nanoprecipitation technique over 99% In vitro

nanoparticles drug 74 times higher

Breviscapine-loaded Breviscapine Prolong the half-life and decreaseCardiovascular and cerebrovascular

Spontaneous emulsification 93.1% Intra Venous

nanoparticles RES uptakesolvent diffusion technique

Zedoary turmeric oil ZedoaryIncrease the drug loading and stability of ZTO

Hepatoprotection Anticancer and High pressure 1.62 ± 0.15% –

nanocapsule turmeric oil anti-bacterialHomogenization method

Loading Capacity

Naringenin-loaded Naringenin Improved the release of NAR and HepatoprotectiveNanoprecipitation method – Oral

nanoparticles improved its solubility

Curcuminoids solid lipid CurcuminoidsProlonged-release of the curcuminoids Anticancer and antioxidant

Micro-emulsion technique 70% In vitro

nanoparticles

CPT-encapsulated CamptothecinProlonged blood circulation and high Anticancer Dialysis method N80% In vitro

nanoparticles accumulation in tumors

Ginkgo biloba nanoparticles Ginkgo biloba

Improving the cerebral blood flow and Brain function activation

High pressure homogenization – Oral

extract metabolism method

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Phytosome

Over the past century, phytochemical and phytopharmacological

sciences established the compositions, biological activities and health

promoting benefits of numerous plant products. Most of the biologically active

constituents of plants are polar or water soluble molecules. However, water

soluble phytoconstituents (like flavonoids, tannins, terpenoids, etc.) are poorly

absorbed either due to their large molecular size which cannot absorb by

passive diffusion, or due to their poor lipid solubility; severely limiting their

ability to pass across the lipid-rich biological membranes, resulting poor

bioavailability (Manach et al., 2004). It has often been observed that the

isolation and purification of the constituents of an extract may lead to a partial

or total loss of specific bio-activity for the purified constituent — the natural

constituent synergy becomes lost. Very often the chemical complexity of the

crude or partially purified extract seems to be essential for the bioavailability

of the active constituents. Extracts when taken orally some constituents may

be destroyed in the gastric environment. As standardized extracts are

established, poor bioavailability often limits their clinical utility due to above

said reasons. It has been observed that complexation with certain other

clinically useful nutrients substantially improves the bioavailability of such

extracts and their individual constituents. The nutrients so helpful for

enhancing the absorption are the phospholipids. Phytosome is a patented

technology developed by a leading manufacturer of drugs and nutraceuticals,

to incorporate standardized plant extracts or water soluble phytoconstituents

into phospholipids(phosphatidylcholine) to produce lipid compatible molecular

complexes, called as phytosomes and so vastly improve their absorption and

bioavailability(Bombardelli et al., 1989). Phospholipids are complex molecules

that are used in all known life forms to make cell membranes. In humans and

other higher animals the phospholipids are also employed as natural digestive

aids and as carriers for both fat-miscible and water miscible nutrients. They

are miscible both in water and in lipid environments, and are well absorbed

orally. Phytosomes are more bioavailable as compared to conventional herbal

extracts owing to their enhanced capacity to cross the lipoidal biomembrane

and finally reaching the systemic circulation. Phytosome has been an

emerging trend in delivery of herbal drugs and nutraceuticals.

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Table 3: Phytosomal herbal formulations. (Ajazuddin & Saraf, 2010)Formulations Active Applications of phytosomal formulations Biological activity Method of Dose Route of

ingredients preparation administration

Ginkgo biloba Flavonoids Flavonoids of GBP stabilize the ROS Cardio-protective, Phospholipids 100 mg Subcutaneous

phytosomes antioxidant complexation and

activity 200 mg/

kg

Ginkgoselect Flavonoids Inhibits lipid peroxidation (LPO), Hepatoprotective, Phospholipids 25 and Oral

phytosome stabilize the ROS antioxidant complexation 50 mg/

kg

Silybin Flavonoids Absorption of silybin phytosome Hepatoprotective, Silybin- 120 mg Oral

phytosome from silybin is approximately antioxidant for phospholipid

seven times greater liver and skin complexation

Ginseng Ginsenosides Increase absorption Nutraceutical, Phospholipids 150 mg Oral

phytosome immunomodulator complexation

Green tea Epigallocatechin Increase absorption Nutraceutical, Phospholipids 50– Oral

phytosome systemic complexation 100 mg

antioxidant, anti-

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cancer

Grape seed Procyanidins The blood TRAP nTotal Radical-trapping Systemic Phospholipids 50– Oral

phytosome Antioxidant Parameter) were significantly antioxidant, complexation 100 mg

elevated over the control cardio-protective

Hawthorn Flavonoids Increase therapeutic efficacy Cardio-protective Phospholipids 100 mg Oral

