INTRODUCTION:

Nanoparticles (NPs) are defined as particulate dispersions or solid particles drug carrier that may or may not be biodegradable, where the drug are dissolved, entrapped, encapsulated or attached to a nanoparticle matrix. The term nanoparticle is a combined name for both nanospheres and nanocapsules in which the drug is confined to an aqueous or oily core surrounded by a shell like wall, while nanospheres are matrix systems in which the drug is physically and uniformly dispersed. Where conventional techniques reach their limits, nanotechnology provides opportunities for the medical applications¹.

The major goals in designing nanoparticles as a delivery system are to control particle size, surface properties and release of pharmacologically active agents in order to achieve the site-specific action of the drug at the therapeutically optimal rate and dose regimen².

Simvastatin is well absorbed from the gastrointestinal tract but is highly extracted by the liver and only 7 percent of the dose reaches the general circulation. It possesses biological half-life of 2 hours.

Simvastatin also undergoes extensive first pass metabolism in intestinal gut and liver. Hence the oral bioavailability of Simvastatin in its intact form is only five percent. All Statins act by Inhibiting 3 Hydroxy 3- Methyl Glutaryl Coenzyme A, A HMG- COA reductase, the rate limiting enzyme of the HMG-COA reductase pathway which is responsible for the endogenous production of cholesterol, hence this study focuses on converting Simvastatin in form of polymeric nanoparticles which could improve the dissolution rate and bioavailability of drug³.

The main aim of present study was preparation of nanoparticles using different polymers like sodium alginate and chitosan and different concentrations by High pressure homogenization method.

MATERIALS AND METHODS:

Materials:

Simvastatin obtained as gift samples MITS Healthcare Pvt Ltd, Panchkula. Glyceryl Monostearate Lipoid manufacturers, Germany. Chitosan, Sodium Alginate and tween 80 Merck specialities private limited, Mumbai.

Methods: Drug excipient interaction study^4,5

FTIR spectroscopy studies:

Drug-excipients compatibilty studies were carried out using FT-IR spectroscopy. All the excipients such as Chitosan, Sodium Alginate, Glyceryl monostearate and Tween 80 individually, physical mixture of excipients, pure drug Simvastatin individually, physical mixture of excipients and drug were mixed separately with infrared (IR) grade KBr in the ratio of 1:100 and corresponding pellets were prepared by applying 15000 lb of pressure in a hydraulic press. The pellets were scanned in an inert atmosphere over a wave number range of 3500-500 cm-1 in FTIR instrument.

Formulation of simvastatin nanoparticles:

Table1: Formulation of Simvastatin Nanoparticles

INGREDIENTS	F1	F2	F3	F4	F5	F6	F7	F8
Simvastatin (mg)	40	40	40	40	40	40	40	40
Glyceryl Monostearate (ml)	100	100	100	100	100	100	100	100
Sodium Alginate(mg)	40	60	80	100	--	---	---	---
Chitosan (mg)	---	---	---	---	40	60	80	100
Tween 80 (ml)	100	100	100	100	100	100	100	100
Methanol(ml)	1	1	1	1	1	1	1	1
Water(ml)	50	50	50	50	50	50	50	50

METHOD OF PREPARATION:

Simvastatin nanoparticles were prepared by High pressure homogenization method by using lipid (Glyceryl Monostearate), polymer (Chitosan, Sodium Alginate) and surfactant (Tween 80) in the formulation. Methanol was used as a solvent. Formulations composition was showed in the table 1.

The calculated amount of polymer, lipid and drug was solubilised in the organic solvent (solvent phase) and surfactant was solubilised in the water (aqueous phase) then while undergo coarse homogenization the solvent phase is added aqueous phase and made it into uniform size of these ingredients then undergoes for the process of High pressure homogenization to get smaller size particles of drug and its excipients. Then formulation was taken into the round bottomed flask and undergo for the rotary evaporation to remove the solvent present in the formulation. Then measure the particle size and zeta potential of the formulation. Observing the formulation physically whether its forms crystals or precipitation in the formulation^6,7.

