INTRODUCTION:

Nanotechnology is the field of investigation since last century, Nanotechnology was first introduced by Nobel laureate Richarard P.Feynmen in his lecture of “There’s plenty of room at the bottom”. Then, there have been different advancements within the field of nanotechnology is made. The development of Nanotechnologies is important in terms of diagnosis, treatment, and prevention of disease. The word nanoparticles come from the Greek word nanus which means dwarf or very small. Nanoparticles (NPs) are the novel invention of modern science in which drug is surrounded by a polymeric membrane where the drugs are dissolved, entrapped, adsorbed, attached and/or encapsulated into or onto a Nano-particulate matrix. The drug delivery vehicles are generally < 100 nm in size with at least one dimension and consist of different biodegradable materials such as natural or synthetic polymers, lipids, or metals. It is composed of three layers i.e. (a) the surface layer, which can be functionalized with a variety of little molecules, metal ions, surfactants, and polymers. (b) The shell layer, which is with chemicals completely different material from the core in all aspects, and (c) The core, that is the central portion of the NPs¹.

Several polymers are being utilized for the fabrication of nanoparticles. Macromolecules include peptides and protein can be easily and effectively deliver through the Nano delivery system. Nanoparticles deliver the drug at a controlled and sustained rate to the site of action².

Nowadays, nanoparticles are more attractive due to some of their unique features such as surface to mass ratio, ability to adsorb and carry compounds such as drugs, probe, and proteins³. Nanoparticles also represent a promising carrier system for the targeting of anti-cancer agents to tumors. They are also reported successfully employed in Brain Drug Targeting. Hexapeptide Dalargin (Tyr-D-Ala-Gly-Phe-Leu-Arg), is the first drug nanoparticle drug that was delivered to the brain as the other medicine that was successfully transported into the brain in a form of nanoparticles are loperamide, phytotoxin, and antibiotic drug³. So, where conventional techniques reach their limits, nanotechnology provides opportunities for medical applications.

ADVANTAGES OF NANOPARTICLES:

1. Site-specific targeting is often achieved by attaching targeting ligands to the surface of particles.

2. Drug release can be controlled or sustained which will increase the therapeutic efficacy of a drug.

3. Side effects and toxicity shall be reduced.

4. Passive and active drug targeting can be easily achieved by manipulating surface and particle size characteristics.

5. Both hydrophilic and hydrophobic drug can be easily delivered.

6. It can be administered through different routes like oral, nasal, parenteral.

Disadvantages:

1. The cost of manufacture is high and encapsulation efficiency is less.

2. The solvent system used during the preparation process may produce toxicity.

3. Particle aggregation and physical handling of nanoparticles in dry and liquid form are difficult.

4. Leakage and sudden release of the drug may be one of the critical problems.

5. The higher surface to volume ratio makes the particles more reactive or catalytic.

Types of Nanoparticles:

Nanoparticles can be classified in various types based on their structures, sizes or physical and chemical properties. A few of them are carbon-based nanoparticles, lipid-based nanoparticles, and polymeric nanoparticles.

Carbon-based Nanoparticles:

These nanoparticles contain carbons. It includes two main materials: carbon nanotubes (CNTs) and fullerenes. CNTs are graphene sheets that are rolled into a tube. These materials are mainly used for structural strengthening as they are 100 times stronger than steel. CNTs are classified into single-walled carbon nanotubes (SWCNTs) and multi-walled carbon nanotubes (MWCNTs). CNTs are one of a kind in a way as they are thermally conductive along the length and non-conductive over the tube.

Fullerenes are the allotropes of carbon. Their structure is like a hollow cage as shown in the Figure 1. below having sixty or more carbon atoms. The structure of C-60 is called Buckminsterfullerene, which looks like a hollow football. In these structures, carbon units are having a pentagonal and hexagonal arrangement.

