Lipid Nanoparticulate system of Simvastatin- A method for solubility enhancement
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1 Research Article ISSN: Akash C et al. / Journal of Pharmacy Research 2017,11(6), Available online through Lipid Nanoparticulate system of Simvastatin- A method for solubility enhancement Akash C * and Preethi Sudheer Department of Pharmaceutics, Krupanidhi College of Pharmacy, #12/1, Sarjapur Road, Chikkabellandur, Carmelaram Post, Varthur Hobli, Bangalore, Karnataka ,India. Received on: ; Revised on: ; Accepted on: ABSTRACT Aim: In the present work, solid lipid nanoparticles (SLNs) of simvastatin was formulated with an aim to increase the solubility, rate of dissolution and drug stability. Method: The formulations were prepared by hot homogenization technique using lipids such as stearic acid, glycerol monostearate and tween 20 as a surfactant. The coarse emulsion was homogenized by using Polytron PT 1600 E homogenizer at rpm. Nanoparticles obtained were characterized for particle size analysis, zeta potential, SEM, DSC, FTIR and also analyzed for drug content and in vitro drug release profile. Results: Out of 12 formulations, selected 5 formulations (F1 to F5) were found to be free flowing. The drug content of the selected formulations was ranging between ± 1.02% to ± 1.03%. The average particle size and zeta potential of selected formulas, F2 was found to be 795.7nm and -33.7mV and F4 was found to be 369nm and -34mV respectively. The comparative release profile of F1 to F5 was found to be in between ± 0.004% to ± 0.005% in comparison to pure drug profile of ± 0.003% at 24hr release study. The DSC thermogram indicates the melting point of the drug was decreased from C to C, due to molecular dispersion of drug in lipids. Conclusion: Solid lipid nanoparticles can be alternate stable cost effective approach for improving dissolution rate of poorly soluble drugs. KEY WORDS: Simvastatin, Solid lipid nanoparticles (SLN), Differential scanning calorimetry (DSC), Scanning electron microscopy (SEM), Zeta potential, Particle size analysis, Fourier transform infrared spectroscopy (FTIR). 1. INTRODUCTION: Oral route is the most common and preferred for drug administration; solubility is a significant physicochemical factor affecting absorption of drug and its therapeutic effectiveness. The low dissolution rate and low solubility of drug substances in water or aqueous G.I.T fluid frequently leads to inadequate bioavailability. In order to overcome the challenges associated with oral drug delivery, nanoparticles (NPs) are considered as alternatives to various conventional drug delivery techniques. [1, 2] Nanoparticles made from lipids provide an attractive means as a novel colloidal drug carrier due to the presence of natural lipids. [3] SLNs are submicron colloidal carriers ( nm) which are composed of physiological lipids, dispersed in water or in aqueous surfactant solution. Generally they are made up of solid hydrophobic core having a monolayer of phospholipid coating. [4,5] *Corresponding author Akash C Department of Pharmaceutics, Krupanidhi College of Pharmacy, Chikkabellandur Village, Carmelaram Post, Varthur Hobli, Bangalore , Karnataka, India. Simvastatin is a typical hypolipidemic (anti-hyperlipidemic) agent classified under BCS Class II, the drug having high permeability but low solubility. Simvastatin is well absorbed from the gastrointestinal tract but is highly extracted by the liver and only 7% of the dose reaches the general circulation. It possesses biological half life of 2 hours. 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. Statins are more effective than other lipid regulating drugs at lowering LDL cholesterol concentration, but they are less effective than the fibrates in reducing triglyceride concentration. [6] So this study focuses on converting simvastatin in form of solid lipid nanoparticles which could improve the solubility and dissolution profile of BCS class II drug. 2. MATERIALS AND METHODS: The pure drug (simvastatin) was gifted from Watson Pharma Ltd. Stearic acid was supplied by Loba Chemie Pvt Ltd, Mumbai. Glycerol monostearate were obtained from Quarigens fine chem, Mumbai. Tween 20 was obtained from Merck specialities Pvt Ltd, Mumbai. All solvents and reagents were of analytical grade.