Evaluation of simvastatin nanoparticles^8,9:

Particle size:

The particle size of the formulation was determined by taking the formulation and making the necessary dilutions of it and measures the particle size by using the zeta sizer (Malvern instruments). For it the particle size of the formulation can be known.

Zeta potential:

Zeta potential shows the charge on the particle surface which indicates the physical stability of dispersed systems. The zeta potential of nanoparticles is measured using Malvern zetasizer ZS 200. The zeta potential were calculated by using the Helmholtz-Smoluchowski equation.

Drug content¹⁰:

Take 1ml of formulation solution from the formulation in 10ml volumetric flask and add phosphate buffer pH 6.8 & methanol mixture to make dissolve and make necessary dilutions with the phosphate buffer pH 6.8.The diluted solutions were analysed at 245 nm in UV-Visible spectrophotometer (Thermo fisher scientific). The drug content of the formulation can be known. It is triplicated.

Total drug content =

Concentration × D.F × total formulation volume

(D.F = dilution factor)

Drug entrapment efficiency^11,12:

Take 5ml of formulation solution in a centrifuge tube and keep this in a ultracentrifugation at 15,000 RPM, at 4 ͦ c temperature for 1¹/₂ hour. From this take 0.5ml of supernatant solution and to this add 0.5ml of methanol and 4ml of phosphate buffer pH 6.8. This diluent solution observance taken by keeping in UV-Visible spectrophotometer from this % drug unentrapment is known. From this %drug entrapment efficiency is known. It is made to triplicated.

% Drug entrappment efficiency =

Total amount of drug – free drug/Total amount of drug

In vitro Drug release studies^13,14:

Take 900ml of pH6.8 phosphate buffer in a USP dissolution test apparatus and maintained at 37±0.5 ͦ C temperature and 75rpm rotation. To this add an equivalent amount of drug (40mg) solution into buffer solution. From this take a 5ml of sample at regular time intervals up to 12hrs. Then filter the samples by stirred cell and collect the filtrate. These samples are further diluted and absorbance was measured in UV-Visible spectrophotometer at 245nm. The drug release rates of Simvastatin in samples were calculated by using the regression equation of the calibration curve.

Scanning electron microscopy^15,16(SEM):

The Surface morphology of Simvastatin Nanoparticles is measured by scanning electron microscopy S-3700N. Scanning electron microscopy is a test process that scans a sample with an electronic beam to produce a magnified image for analysis. During Scanning electron microscopy analysis, the signals generated produce a two dimensional image and reveal information about the sample, including chemical composition and external morphology

Comparison of % drug release rate between Marketed Simvastatin tablet and optimized nanoparticles¹⁷:

Take marketed Simvastatin (Simvofix 40mg) and carried out the dissolution study in the Dissolution Apparatus USP II. Take 900ml of dissolution media and to this add 40mg of Simvofix tablet. Then samples are collected at regular intervals of time and made to filter it. Collect the filtrate and made necessary dilutions and measure the Absorbance by keeping in UV-Visible spectrophotometer at 245 nm. The percentage release rates of Simvastatin in samples were calculated by using the regression equation of the calibration curve. Then triplicate it. Then compare the dissolution release rates of tablet and nanoparticles.

The FTIR spectra of the pure drug, and optimized formulation (F8) are shown in figure 1 and 2 respectively, there is no major shifting of functional groups, hence confirmed the absence of interaction between drug and the excipients mixtures.

The particle size and polydispersity index are found to be 97.47±2.22nm to 156.1±1.22nm and 0.087±0.015 to 0.445±0.016 respectively. The results are shown in shown in Table 2 and figure 3

Table.2: Evaluation parameters of Particle size, PDI, Zeta potential:

formulation Code	Particle size (nm)	Polydispersity Index (PI)	Zeta potential (mV)
F1	111.8±3.61	0.268±0.012	-19.13±2.61
F2	105.8±2.48	0.342±0.018	-19.34±2.68
F3	128.3±2.15	0.366±0.013	-20.63±3.61
F4	156.1±1.22	0.445±0.016	-21.23±2.58
F5	146.83±4.73	0.219±0.011	-23.90±3.41
F6	127.75±4.68	0.087±0.015	-14.80±2.48
F7	119.3±3.11	0.297±0.016	-27.24±3.51
F8	97.47±2.22	0.333±0.018	-30.60±4.72