Ceramic Nanoparticles:

They are inorganic solids made up of oxides, carbides, carbonates, and phosphates. These types of nanoparticles are having chemical inertness and high heat resistance. These are useful in drug delivery for many diseases like bacterial infections, glaucoma, and cancer.

Metal Nanoparticles:

These nanoparticles can synthesize by chemical, electrochemical, or photochemical strategies. By chemical methods, we can get metal nanoparticles by reducing the metal-ion precursors in solution by chemical reducing agents. These can adsorb small molecules and have high surface energy. They are widely used in research areas, detection and imaging of biomolecules, environmental and bioanalytical applications.

Semiconductor Nanoparticles:

They are having properties like those of metals and non-metals. They are found in the periodic table in groups II-VI, III-V or IV-VI. Some examples are InP, InAs GaP, GaN , germanium and silicon etc. These are used in electronics devices, photo-optics, photocatalysis, and water splitting applications.

Lipid-Based Nanoparticles:

They are generally spherical in shape and having a diameter ranging from 10 to 100nm. It comprises of a strong center made of lipid and a network containing soluble lipophilic particles. The outside layer of these nanoparticles is stabilized by surfactants and emulsifiers. These nanoparticles have applications in the biomedical field as a drug carrier and delivery and RNA release in cancer therapy.

Carbon-Based Ceranuc Babiopartucles

Metal Nanoparticles Semiconductor Nanoparticles

Lipid Based Nanoparticles

Fig 1: Classification of Nanoparticle

Classification of Nanoparticles Based on their Dimensions:

Nanoparticles are also classified depending on their crystalline forms and chemical composition which are 0D, 1D, 2D and 3D Nanoparticles. This classification is based on the electron movement along the dimensions in the NPs.

In 0D NPs, electrons are entrapped in a dimensionless space.

In 1D NPs, electrons have electrons that can move along the x-axis, which is less than 100 nm.

2D and 3D NPs have electron movement along the x–y-axis, and x, y, z-axis respectively.

Spheres and clusters nanofivers wires and rods

thin films, plates bulk nanomaterials

Fig 2: Classification of Nanoparticle based on dimension

Mechanisms of Drug Release:

The drug from the polymeric drug carriers deliver at the site of the tissue by anyone of the three general physico-chemical mechanisms which are explained below:

1. By hydration which causes the swelling of the polymer nanoparticles followed by release through diffusion.

2. By an enzymatic reaction that leads to rupture or degradation or cleavage of the polymer at the site of delivery and results in the release of the drug from the entrapped inner core.

3. Dissociation of the drug from the polymer and it does de-adsorption/release from the swelled nanoparticles.

Polymer Used for the Preparation of Nanoparticles:

Polymers based nanomaterials are the vehicles for control release of the drug, in which drug can be either adsorbed on their surface or entrapped inside. Polymers utilized in preparation of nanoparticles should be biocompatible and biodegradable. It can be either from natural source or synthetic source^4,5,6. List of polymers can be ulilized are given below:

· Natural polymers

a) Chitosan

b) Gelatin

c) Alginate

· Synthetic polymers

a) Poly lactide (PLA)

b) Poly acrylate

c) Poly mathacrylate

d) polycaprolactones

e) Poly lactide-co-glycolide (PLGA)

METHODS OF PREPARATIONS:

Fig 3: Method of preparations

1. Nanoparticles Preparation by Cross Linking Techniques:

These nanoparticles are prepared from amphiphilic macromolecules, proteins and polysaccharides which have an affinity for aqueous and lipid solvents. The technique of their preparation involves the aggregation of amphiphiles followed by further stabilization is done either by heat denaturation or chemical cross-linking.

The method involves the emulsification of bovine serum albumin/human serum albumin or protein aqueous solution in oil using high-pressure homogenization or high-frequency sonication. The w/o emulsion so formed is poured into preheated oil (heat cross-linking). The suspension in preheated oil maintained above 100-degree centigrade is held stirred for the time to denature and aggregate the protein contents and to evaporate water. The particles formed were washed with natural solvent to expel any oil traces and collected through centrifugation. For heat-sensitive substance, chemical cross-linking are done.