2 Akash C et al. / Journal of Pharmacy Research 2017,11(6), 2.1. Saturation solubility studies: A saturated solution of drug in melted lipids was prepared by dissolving drug in 250mg of lipids/lipids combinations. Approximately 100 mg drug lipid melt was dissolved in methanol and diluted to Beer s range; UV spectral analysis was carried out and the absorbance was noted at 237.5nm Preparation of simvastatin loaded nanoparticles: Hot homogenization technique: Required quantities of lipids were melted; the drug was dissolved in the melted lipids. The drug loaded lipid was added to the hot aqueous surfactant (tween 20) under mechanical stirring at 700 rpm for 10min. The coarse emulsion obtained was homogenized using Polytron PT 1600E homogenizer. The o/w nanoemulsion was cooled to room temperature, resulted in SLNs. The formulations are given in table 1 and selected formulations based on free flowing properties are given in table 2. [7,8] Table 1: Formulation table concentration of 40μg/ml. Absorbance of the solution was measured at 237.5nm spectrophotometrically. 50mg of blank formulation treated in the same way was used as blank for estimation of drug Differential scanning calorimetry (DSC): This study was carried out for pure drug and formulations using differential scanning calorimeter (MDSC 2920 Mettler-Toledo, USA). For this, the powder sample (1to5mg) was packed in aluminum lid was placed on the top of the pan and was crimped. The crimped pan was placed in the sample cell along with an empty pan as a reference. Temperature was increased from 0 C till the powder sample undergoes decomposition at a rate of 5 C/min. The thermal cell was purged with dry nitrogen at 50ml/min. [9] 3.3. FTIR spectroscopy: FTIR spectra of simvastatin and the formulations were recorded in KBr medium pellets using FTIR spectrophotometer. The scan was performed in the range of cm -1. [10] 3.4. Particle size distribution and zeta potential measurement: Particle size analysis was performed by dynamic light scattering (DLS), also known as photon correlation spectroscopy (PCS), using a Malvern Zetasizer Nano ZS (Malvern Instruments, UK). Prior to the measurements, the samples were diluted using ultra-purified water to yield a suitable scattering intensity. DLS data were analyzed at 25 C using the general purpose mode. DLS yields the hydrodynamic diameter (Z-ave and polydispersity index PI) as a measure of width of particle size distribution. [11] Table 2: Selected formulations based on free flowing nature The zeta potential was determined by the measurement of the electrophoretic mobility using Malvern Zetasizer Nano ZS (Malvern instruments, UK). The field strength applied was 20V/cm. To avoid fluctuation occurring due to variations in the conductivity of the water, range from 1 to 10μS/cm. The conductivity of the water was adjusted to 50μS/cm using 0.9% (w/v) sodium chloride solution. Each sample was measured three times, mean value is determined. [12,13] 3. EVALUATION OF SOLID LIPID NANOPARTICLES: 3.5. Scanning electron microscopy (SEM): The morphology of NPs was determined by using scanning electron microscope (JSM 840 A, Jeol, Japan) operating at 15 KV. The samples were mounted on a metal stub with a double adhesive tape and coated with platinum/palladium alloy under vacuum. [14] 3.1. Drug content: 3.6 In vitro release of simvastatin in phosphate buffer ph % The formulation equivalent to 50 mg of drug was taken and dissolved in SLS: methanol; volume was made up to 100ml with methanol. 1ml of the above solution was diluted suitably with methanol to get a Formulations equivalent to 20 mg of drug was taken and studies