The values represent mean ± SD

Table 3: Evaluation of Total drug content and % Drug Entrapment efficiency

Formulation code	Total drug content (mg)	Free drug (mg)	% Drug Entrapment efficiency
F1	39.75±0.90	7.09±0.56	82.16±2.29
F2	39.58±1.10	7.83±0.73	80.21±3.88
F3	39.63±0.68	7.92±0.59	78.11±5.45
F4	39.22±1.20	6.89±0.59	80.55±4.75
F5	39.52±1.19	6.63±0.58	83.22±2.62
F6	39.67±0.91	7.98±0.61	79.88±3.43
F7	39.78±1.11	6.79±0.73	82.93±4.60
F8	39.92±1.45	5.71±0.82	85.69±5.23

The values represent mean ± SD, n = 3.

The total drug content in the formulation, F1 to F8 were found to be 39.22mg to 39.92mg and it was observed that total drug content were uniformity. The results are shown in Table 4.3. The %Drug Entrapment efficiency in the formulations F1 to F8 are known by taking formulation in the centrifuge tube and undergoing ultracentrifugation. Then calculate free drug in the formulations. For the prepared Simvastatin nanoparticles F1 to F8, the % drug Entrapment efficiency are found to be 78.11% to 85.69%. The results are shown in Table 3

In vitro dissolution studies:

Figure 4: In-vitro dissolution profiles of Simvastatin nanoparticles (F1-F4)

Figure 5: In-vitro dissolution profile Simvastatin nanoparticles(F5-F8)

The in vitro release study of Simvastatin nanoparticles formulations F1 to F8 were conducted at pH6.8 phosphate buffer for 12 hours. The formulation, F1 to F4 by using Sodium Alginate were found to be 80.87% to 89.87% at the end of 12 hrs. Similarly, the drug release study of formulations F5 to F8 by using Chitosan were found to be 82.81% to 97.28% at the end of 12 hrs It was found that the amount of polymer increased, drug release was sustained. The formulation, F8 showed the maximum 97.28% drug release at the end of 12hours. So, formulations F8 showed better sustain the drug release for 12hrs period of time as compared to other prepared formulations. The in-vitro dissolution drug release of all the formulations as shown in and Figure 4 and figure 5. Among all the formulations, F8 was best formulation based on dissolution studies, hence release kinetics was applied.

Table 4: Drug release kinetic of all formulations of Simvastatin nanoparticles (F1 to F8)

Formulations	Zero R²	First order R²	Higuchi R²	Korsmeyer-Peppas R²	N-values
F1	0.934	0.728	0.732	0.998	0.657
F2	0.950	0.845	0.917	0.982	0.527
F3	0.989	0.805	0.904	0.983	0.554
F4	0.965	0.984	0.922	0.996	0.657
F5	0.982	0.744	0.908	0.995	0.706
F6	0.975	0.645	0.899	0.993	0.782
F7	0.896	0.890	0.909	0.956	0.858
F8	0.912	0.910	0.953	0.985	0.781

The data obtained from dissolution study of F8 formulation was fitted into various release kinetics models and the value of resulting regression coefficient is calculated. The data was also fitted into Korsmeyer-Peppas model in order to obtain the ‘n’ value to describe the mechanism of drug release. The ‘n’ value of 0.781 indicates that the drug release follows anomalous (non-fickian) diffusion mechanism which signifies that the drug release is diffusion-controlled. From these results, we concluded that optimized formulation, F8 showed Zero order with non-fickian mechanism. The results are shown in Table 4

SCANNING ELECTRON MICROSCOPY (SEM):

The SEM photograph of formulation reveals that Simvastatin nanoparticles were spherical and moderately uniform shown in figure 6

Figure 6: SEM of Simvastatin nanoparticles optimized Formulation (F8)

Figure 7: Comparission in-vitro dissolution profile F8 and marketed tablet (SIMVOFIX)

CONCLUSION:

Simvastatin an anti-lipidemic drug were formulated as nanoparticles an approach to increase in %drug Entrapment efficiency, improve the stability and better drug release there by its improve bioavailability. The formulation (F5) showed particle size 97.47nm, PDI as 0.333, zeta potential -30.60mV and %Entrapment efficiency 85.69%. All the formulations shown %drug release shows more 12hrs. The formulation, F8 showed the maximum 97.28% drug release. So, formulations F8 showed better sustain the drug release for 12hrs period of time as compared to other prepared formulations. Therefore polymeric nanoparticles can be good candidates to encapsulate Simvastatin for the lowering of cholesterol.