Cross-linking can be done by two method:

A) Heat denaturation

Fig 4: Heat denaturation method

A) By chemical cross-linking agent

Fig 5: Chemical cross-linking method

2. POLYMERIZATION BASED METHODS:

A) Emulsion polymerization:

Emulsion polymerization includes emulsion containing water, monomer, and surfactant. The oil-in-water emulsion is the most common type of emulsion polymerization, in which droplets of monomer are emulsified in a continuous stage of water. Emulsion polymerization is one of the quickest methods for the preparation of nanoparticle. It involves the dispersion of the monomer into a solvent in which the monomer is not soluble (non-solvent). Surfactants or protective soluble polymers are used to prevent aggregation in the early stages of polymerization. Then the polymerization process can be initiated by different mechanisms like by applying high energy radiation like UV or visible light, monomers can be changed into initiating radicals. When a monomer collides with these radicals then, initiation takes place. Phase separation and formation of solid particles can take place before or after the termination of the polymerization reaction⁷.

Fig 6: Emulsion polymerization method

B) Dispersion polymerization:

It involves a homogenous system formed when monomers, initiators and a stabilizer dissolved in a solvent which will result in the formation of polymer particles. In this method, the monomers and initiators are easily dissolvable in a solvent used for a reaction medium but is a non – solvent. Nucleation is directly induced in aq. monomer solution. So stabilizer/surfactant is not needed. Initiation is achieved by the same mechanism as in emulsion polymerization by high energy radiation. Polymerization is started by including a catalyst and continues with the nucleation stage taken after by development stage^8,9.

Fig 7: Polymerization dispersion method

C) Interfacial polymerization:

It is a type of step-growth polymerization in which polymerization takes place at the interface between two immiscible phases (generally two liquids). It is one of the well-established strategies utilized for making nanoparticles. It involves polymerization of two reactive monomers or agents, which are dissolved respectively in two phases (i.e., continuous- and dispersed-phase), and the reaction takes place at the interface of the two liquids. Oil-containing Nanocapsules were obtained by the polymerization of monomers at the oil/water interface of a very fine oil-in-water microemulsion^10,11.

Fig 8: Interfacial polymerization method

3. POLYMER PRECIPITATION METHODS:

a) Solvent evaporation:

In this method, polymers are dissolved in an organic solvent such as chloroform, dichloromethane, and then the drug is dispersed in this solution. Then, this mixture emulsified in an aqueous phase containing surfactant e.g. sodium dodecyl sulfates. With the help of mechanical stirring, sonication, or micro fluidization (high-pressure homogenization), make oil in water emulsion.

Then, the organic solvent is evaporated by increased temperature and reduced pressure with continuous stirring. The size of nanoparticles can be controlled by adjusting the stirring rate, type and amount of dispersing agent, a viscosity of organic and aqueous phases and temperature. Polymers which are used in this method are PLA, PLGA, cellulose acetate phthalate, Poly β-hydroxybutyrate (PHB).¹².

Fig 9: Solvent evaporation method

b) Double emulsion method:

This method is useful for incorporating hydrophilic drugs because of poor entrapment of hydrophilic drugs by emulsification and evaporation methods, so double emulsification technique is utilized. In this method, w/o emulsion is prepared by the addition of aqueous drug solution to organic polymer solution with continuous stirring. This prepared emulsion is further added into another aqueous phase with vigorous stirring, which will give w/o/w emulsion, then the organic solvent removed by high centrifugation^12,13,14.

