Formulation And Evaluation of Anthralin Nano Particle Loaded Shampoo for Alopecia Areata

The history of diseases dates back to ancient civilizations, with natural remedies used to treat various            malfunctions. Scientific progress has led to the recognition and separation of active compounds in raw materials, their modes of action, and their chemical processes for lab-based synthesis. Drug delivery aims to transport the medicament to the target site at an adequate concentration for an adequate duration. Carrier systems have been continuously developed since their inception in the early 20th century to ensure that medicaments are delivered to the targeted site of action with regulated release. Development in polymer technology and nanotechnology led to the development of biodegradable and biocompatible polymers and a reduction in size or miniaturization techniques to see the path for drugs and their carrier system at the colloidal level. Nanotechnology has significantly transformed the strategies and emphasis of research and applications regarding disease diagnosis, treatment, monitoring, and management.

1.1 Nano Particle^6-8

The term "nanoparticle" in pharmaceutics implies structures less than 1μm in diameter.
Nanoparticles are particles having size range under 100 nm. These materials have significant roles in contemporary medicine in recent years, with uses varying from contrast agents in medical imaging to carriers for delivering genes to specific cells. The benefits of nanoparticles in modern medicine are numerous. Colloidal/Particulate system, developed through the colloidal method, offers potential for drug delivery in pharmaceutical-healthcare systems.The system includes lipid nanoparticles, liposomes, nanosomes, conjugates, nanoconjugates, nanocrystals, drugs, implants, nano constructs, nano emulsion, and nanosuspension.

1. 2. Shampoo⁹

Shampoo is a hair care product used to get rid of contaminants such oils, grime, dandruff, skin cells, and other skin and skin-related debris that slowly accumulate in hair. The concentration of shampoo is around 8% because it must be gentle on the skin and cause less discomfort to the eyes. Alcohol ether sulphates and salts like sodium are frequently employed as surfactants in shampoo.

1.3 Anthralin^10-13

Anthralin, a hydoxanthrones, is used for treating alopecia by causing redness or irritation of the scalp due to free radicals. It is applied as short-contact therapy, causing itching, redness, scaling, and staining of skin, hair, and clothes. Unstable anthralin is oxidized to anthralin dimmer, which is insoluble and eliminated in urine. Oxidized products cause violet-brown staining of cloths and skin, and do not possess hair regrowth activity.  It is also known as Anthralin, Dihydroxyanthranol, Anthranol and 1,8 – Dihydroxy 9Anthrone, Anthralinum (Latin), Anthralin (German), Anthralin (French), Ditranol (Spanish). It is an yellow or orange-yellow, fine crystalline, odourless or almost odorless powder. It is soluble in chloroform and oils, slightly soluble in ethanol (95%). Its absorption is more in skin with impaired stratum corneum barrier in 30 minutes, about 16 hours into intact skin. Its metabolism is oxidized to dianthrone, insoluble polymerization products. It is used for various psoriatic conditions and Alopecia areata. Its toxicity and side effects are directly proportional to the amount of drug product being used.

2. MATERIALS AND METHODS

Materials used in this work were procured from different manufactures and used without purification. All chemicals and solvents used was of analytical grade. Formulation components were examined on physicochemical ground. Pre-formulation studies were designed to fix optimum conditions for a stable formulation such as physical appearance, solubility analysis, melting point determination.^14-16

2.1 Preparation of Nanoparticle Loaded Shampoo

The nanoparticles were prepared by spontaneous emulsification method at various ratio of chitosan, oil and drug as mentioned in the table 4.3. The drug was dissolved in glacial acetic acid and kept aside. Then dissolve chitosan in glacial acetic acid and stirred for 2 hours and kept overnight. After the dissolution of chitosan, the drug was added into it by continuous stirring for 2 hours. Further the mixture of drug and chitosan nanoparticle were added drop by drop into oil phase to get a emulsion. The mixture was stirred continuously for 1 hour. Then glutaraldehyde was added into it and stirred for 2 hours. Add EDTA and ascorbic acid and stirred for 2 hours. Finally, the nanoparticle was collected by centrifugation at 10000rpm for 1 hour and dried nanoparticle were kept in desiccator.^17-19