REFERENCES:

1. Shiva kumar yellanki, Sai manoj A and Mangilal T (2021) Preparation and in vitro evaluation of metoprolol-loaded bovine serum albumin nanoparticle. Asian journal Pharm Clin Res, 14(1), 213-217

2. Monowar Hussaini et al. (2020) Formulation and Evaluation of Ethambutol Polymeric Nanoparticles. Int J App Pharm, 12(4), 207-217.

3. Alam Zeba et al. (2019) Simvastatin- loaded solid lipid nanoparticles for enhanced anti-hyperlipidemic activity in hyperlipidemia animal model. International Journal of Pharmaceutics, 560, 136–143.

4. Vipan kumar kamboj and Prabhajar kumar verma (2018) Preparation and characterization of metformin loaded steric acid coupled F127 Nanoparticles. Asian J Phar Clin Res, 11(8), .212-2

5. Surendranath Betala1 et al. (2018) Formulation and evaluation of polymeric nanoparticles of an anti-hypertensive drug labetalol. Journal of Drug Delivery and Therapeutics, 8(6-s), 187-191.

6. Jin Sun a, d et al. (2017) The studies of PLGA nanoparticles loading atorvastatin calcium for oral administration in vitro and in vivo. asian journal of pharmaceutical sciences, 12, 285-291

7. N. Selvasudha et al. (2017) Optimization of formulation parameters and characterization of simvastatin loaded chitosan nanoparticles. Int.Jou.Adv.Res, 5(2), 2386-2400.

8. Shanmuganathan Seetharaman1 et al. (2016) Preparation and Evaluation of Cefixime Nanoparticles Prepared Using Fenugreek Seed Mucilage and Chitosan as Natural Polymers. IJPCR, 8(3)

9. K. Masilamanib et al. (2015) Development and Evaluation of Chitosan Based Polymeric Nanoparticles of an Antiulcer Drug Lansoprazole. Journal of Applied Pharmaceutical Science, 5 (04), 020-025

10. Vijaya Bhaskar N et al. (2015) Formulation and characterisation of Simvastatin nanoparticles loaded transdermal patch. Journal of Chemical and Pharmaceutical Research, 7(7), 1084-1093

11. P.Swathi et al. (2014) Formulation of ibuprofen loaded ethyl cellulose nanoparticles by nanoprecipitation technique. Asian.Jou.Pharm.Clin.Res, 7(3), 44-48

12. Tiruwa R (2015) A review on nanoparticles preparation and evaluation parameter. Indian Journal of Pharmaceutical and Biological Research. 4(2).27-31.

13. Pal SL, Jana U and manna Pk (2011) Nanoparticles: An overview of preparation and characterization. Journal of Applied Pharmaceutical Science. 1(6), 228-259.

14. Khosla G, Goswami L and kothiyal P (2012) Nanoparticles: A Novelistic Approach for CNS disorders. Journal of Advanced Pharmaceutical Sciences, 2(2), 220-259.

15. Lam P-L et al. (2017) Recent advances in green nanoparticulate systems for drug delivery: efficient delivery and safety concern. Nanomedicine. 12, 357–85.

16. Jahangirian H et al. (2017) A review of drug delivery systems based on nanotechnology and green chemistry: green nanomedicine. Int J Nanomed. 12, 2957.

17. Siddiqui AA, Iram F, Siddiqui S and Sahu K (2014) Role of natural products in drug discovery process. Int J Drug Dev Res. 6(2), 172–204

Received on 31.03.2023 Modified on 22.05.2023

Asian J. Pharm. Tech. 2023; 13(3):183-188.

DOI: 10.52711/2231-5713.2023.00033