Phytosome and absorption and Complexation

antihypertensive

Quercetin Quercetin Exerted better therapeutic efficacy Antioxidant, Quercetin– – Oral

phytosome anticancer phospholipid

complexation

Curcumin Curcumin Increase antioxidant activity and Antioxidant, Curcumin– 360 mg/ Oral

phytosomes Increase bioavailability anticancer phospholipid kg

complexation

Naringenin Naringenin Prolonged duration of action Antioxidant Naringenin– 100 mg/ Oral

phytosomes activity phospholipid kg

complex

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Emulsions

Emulsion refers to a non-homogeneous dispersion system that is

composed of two kinds of liquids unable to dissolve each other, and one of

which disperse in the other one in a form of droplets. Generally, emulsion is

composed of oil phase, water phase, surfactant and sub-surfactant. Its

appearance is translucent to transparent liquid. Emulsion can be classified

into ordinary emulsion (0.1–100 μm), micro-emulsion (10–100 nm), sub-micro-

emulsion (100–600 nm), etc. (Table 4). Among them, themicro-emulsion is

also called nanoemulsions, and the sub-micro-emulsion is also called lipid

emulsion. As a drug delivery system, emulsion distributes in vivo in the

targeted manner due to its affinity to the lymph. In addition, the drug can be

sustained release in a long time because the drug is packaged in the inner

phase and kept off direct touch with the body and tissue fluid(Lu et al., 2005).

After the oily drugs or lipophilic drugs being made into O/W or O/W/O

emulsion, the oil droplets are phagocytosised by the macrophage and get a

high concentration in the liver, spleen, and kidney in which the amount of the

dissolved drug is very large.Whilewater soluble drug is produced

intoW/OorW/O/W emulsion, it can be easily concentrated in the lymphatic

system by intramuscular or subcutaneous injection. The size of the emulsion

particle has an impact on its target distribution. Apart from its targeted

sustained release, producing the herbal drug into emulsion will also

strengthen the stability of the hydrolyzedmaterials, improve the penetrability of

drugs to the skin and mucous, and reduce the drugs' stimulus to tissues.

So far, somekinds of herbal drugs, suchas camptothecin, Brucea

javanica oil, coixenolide oil and zedoary oil have been made into emulsion.

For example, Zhou et al., (2004) studied the influence of the elemenum

emulsion on the human lung adenocarcinoma cell line A549 and protein

expression. Results showed that the elemenum emulsion has a significant

inhibition on the growth and proliferation of the A549 in vitro and it showed a

time and dose-dependent relationship. Elemenum emulsion is a type of new

anti-cancer drug with great application prospects. Furthermore, it has no

marrow inhibition and no harm to the heart and liver.

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Table 4: Emulsion herbal formulations. (Ajazuddin & Saraf, 2010)

Formulations Active Applications of Biological activity Method of preparation Droplet Drug Route ofingredients emulsion size loading administration

formulations

Self-nanoemulsifying Zedoary Improved aqueous Hepatoprotection Drawing ternary phase 68.3± 30% Oral

Zedoary essential oil turmeric dispersibility, anticancer and Diagram 1.6 nm

oil stability and oral anti-bacterial

bioavailability.

Triptolide micro- Triptolide Enhance the Anti- High pressure b100 nm – Topical

emulsion penetration of inflammatory Homogenization method

drugs through the

stratum corneum

by increased

hydration

Docetaxel submicron Docetaxel Improve residence Anticancer High pressure 166.00 nm 90% Intravenous

emulsion time Homogenization method

Berberine Berberine Improve residence Anticancer Drawing ternary phase 56.80 nm 0.50% Oral

nanoemulsion time and diagram

absorption

Silybin nanoemulsion Silybin Sustained release Hepatoprotective Emulsification method 21.20 nm – Intramuscular

formulation

Quercetin micro- Quercetin Enhance Antioxidant High speed 10– 0.3% Topical

emulsion penetration into Homogenization method 100 nm solution

stratum corneum

and epidermis

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Other novel vesicular herbal formulations