3 Akash C et al. / Journal of Pharmacy Research 2017,11(6), were carried out using type II apparatus (paddle type). SLN formulations were placed in the dissolution flask containing 900ml of phosphate buffer ph 7 containing 0.1% SLS as the dissolution medium. The temperature of the medium maintained at 37±0.5 C, the samples were withdrawn at predetermined time intervals for 24 hr. The absorbance was noted and % CDR was calculated. Table 3: Percentage drug content 3.7. Kinetics of drug release: The data obtained were checked for its suitability to be fitted into Zero order, First order, Higuchi model, and Korsmeyer-Peppas model. Based on R 2 value, the best fit model was selected. [15] 3.8. Short term stability studies: Short term stability studies were carried out at 40 C ± 2 C /75% RH ± 5% RH for 1 month to determine physical and chemical stability. 4. RESULTS AND DISCUSSION: 4.1. Saturation solubility studies of simvastatin in lipids: Among the lipids used such as stearic acid 12.00mg/250mg, glycerol monostearate 8.255mg/250mg and combination of stearic acid and glycerol monostearate 12.11mg/250mg, combination of stearic acid and glycerol monostearate showed highest saturation solubility of drug when compared to other lipids as shown in figure 1. Saturation solubility of simvastatin in lipids (w/w)/250mg Saturation solubility of simvastatin in lipids (w/w) Stearic acid Glycerol monostearate Stearic acid+glycerol monostearate Lipids Figure 1: Saturation solubility of simvastatin in lipids (w/w)/250mg (C) 4.2. Characterization of solid lipid nanoparticles: Drug content (%) of selected formulations was found to be 61.56±1.02%, 65.50±1.00%, 77.80±1.01%, 78.34±1.03% & 77.60±1.05%, for F1, F2, F3, F4, F5 formulations, here F4 formulation showed better drug content when compared to other formulation shown in table 3. DSC thermogram of simvastatin exhibits an endothermic peak at < C as shown in figure 2. In case F2 and F4 formulation the endothermic peak shifted towards lower temperature i.e C and C respectively. Shifting of the peaks might have occurred as the drug is molecularly dispersed in lipids. Figure 2: DSC thermogram of pure drug simvastatin, F2 formulation, (C) F4 formulation
4 Akash C et al. / Journal of Pharmacy Research 2017,11(6), FTIR spectra (figure 3) of simvastatin and formulations reveal the characteristic absorption peaks of simvastatin at cm -1 (OH strong, free, stretch), cm -1 (aromatic C=C Stretch, medium weak, multiple bands), cm -1 (methyl C-H symmetric stretch; methylene C-H asymmetric stretch), cm -1 (secondary alcohol C-O stretch). The FTIR study revealed the characteristic peaks of simvastatin were also present in the formulations F2 and F4. It showed that there is no interaction between the drug and excipients. (D) Figure 3: FTIR spectrum of simvastatin pure drug, physical mixture, (C) F2 formulation, (D) F4 formulation 4.3. Particle size distribution and zeta potential measurement: The particle size analysis data (figure 4) of formulations F2 and F4 obtained by Malvern zetasizer indicates the average particle size which was found to be 795.7nm with poly dispersity index of and 369nm with poly dispersity index of respectively. The poly dispersity index less than 1 indicates that the particle size distribution is uniform. The particles fell in an acceptable nanometer range. (C) Figure 4: Particle size distribution of F2 formulation, F4 formulation Zeta potential of the SLNs (figure 5) was measured by Malvern zetasizer. The zetapotential value greater than -30mV less than -40mV is found to be stable (Riddick TM 1968). The zeta potential of F2 and F4 was found to be and -34mV respectively. This indicates that the formulations are stable.