Fig 10: Double emulsion method

c) Emulsions- Diffusion Method:

It is a modified version of the solvent evaporation method. This method is an alternative method for avoiding the toxicity-solvent problems caused by emulsification-evaporation methods. It has simple implementation and high reproducibility. It is also used to encapsulate several kinds of drugs, including peptides and proteins. In this method, the encapsulating polymer is dissolved in a partially water-soluble solvent such as propylene carbonate and saturated with water to ensure the initial thermodynamic equilibrium of both liquids. Subsequently, the polymer-water saturated solvent phase is emulsified in an aqueous solution containing a stabilizer, which leads to solvent diffusion to the external phase and the formation of Nanospheres or Nanocapsules, according to the oil-to-polymer ratio. Finally, the solvent is eliminated by evaporation or filtration, according to its boiling point.^15,16.

Fig 11: Emulsion diffusion method

d) Salting Out Method:

The salting out procedure is a modification of the emulsification/solvent diffusion method. Polymer and drug are initially dissolved in a solvent. In this method, toxic solvents are not used. Generally, acetone is used as it is miscible with water and easily removed. Then the above mixture is subsequently emulsified into an aqueous gel containing the salting-out agent such as electrolytes, magnesium chloride, calcium chloride, and magnesium acetate, or non- electrolytes like sucrose along with a colloidal stabilizer (polyvinyl pyrrolidone or hydroxyethylcellulose). The oil/water emulsion is diluted with a sufficient volume of water or an aqueous solution to enhance the diffusion of acetone into the aqueous phase, thus inducing the formation of nanospheres.^17,18.

Fig 12: Salting out method

EVALUATION:

1) Particle size:

Particle size distribution and morphology are the most important parameters of the characterization of nanoparticles. Morphology and size determination is done by using photon correlation spectroscopy or dynamic light scattering. The results obtained by the photon correlation spectroscopy is verified by utilizing scanning electron microscopy (SEM). SEM technique is based on an electron scanning principle, which will give available information about the NPs at the Nanoscale level. Wide literature is available, where people used this technique to study not only for determining the morphology of their nanomaterial but also the dispersion of NPs in the bulk or matrix [1]. It was found that 100nm nanoparticle is having grater uptake as compare to 1m particle ¹⁹.

2) In-vitro release Study:

It is performed in USP Type II dissolution apparatus at a rotation speed of 50 rpm. The prepared immersed in 900ml of phosphate buffer solution in a vessel, and temperature should be maintained at 37±0.20°C. Required quantity 5ml of the medium was withdrawn at specific periods and the same volume of dissolution medium was replaced in the vessel to maintain a constant volume. The withdrawn samples are analyzed using uv- spectrophotometer²⁰.

3) Stability of Nanoparticles:

Stability studies of prepared nanoparticles determined by placing the formulation at 4°C ±1°C and 30°C ± 2°C in the stability chamber for 90 days. Then, the samples were analyzed after some time like at 0, 1, 2, and 3 months to check any changes in their physical appearance or for their drug content, drug release rate²¹.

4) Structure and Crystanillity:

There are many methods for the determination of structure and crystallinity. The x-ray diffraction method is most widely used to determine the structure and crystallinity. X-ray imaging begins with a beam of high energy electrons crashing into a metal target and x-rays are produced. A filter near the x-ray source blocks. These low energy rays, which mean only the high energy rays pass through a patient toward a sheet of photographic film. The x-ray can penetrate liquids, gas, and solids. The point of penetration is based on the intensity, quality, and wavelength of the X-ray beams.

5) Yield of Nanoparticles:

The yield of nanoparticles can be obtained by comparing the whole weight of nanoparticles formed with the combined weight of the copolymer and drug.

% Yield = Amount of Nanoparticle ×100/Amount of drug + Polymer

6) Drug Content, Surface entrapment, Drug entrapment:

Amount of drug present in the supernatant (w) determined by UV spectrophotometer. After that standard calibration curve plotted. Then the amount of drug present in supernatant subtracted from the total amount used in the preparation of nanoparticles (W). (W-w) is the amount of drug entrapped. % drug entrapment calculated by.