Transferosomes are applied in a non-occluded method to the skin,

which permeate through the stratum corneum lipid lamellar regions as a result

of the hydration or osmotic force in the skin. The carrier aggregate is

composed of at least one amphiphat (such as phosphatidylcholine), which in

aqueous solvents self-assembles into lipid bilayer that closes into a simple

lipid vesicle. By addition of at least one bilayer softening component (such as

a biocompatible surfactant or an amphiphile drug) lipid bilayer flexibility and

permeability are greatly increased. The resulting, flexibility and permeability

optimised, Transfersome vesicle can therefore adapt its shape to ambient

easily and rapidly, by adjusting local concentration of each bilayer component

to the local stress experienced by the bilayer. In its basic organization broadly

similar to a liposome), the Transfersome thus differs from such more

conventional vesicle primarily by its "softer", more deformable, and better

adjustable artificial membrane.Another beneficial consequence of strong

bilayer deformability is the increased Transfersome affinity to bind and retain

water. An ultradeformable and highly hydrophilic vesicle always seeks to

avoid dehydration; this may involve a transport process related to but not

identical with forward osmosis. For example, a Transfersome vesicle applied

on an open biological surface, such as non-occluded skin, tends to penetrate

its barrier and migrate into the water-rich deeper strata to secure its adequate

hydration. Barrier penetration involves reversible bilayer deformation, but

must not compromise unacceptably either the vesicle integrity or the barrier

properties for the underlying hydration affinity and gradient to remain in place.

Since it is too large to diffuse through the skin, the Transfersome needs to find

and enforce its own route through the organ. The Transfersome vesicles

usage in drug delivery consequently relies on the carrier’s ability to widen and

overcome the hydrophilic pores in the skin or some other (e.g. plant cuticle)

barrier. The subsequent, gradual agent release from the drug carrier allows

the drug molecules to diffuse and finally bind to their target. Drug transport to

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an intra-cellular action site may also involve the carrier’s lipid bilayer fusion

with the cell membrane, unless the vesicle is taken-up actively by the cell in

the process called endocytosis. It can be applicable as drug carriers for a

range of small molecules, peptides, proteins and herbal ingredients.

Transferosomes can penetrate stratum corneum and supply the nutrients

locally to maintain its functions resulting maintenance of skin (Benson, 2006)

in this connection the transferosomes of Capsaicin has been prepared by

Xiao-Ying et al., (2006) which shows the better topical absorption in

comparison to pure capsaicin.

Ethosome, as a novel liposome ie. Ethosomes are the modified forms

of liposomes that are high in ethanol content. The ethosomal system is

composed of phospholipid, ethanol and water, is especially suitable as a

topical or transdermal administration carrier (Jain et al., 2007; Fang et al.,

2008). The size of ethosomes vesicles can be modulated from tens of

nanometers to microns. Ethosome has a high deformability and entrapment

efficiency and can penetrate through the skin completely and improve drug

delivery through the skin. In contrast to liposomes, ethosomes have been

shown to exhibit high encapsulation efficiency for a wide range of molecules

including lipophilic drugs, and are selective at delivering molecules to and

through the skin, the physical and chemical properties of ethosomes make the

delivery of the drug through the stratum corneum into a deeper skin layer

efficiently or even into the blood circulation (Dayan and Touitou, 2000). This

property is very important as the topical drug carrier and transdermal delivery

system. Moreover, the ethosomes carrier also can provide an efficient

intracellular delivery for both hydrophilic and lipophilic drugs (Touitou et al.,

2001), percutaneous absorption of matrine an anti-inflammatory herbal drug is

increased. It also permits the antibacterial peptide to penetrate into the

fibrocyte easily. The roles of these types of novel vasicular system over herbal

drug delivery are summarized in (Table 5).

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Table 5: Other novel vesicular herbal formulations. (Ajazuddin & Saraf, 2010)

Formulations Active ingredients Applications Biological Droplet size Route of

activity administration

Capsaicin transferosomes Capsaicin Increase skin penetration Analgesic 150.6 nm Topical

Colchicine transferosomes Colchicine Increase skin penetration Antigout – In vitro

Vincristine transferosomes Vincristine Increase entrapment efficiency and skin Anticancer 120 nm In vitro

permeation y

Matrine ethosome Matrine Improve the percutaneous permeation Anti- 110±8 nm Topical

inflammatory

Ammonium glycyrrhizinate Ammonium Increase of the in vitro percutaneous Anti- 350 nm to Topical

ethosomes glycyrrhizinate permeation inflammatory 100 nm

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Microspheres

Administration of medication via micro particulate systems is

advantageous because microspheres can be ingested or injected and; they

can be tailored for desired release profiles and used site-specific delivery of

drugs and in some cases can even provide organ-targeted release (Sanli et

al., 2009). So far, a series of plant active ingredients, such as rutin,

camptothecin, zedoary oil, tetrandrine, quercetine and Cynara scolymus

extract has been made into microspheres (Table 6). In addition, reports on

immune microsphere and magnetic microsphere are also common in recent

years. Immune microsphere possesses the immune competence as a result

of the antibody and antigen was coated or adsorbed on the polymer

microspheres.