Table 1.1 Formulation of drug loaded nanoparticle

Table no. 1.2 Ingredients and their composition of shampoo

2.2 Evaluation of nano shampoo²⁰

• Organoleptic properties: The colour of shampoo was visually examined against black and white background. The odour of the shampoo was also analysed.
• pH Measurement: Dissolved 5ml of shampoo in 100ml distilled water and then measured the pH meter.
• Viscosity: A Brookfield viscometer RV series model was used to calculate the viscosity of the produced mixtures. At 25°C, the viscosity of the chosen formulations was measured by pouring them into the viscometer's sample adaptor. The viscosity was tested five minutes following the spindle's rotation using spindle number 3 and 4 at a speed of ten revolutions per minute.
• Drug Content Determination: The drug content was determined using UV Spectrophotometer. 0.5g of nanoparticle was diluted with 10ml chloroform followed by continuous shaking until a clear solution and it is filtered with 0.45micron filter paper, from the filtered solution 5ml is taken and diluted to 10ml. Drug concentration in the solution was determined by UV Spectrophotometer at 254nm using formula ^[70]:

Drug Loading/ Encapsulation Efficiency: A small portion of Nano dispersion centrifuged at 10000rpm for 1 hour the supernatant was removed amount of unincorporated drug was measured by taking the absorbance of appropriately diluted supernatant solution at 254nm using VU Spectrophotometer, drug loading efficiency calculated using equation:

3. RESULTS AND DISCUSSION

Analysis of Drug

Table 1.3 Physical appearance of drug

Table 1.4 Solubility of Dithranol

Melting Point

Table 1.5 Melting point of dianthrol

Determination of λmax: The absorbance maxima of drug were found using AN-UV7000 UV-Visible spectroscopy and the maximum absorbance of the drug was found at 254nm,284nm, and 354 nm.

Figure 1.1 Absorbance maxima of dithranol

Table 1.6 Standard plot of dithranol

Figure 1.2 Calibration curve of dithranol

Figure No. 1.3 HPLC of Dithranol

pH: The pH of the F1, F2, F3, F4 was performed using MKVI,19370 MODEL pH meter and concluded that the F3 formulation shows the similar pH range of psoriatic skin. Hence F3 formulation is considered to be best for the psoriatic skin.

Table 1.7 pH of Dithranol Nano shampoo using different formulation

Viscosity: The Viscosity was performed for F1, F2, F3 and F4 formulation using Ostwald Viscometer and concluded that the formulations show good flow property which are needed for the shampoo.

Table 1.8 Viscosity of dithranol Nanoshampoo using different formulation

Drug Content Determination: The drug content determination was performed on various formulation using AN-UV7000 UV-Visible spectroscopy and the best drug content was found in the F3 formulation.

Table 1.9 Drug Content Determination of Dithranol Nanoshampoo using different formulation

Drug Content Determination: The drug content determination was performed on various formulation using AN-UV7000 UV-Visible spectroscopy and the best drug content was found in the F3 formulation.

Table 1.9 Drug Content Determination of Dithranol Nanoshampoo using different formulation

Drug Loading/ Encapsulation Efficiency: AN-UV7000 UV-Visible spectroscopy was used to perform drug loading efficacy of polymer (chitosan). The best encapsulation efficiency of chitosan: drug was found to be 2:1 ratio (F3 formulation).

Table 1.10 Drug Loading Efficiency of Dithranol Nano shampoo using different formulation

Particle Size and Surface Charge: The average particle size, zeta potential and polydispersity index of the developed nanoparticle were determined by using Litesizer 500. The particle size was found to be in the range of 380 to 500nm. The average particle size, zeta potential and polydispersity index of the developed nanoparticle were determined by using Litesizer 500. The zeta potential was found to be 31.2mV.      The F3 shows an average particle size and zeta potential of 382.2 nm and 31.2 mV respectively.

Figure 1.4 Particle size of dithranol nanoparticle

Figure 1.5 Zeta potential of dithranol particle

Figure 1.6 ATR interpretation of dithranol nanoparticle

Figure 1.7 Combined   ATR spectrum

Sem: The Scanning Electron Microscopy was performed by using JSM-IT710HR. The SEM image was disclosed, the F3 nanoparticle was spherical in shape and they have particle size of less than 10μm.

Figure 1.9 Scanning electron microscope image of NPs

Figure No. 1.10 X-Ray Diffraction of Nanoshampoo