W - W

% durg entrapment = ---------------- ×100

APPLICATION:

A nanoparticle has great prospects for the diagnosis and treatment of diseases. It has been widely employed for different therapeutic applications, some of which are listed below:

· Nanoparticles are reported to effectively combat against intracellular infections of the human body.

· Nanoparticles can effectively deliver chemotherapeutics drugs with reduced toxicity and increase therapeutic activity.

· Nanoparticles are successfully served the purpose of drug targeting to specific areas with minimum side-effects.

· Nanoparticle systems can improve the delivery of the drug to eyes through the increase retention time.

· It can be used as carriers for radio nucleotides for diagnostic purposes.

· It is very useful for improving the solubility and bioavailability of poorly soluble drugs and protects the drug from gastrointestinal enzymes.

· Solid nanoparticles are useful for hair and skin care therapy.

· Nanoparticle drug delivery systems can effectively deliver drugs across the blood-brain barrier (BBB).

· Many macromolecules such as protein and peptides are effectively delivered through nanoformulations.

· It is useful as a vaccine adjuvants, which increases the uptake of the drug and increases the systemic circulation of the drug¹²^,¹³^,²⁰.

Inspite of various preclinical research, only some of them approved for marketing, some of them are discussed below:

Nanoparticles are useful for targeting tumours due to its hight retentiability like, doxorubicin nanospheres showed greater concentration in the liver , spleen and lungs as compared to free doxorubicin²².

Also, now- a- days gold nanoparticles (AU NPs) are being utilizing in medical imaging for early diagnosis or detection and treatment of diseases which includes tumour targetting. Gold NPs composed gold atoms surrounded by negative reactive groups so they can be easily biofuntionalized with a wide range of biomolecules. These nanoparticles are having surface plasmon resonance (SPR) bands from which light is converted into heat and scateer the heat for killing cancer cell.

There are also some polymers like degrabale hydrophobic polymers e.g PLA, PLGA are useful for development of nanoformulation because they slowly decompose into their monomeric units over certain period of time. For e.g Leuprolide ( A testosterone –inhibiting drug) incorporated with polylactide –co-glycolic acid (PLGA) nanoparticles was found to be very useful for treating prostate cancer.

Polyethylene glycol is a polymer which is now useful for preventing going degradation of drug by the immune system. For e.g Adynovate formulation is useful for treating hemophilia- A, which was prepared by PEGylation of antihemophilic factor VIII²³.

Some of marketed nanoparticle formulation to treat diseases is given in Table No.1.

Table no 1 : Marketed preparations of Nanoparticles

Sr No	Name	Active ingredient	Used for	RA
1	Megace ES®	Megestrol acetate	Anorexia	Oral
2	Emend®	Aprepitant	Emesis, antiemetics	Oral
3	Rapamune®	Rapamycin	Immunosuppressant	Oral
4	Copaxone®	Polypeptide composed of four amino acids (glatiramer)	Multiple sclerosis	SC
5	Abraxane®	Nanoparticles (130 nm) formed by albumin with conjugated paclitaxel	Metastatic breast cancer	I.V
6	Ferumoxytol	Superparamagnetic iron oxide nanoparticles coated with dextran.	Treatment of iron deficiency anemia in adults with chronic kidney disease	I.V

RA= Route of administration

CONCLUSION:

Nanoparticles may be considered as a part of a novel drug delivery system, which can play a vital role in achieving effective therapeutic benefits, better bioavailability with minimum side effects and toxicity. It has made major contributions to the modern medical system and expected to create a milestone for the emerging health care system.

ACKNOWLEDGEMENT:

Author would like to acknowledge Assam down town University for providing library facilities and other necessary facilities to carry out the task.

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Received on 18.04.2020 Modified on 27.05.2020

Research J. Pharm. and Tech. 2021; 14(3):1815-1822.

DOI: 10.5958/0974-360X.2021.00322.X