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Table 6: Microspheres encapsulated herbal formulations. (Ajazuddin & Saraf, 2010)

Formulations Active Applications of Biological activity Method of Size in Route of

ingredients formulations preparation µm administration

Rutin–alginate– Rutin Targeting into cardiocascular Cardiovascular and Complex- 165.00– In vitro

chitosan and cerebrovascular region Cerebrovascular coacervation method 195.00

microcapsules diseases

Zedoary oil Zedoary oil Sustained release and Higher Hepatoprotective Quasi-emulsion– 100– Oral

microsphere bioavailability solvent diffusion 600

method

CPT loaded Camptothecin Prolonged-release of Anticancer Oil-in-water 10 Intraperitoneally

microspheres camptothecin evaporation method and intravenously

Quercetin Quercetin Significantly decreases the Anticancer Solvent evaporation 6 In vitro

microspheres dose size

Cynara scolymus Cynara Controlled release of Nutritional Spray-drying 6–7 Oral

microspheres scolymus neutraceuticals supplement technique

extract

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Proprietary novel drug delivery system of plant actives and extracts

Cosmetochem International AG is a Swiss-based company, specialized

in the production of high quality, customized botanical extracts and actives,

launch botanical, standardized, liposomal powders named Liposome

Herbasec® [86] a novel range of standardized botanical extracts in a

liposomal-based powder form. As the liposome carriers are very effective

penetration enhancers which serve as carriers to the skin, increasing the

bioavailability of the plant extracts. In present formulation the freeze-dried

dispersion of Liposome Herbasec ® is reformed when dispersed in water, re

encapsulating the concentrated plant extract. Phospholipids used for the

preparation of formulation are the safest, mildest substances which allow the

penetration of the plant actives into the deeper layers of the epidermis and

avoid the use of solvents.There are five extracts in the current Liposome

Herbasec® range (Table 7) which are standardized for specific

phytochemicals. White and green tea are standardized for caffeine and total

polyphenols, white hibiscus for fruit acids, guarana for caffeine and aloe vera

is aloin-free. Liposome Herbasec® can be used in a wide range of personal

care applications. Smilarly based on Phytosome® technology, a line of

products has been developed and commercialized by Indena (Table 7). The

Phytosome® formulation increases the absorption of active ingredients when

topically applied on the skin (Bombardelli et al.,1991) and improves systemic

bioavailability when administered orally (Marczylo et al., 2007). A

Phytosome® is generally more bioavailable than a simple herbal extract due

to its enhanced capacity to cross the lipid-rich biomembranes and reach

circulation (Rossi et al., 2009). To overcome the poor bioavailability of silybin,

Indena has complexed it with soy phospholipids exploiting the Phytosome®

technology. As demonstrated by comparative pharmacokinetic studies,

Silipide® represents the most absorbable oral form of silybin known. The

pharmacokinetics of Silipide® in healthy human subjects showed that

complexation with phosphatidylcholine improved the oral bioavailability of

silybin 4-6 fold compared with silymarin, presumably because of a facilitated

passage across the gastrointestinal mucosa. The good bioavailability of

Siliphos® was confirmed in a human pharmacokinetic study in prostate

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cancer patients. The study employed high dosages, and was aimed at getting

information on toxicity and phase II dosage of the product. Siliphos® at a daily

oral dose of 13 g in 3 divided doses, was well tolerated in all patients, and this

dosage was recommended for the phase II study (Flaig et al., 2007). The

results, including the optimal tolerability obtained in these “extreme” clinical

situations, provide strong support for the use of Siliphos® also in less severe

pathologies associated with liver damage. Ginkgoselect® Phytosome® was

administered at a dosage of 360 mg/day (120 mg three times per day) to 22

subjects affected by the Raynaud's disease in a double-blind, placebo-

controlled trial. Patients were required to record the frequency and duration of

any vasospastic attack, also completing a scoring scale of the overall

perception of the severity of the episodes. Patients were reviewed after two,

four and ten weeks of treatment. This pilot study showed the efficacy of

Ginkgoselect® Phytosome® in promoting a clear and highly statistically

significant reduction in the frequency (56%) and severity of Raynaud's attacks

per day (Muir et al., 2002). Meriva® is a patented complex of curcumin, a

dietary phenolic, with soy phosphatidylcholine (Kidd, 2009). A lot of work that

has been published in the journal Cancer Chemotherapy and Pharmacology

(Marczylo et al., 2007) demonstrated Meriva®'s superior bioavailability

compared to a standardized curcumin extract in rats, while very promising

initial preclinical results in terms of improved hydrolytical stability and human

pharmacokinetics have been shown more recently. Including the advantages

of these above mentioned commercialized NDDS preparation of plant

actives/extracts a variety of other preparations is also available (Table 7)

which show the remarkable advantages over pure plant actives/extracts.

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Table 7: Marketed novel drug delivery formulations of plant active and extracts. (Ajazuddin & Saraf, 2010)

SN Brand name Plant active/extracts Type of Company

NDDS name

1 White tea liposome Herbasec® Camellia sinensis extract Liposome Cosmetochem

2 Green tea liposome Herbasec® Camellia sinensis Extract Liposome Cosmetochem

3 White hibiscus liposome Herbasec® White hibiscus extract Liposome Cosmetochem

4 Aloe vera liposome Herbasec® Aloe vera Extract Liposome Cosmetochem

5 Guarana liposome Herbasec® Guarana extract Liposome Cosmetochem

6 18ß-glycyrrhetinic acid Phytosome® 18ß-glycyrrhetinic acid from licorice rhizome Phytosome Indena

7 Centella Phytosome® Triterpenes from Centella asiatica leaf Phytosome Indena

8 Crataegus Phytosome® Vitexin-2″-O-rhamnoside from Hawthorn flower Phytosome Indena

9 Escin ß-sitosterol Phytosome® Escin ß-sitosterol from horse chestnut fruit Phytosome Indena

10 Ginkgoselect® Phytosome® Ginkgoflavonglucosides, ginkgolides, bilobalide from Phytosome Indena

Ginkgo biloba leaf

11 Ginselect® Phytosome® Ginsenosides from Panax ginseng rhizome Phytosome Indena

12 Ginkgo biloba terpenes Phytosome® Ginkgolides and bilobalide from Ginkgo biloba leaf Phytosome Indena

13 Ginkgo biloba dimeric flavonoids Phytosome® Dimeric flavonoids from Ginkgo biloba leaf Phytosome Indena

14 Greenselect® Phytosome® Polyphenols from green tea leaf Phytosome Indena

15 Leucoselect® Phytosome® Polyphenols from grape seed Phytosome Indena

16 Meriva® Curcuminoids from turmeric rhizome Phytosome Indena

17 PA2 Phytosome® Proanthocyanidin A2 from horse chestnut bark Phytosome Indena

18 Sericoside Phytosome® Sericoside from Terminalia sericea bark root Phytosome Indena

19 Siliphos® Silybin from milk thistle seed Phytosome Indena

20 Silymarin Phytosome® Silymarin from milk thistle seed Phytosome Indena

21 Virtiva® Ginkgoflavonglucosides, ginkgolides, bilobalide from Phytosome Indena

Ginkgo biloba leaf

22 Visnadex® Visnadin from Ammi visnaga umbel Phytosome Indena

24

Phytosomes as a boon for herbal drug delivery

Herbal drugs containing bioactive constituents are mainly water soluble

molecules. However, many flavonoid which are water soluble

phytoconstituents like to be poorly absorbed (Manach et al., 2004) due to their

poor miscibility with oils and other lipids or due to their multiple-ring large size

molecules which cannot be absorbed by simple diffusion, severely limiting

factors are available for their ability to pass across the lipid-rich outer

membranes of the enterocytes of the small intestine. Polyphenols (Water-

soluble phytoconstituents) molecules can be converted into lipid-compatible

molecular complexes, which are called Phytosomes. Phytosomes are more

bioavailable in comparison to simple herbal extracts. They have enhanced

capacity to cross the lipid rich biomembranes and finally reaching the blood

(Bombardelli et al., 1989). The lipid-phase substances employed to make

phytoconstituents, lipid compatible are phospholipids from soy, mainly

phosphatidylcholine (PC). Phospholipids are complex molecules that are used

in all known life forms to make cell membranes. The term “Phyto” means plant

while “some” means cell-like. The Phytosomes process itself produces a little

cell whereby the valuable component of the herbal extract is protected from

destruction by digestive secretions and gut bacteria. Many popular

standardized herbal extracts comprising of flavanoids, polyphenolics,

terpenes, alkaloids, volatile oils are employed for the preparation

ofphytosomes. Flavonoids are the most important group of phytochemicals.

Flavonoids are the class of compounds that have referred to be a natural

biological response modifier which acts as powerful antioxidants that

providing remarkable protection against oxidative and free radical damage.

Various flavonoids which have shown antioxidant activity 50 to 200 times

more potent than vitamin C or E. we can use certain flavonoids-rich extracts

that referred as “tissue specific antioxidants” due to their ability of

concentrated in specific body tissue. There are many plant drugs that are

incorporated to Phytosomes process as herbal extracts including Ginkgo

biloba, grape seed, hawthorn, milk thistle, green tea, and ginseng

.Phytosomes are more bioavailable as compared to conventional herbal

extracts owing to their enhanced capacity to cross the lipoidal biomembrane

and finally reaching the systemic circulation. So, Phytosomes has been a

novel approach for the herbal drug delivery (Bhattacharya, 2009).

Method of preparation:

25

Phytosomes are complexes chemical mixtures which are prepared by

reacting from with one or two mole of natural or synthetic phospholipids

phosphatidyl ethanolamine or phosphatidyiserine with one mole of

component. For example, flavolignanans, either alone or in the natural mixture

in aprotic solvent such as- dioxane or acetone from which complex can be

isolated by precipitation with non solvent such as aliphatic hydrocarbons or

lyophilization or by spray drying. In the complex formation of Phytosomes the

ratio between these two moieties is in the range from 0.5-2.0 moles. The most

preferable ratio of phospholipids to flavonoids is 1:1. In the Phytosomes

preparations, phospholipids are selected from the various group such as,

phosphatidyl, ethanolamine, phosphatidylcholine, soy lecithin, from bovine or

swine brain or dermis, phosphatidyiserine in which acyl group may be same

or different and mostly derived from palmitic, stearic, oleic and linoleic acid.

Flavonoids are selected from the group consisting of quercetin, kaempferol,

quercretin-3, rhamnoglucoside, quercetin- 3- rhamnoside, hyperoside,

vitexine, diosmine, 3- rhamnoside, (+). Some liposomal drugs complex

operate in the presence of the water or buffer solution where as phytosomes

operate with the solvent having a reduced dielectric constant. Flavonoid which

is the Starting material of component is insoluble in chloroform, ethyl ether or

benzene. They become extremely soluble in these solvents after forming

phytosomes. This chemical and physical property change is due to the

formation of a true stable complex (Sharma and Sikarwar, 2005).

How Phytosomes differ from liposome ?

Likewise Phytosomes, a liposome is formed by mixing

phosphatidylcholine with water soluble substance in definite ratio. The

phosphatidylcholine molecules surround the water soluble substance in which

no chemical bond is formed. There are hundreds or even thousands of

phosphatidylcholine molecules surrounding the water-soluble compound. In

contrast, with the Phytosomes process the plant components and the

phosphatidylcholine actually form a 1:1 or a 2:1 molecular complex depending

on the substance(s) complexed, in which chemical bond is formed. This

difference shows that Phytosomes being much better absorbed than liposome

showing better bioavailability. Phytosomes have also been found superior

than liposome in topical and skin care.

26

FIG. 1: PHYTOSOMES DIFFER FROM LIPOSOMES

Properties of Phytosomes

Chemical properties: Phytosomes are novel complexes formed between the

natural product and natural phospholipids, like soy phospholipids. Such a

complex is obtained by reaction of stoichometric amounts of phospholipid and

the substrate in an appropriate solvent. On the basis of spectroscopic data it

has been shown that the interaction of phospholipid-substrate is due to the

formation of hydrogen bonds between the polar head of phospholipids (i.e.

phosphate and ammonium groups) and the polar functionalities of the

substrate.

When it treated with water, phytosomes assumes a micellar shape which

formed the liposomal-like structures, In liposome the active principle is floating

in the layer membrane, while in phytosomes the active principle is anchored

to the polar head of phospholipids, becoming an integral part of the

membrane for example in the case of the catechindistearoyl

phosphatidylcholine complex, there is the formation of H-bonds between the

phosphate ion on the phosphatidylcholine side and the phenolic hydroxyls of

the flavone moiety.

27

FIG. 2: PHOSPHATIDYLCHOLINE COMPLEX

Phosphatidylcholine: This can be assumed from the comparison of the

NMR of the complex with the precursors of complex.The signals of the fatty

chain are almost unchanged. Such evidences inferred that the two long

aliphatic chains are wrapped around the active principle, producing a lipophilic

envelope, which shields the polar head of the phospholipid and the catechin.

Biological Properties: Phytosomes are advanced botanical technology that

offers improved absorption, enhanced delivery and increased bioavailability of

herbal extracts phytosomes over the non complexed botanical derivatives has

been demonstrated by pharmacokinetics studies or by pharmacodynamic

tests in experimental animals and in human subjects.

The Phytosome Technology: The flavonoid and terpenoid constituents of

plant extracts provide them for the direct binding to phosphatidylcholine.

Phytosomes results from the reaction of a stoichometric amount of the

phospholipid (phosphatidylcholine) with the standardized extract or

polyphenolic constituents in a non polar solvent (Bombardelli et al., 1989).

Phosphatidylcholine is a bifunctional compound, the phosphatidyl moiety

being lipophilic and the choline moiety being hydrophilic in nature. In

particular, the choline head of the phosphatidylcholine molecule binds to

28

these compounds while the lipid soluble phosphatidyl portion comprising the

body and tail which then envelopes the choline bound material

Hence, the Phytoconstituents produce a lipid compatible molecular complex

with phospholipids, also called as phytophospholipid complex. By specific

spectroscopic techniques, it can be demonstrated that the molecules are

anchored through chemical bonds to the polar choline head of the

phospholipids (Bombardelli, 1991). Precise chemical analysis designate that

the unit phytosome is usually a flavonoid molecule linked with at least one

phosphatidylcholine molecule. The result is a little micro sphere or cell is

produced. The term "Phyto" means plant while "some" means cell-like. The

phytosome technology produces a little cell, whereby the plant extract or its

active constituent is protected from destruction by gastric secretions and gut

bacteria owing to the gastro protective property of phosphatidylcholine.

Characterization of Phytosomes: There are many factors which govern the

behavior of phytosomes in both physical and biological system, such as

physical size membrane permeability; percent entrapped solutes, chemical

composition as well as the quantity and purity of the starting materials.

Therefore, the phytosomes are characterized for physical attributes i.e. shape,

size, its distribution, percentage drug capture entrapped volume, percentage

drug released and chemical composition (Jain, 2005).

Enhanced bioavailability: Recent researches showed that most of the

phytosomal studies are focused to Silybum marianum (milk thistle) which

contains premier liver-protectant flavonoids. The fruit of the milk thistle plant

contains flavonoids known for hepatoprotective effects (Bombardelli et al.,

1991). Silybin is the chief and most potent constituent of silymarin, the

flavonoid complex from milk thistle. A standardized extract from Silybum

marianum (milk thistle) is an excellent liver protectant but very poorly

absorbed orally.

Yanyu et al., (2006) prepared the silymarin phytosome and show its

pharmacokinetics in rats. In the study after oral administration of prepared

Silybin phospholipid complex, the bioavailability of Silybin in rats was

29

increased remarkably due to an impressive improvement of the lipophilic

property of Silybin-phospholipid complex and the biological effect of Silybin

was improved. Tedesco et al., (2004) reported Silymarin phytosome show

better anti-hepatotoxic activity than silymarin alone. Silymarin phytosome

provide protection against the toxic effects of aflatoxin B1 on performance of

broiler chicks.

Moscarella et al., (1993) perform a human study of 232 patients with

chronic hepatitis (viral, alcohol or drug induced). They are treated with Silybin

phytosome at a dose of 120 mg either twice daily or thrice daily function

returned to normal faster in patients taking Silybin phytosome compared to a

group of commercially available silymarin, 117 controls (49 treated with for up

to 120 days, liver untreated or given placebo). Studies have shown ginkgo

phytosome (prepared from the standardized extract of Ginkgo biloba leaves)

produced better results than the conventional standardized extract from the

plant.

Grape seed phytosome is prepared from grape seed extract containing

oligomeric polyphenols (grape proanthocyanidins or procyanidinsrom) of

varying molecular size, complexed with phospholipids. The main properties of

procyanidin flavonoids of grape seed that they increase the total antioxidant

capacity and stimulation of physiological antioxidant defenses of plasma,

protective effects against atherosclerosis thereby offering marked protection

for the cardiovascular system, protection against ischemia/reperfusion

induced damages in the heart, and other organs through a network of

mechanisms that extend beyond their great antioxidant potency (Schwitters

and Masquelier, 1993).

Despite such potential actions green tea polyphenols have very poor

oral bioavailability from conventional extracts. The complexation of green tea

polyphenols with phospholipids strongly improves their poor oral

bioavailability. A study on absorption of phytosomal preparations was

performed in healthy human volunteers along with complexed green tea

extract following oral administration. Over the study period of 6 hours the

plasma concentration of total non flavonoids was more than doubled when

30

coming from the phytosomal versus the nonphytosomal extract. Antioxidant

capacity was measured as TRAP (Total Radical-trapping Antioxidant

Parameter). The peak antioxidant effect was a 20% enhancement and it

showed that the phytosome formulation had about double the total antioxidant

effect. Maiti et al., (2005) developed the quercetin phospholipid phytosomal

complex which showed that the formulation exerted better therapeutic efficacy

than the molecule in rat liver injury induced by carbon tetrachloride. Recently

they developed the phytosomes of curcumin (flavonoid from turmeric,

Curcuma longa) and naringenin (flavonoid from grape fruit, Vitis vinifera) in

two different studies (Maiti et al., 2006). The antioxidant activity of the

quercetin phospholipid phytosomal complex was significantly higher than pure

curcumin in all dose levels tested.

Advantages of Phytosomes (Kidd and Head, 2005) : Phytosomes have the

following advantages;

Phosphatidylcholine used in preparation of phytosomes, besides acting

as a carrier also acts as a hepatoprotective, hence giving the

synergistic effect when hepatoprotective substances are employed.

They enhance the absorption of lipid insoluble polar phytoconstituents

through oral as well as topical route showing better bioavailability,

hence significantly greater therapeutic benefit.

As the absorption of active constituent(s) is improved, its dose

requirement is also reduced.

Chemical bonds are formed between phosphatidylcholine molecule

and phyto constituents, so the phytosomes show better stability profile.

Added nutritional benefit of phospholipids

31

FIG. 3: ORGANIZATION OF THE PHYTOSOME MOLECULAR COMPLEX

Applications of Phytosomes: There are many plant drugs that are

incorporated to Phytosomes process as herbal extracts including Ginkgo

biloba, grape seed, hawthorn, milk thistle, green tea, and ginseng. Most of the

phytosomal studies are focused to Silybum marianum which shows that it

contains premier liver-protectant flavonoids. The fruit of the milk thistle plant

(S. marianum, Family steraceae) contains flavonoids known for

hepatoprotective effects. It was found that Silymarin has been shown to have

positive effects in treating liver diseases of various kinds, including

inflammation of the bile duct, hepatitis, cirrhosis and fatty infiltration of the

liver. The antioxidant capacity of silymarin significantly boosts the liver’s

resistance to toxic insults (Valenzuela et al., 1989). Silymarin primarily

contains three flavonoids of the flavonol subclass. Silybin predominates,

followed by silydianin and silychristin. Silybin is a flavonolignan which is

probably produced within the plant by the combination of a flavonol with a

coniferyl alcohol. It is now known that Silybin is the most potent of the three

(Hiking et al., 1984). Silybin protects the liver by conserving glutathione in the

parenchymal cells, while PC helps repair and replace cell membranes (Kidd,

1996). These constituents offer the synergistic benefit of sparing liver cells

from destruction. In its native form within the milk thistle fruit, Silybin occurs

primarily complexed with sugars, as a flavonyl glycoside or flavonolignan

Silybin has been extensively researched and found to have impressive

bioactivity, albeit limited by poor bioavailability.

TABLE 8: COMMERCIAL PHYTOSOME PREPARATIONS

Phytosomes Phytoconstituent complexed Indication

32

with Phosphatidylcholine

Silybin

Phytosome TM

Silybin from Silymarin Food Product,

antioxidant for Liver

and skin.

Ginkgo

Phytosome TM

24 % ginkgoflavonglycosides

from Ginkgo biloba

Protects brain and

vascular lining, Anti-

skin ageing agent.

Panax ginseng

Phytosome TM

37.5 % ginsenosides from roots

of Panax ginseng

Food Product.

Green Tea

Phytosome TM

Epigallocatechin 3-O- gallate

from Camelia sinensi

Food Product,

Systemic antioxidant,

Cancer protectant.

Super Milk thistle

Extract

Silybin from Silymarin Food Product;

antioxidant for liver

and skin.

Grape seed

(PCO)

Phytosomes

Procyanidolic oligomers (PCOs)

from grape Seeds

Food Product; protects

against heart

Hawthorn

Phytosomes

Flavonoids Food Product.

Centella

Phytosome

Terpenes Vein & skin disorders.

CONCLUSION:

An extensive research is going on in the area of novel drug delivery

and targeting for plant actives and extracts. Herbal drugs have enormous

33

therapeutic potential which should be explored through some value added

drug delivery systems. Lipid solubility and molecular size are the major

limiting factors for drug molecules to pass the biological membrane to be

absorbed systematically following oral or topical administration. Standardized

plant extracts or mainly polar phytoconstituents like flavonoids, terpenoids,

tannins, xanthones when administered through novel drug delivery system

show much better absorption profile which enables them to cross the

biological membrane, resulting enhanced bioavailability. Phytosomes forms a

bridge between the convectional delivery system and novel delivery system.

The Phytosome process has been applied to many popular herbal extracts

including Ginkgo biloba, grape seed, hawthorn, milk thistle, green tea, and

ginseng. The flavonoid and terpenoid components of these herbal extracts

lend themselves quite well for the direct binding to phosphatidylcholine.

Through study of literature reveals that phytosome show promise in reliving

the pain and symptoms associated with asthma, arthritis, rheumatism, ulcers,

phlebitis, edema, varicose veins, premenstrual syndrome, diabetic retinopathy

and hemorrhoids. Phytosomes are used as a medicament and have wide

scope in cosmetology. Many areas of phytosome are to be revealed in future

in the prospect of pharmaceutical application.

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