5 Akash C et al. / Journal of Pharmacy Research 2017,11(6), (C) Figure 6: SEM of F2 formulation, F4 formulation, (C) pure drug simvastatin 4.5 In vitro release of simvastatin in phosphate buffer ph % SLS: The in vitro release study of formulations F1 to F5 and pure drug in phosphate buffer ph % SLS revealed that the formulations exhibited a drug release of 92.05± to 97.65± % in comparison to pure drug simvastatin which showed release rate of ± % in a 24 hr release study as shown in figure 7. Figure 5: Zetapotential of F2 formulation, F4 formulation 4.4. Scanning electron microscopy (SEM): Scanning electron microscopy (SEM) studies were carried out for selected formulations F2, F4 and pure drug shown in figure 6. It was seen that the particles were discrete without much clumping. The surfaces were appeared to be uneven. Figure 7: In vitro release of simvastatin in phosphate buffer ph % SLS 4.6. kinetics of drug release: The release data obtained from selected F4 formulation is subjected for curve fitting shown in table 4. The release data was best fitted to first order release kinetics. Table 4: Release kinetics of simvastatin loaded SLNs (F4 formulation)
6 Akash C et al. / Journal of Pharmacy Research 2017,11(6), 4.7. Short term stability studies: The selected F4 formulation are subjected to short term stability studies for a period of 1 month at 40 C ± 2 C /75% ± 5% RH change in the release profile was analyzed as shown in figure 8. The results revealed that there was no significant change in the release profile. Figure 8: Release profile of selected F4 formulation after 1 month 5. CONCLUSION: In present study simvastatin loaded solid lipid nanoparticles were formulated with an aim to increase the solubility, release rate. Solid lipid nanoparticles were formulated by hot homogenization technique. This technique was found to be successful in preparing free flowing discrete nanosized particles, which was evident from particle size distribution data and zeta potential. In DSC thermogram, a melting point change from C (pure drug) to C (SLN) was observed, which indicates the molecular dispersion of drug in lipids resulted in increased rate of drug release. The in vitro drug release was found to be higher from solid lipid nanoparticles and F4 formulation showed high release rate compared to pure drug (simvastatin). The formulation can be economically manufactured from relatively cheap raw materials like stearic acid, GMS (glycerol monostearate), tween 20 etc. The developed SLNs offer the advantage of nanorange particle with uniform particle size distribution and higher stability. ACKNOWLEDGEMENTS: It is with great pleasure that I place on record a deep sense of gratitude and heartfelt thanks to my research guide Preethi Sudheer, for her help, support throughout the progress of this work and I also thank Watson Pharma Ltd for providing gift sample of simvastatin (Pure drug). REFERENCES: 1. Kadam SV, Shinkar DM, Saudagar RB. Review on solubility enhancement techniques. Int J Pharm Bio Sci. 2013; 3(3): Charade VV, Chaudhar PD. Development and evaluation of self emulsifying drug delivery system for lornoxicam. Int J Res Dev Pharm L Sci 2013; 2(4): Phaechamud T, Tuntarawongsa S. Clotrimazole soft lozenges fabricated with melting and mold technique. Res J Pharm Bio Chem Sci. 2010; 1(4): Subedi RK, Kang KW, Choi HK. Preparation and characterization of solid lipid nanoparticles loaded with doxorubicin. Eur J Pharm Sci. 2009; 37(3): Vyas SP, Khar RK. Targeted & controlled drug delivery novel carrier system. 1st edi. 2002; p Drug bank. Simvastatin [Internet] [cited 2017 Mar 11]. Available 7. Parhi R, Suresh P. Production of solid lipid nanoparticles drug loading and release mechanism. J Chem Pharm Res. 2010; 2(1): Jain N, Jain R, Thakur N, Gupta BP, Jain DK, Banveer J, Jain S, et al,. Nanotechnology: a safe and effective drug delivery system. Asian J Pharm Clin Res. 2010; 3: Castelli F, Puglia C, Sarpietro MG, Rizza L, Bonina F, et al,. Characterization of indomethacin loaded lipid nanoparticles by differential scanning calorimetry. Int J Pharm. 2005; 304(1): Soltani S, Zakeri-Milani P, Barzegar-Jalali M, Jelvehgari M, et al,. Fabrication and invitro evaluation of ketotifen fumarate loaded PLGA nanoparticles as a sustained delivery system. Iranian J Pharm Res. 2017; 16(1): Liu W, Hu M, Liu W, Xue C, Xu H, Yang X, et al,. Investigation of the carbopol gel of solid lipid nanoparticles for the transdermal iontophoretic delivery of triamcinolone acetonide acetate. Int J Pharm. 2008; 364(1): Riddick TM. Control of colloid stability through zeta potential with a closing chapter on its relationship to cardiovascular disease (vol 1). Zeta-meter, Inc Yadav NE, Khatak SU, Sara UV. Solid lipid nanoparticles-a review. Int J Appl Pharm. 2013; 5(2): Ramteke KH, Joshi SA, Dhole SN. Solid lipid nanoparticle: a review. IOSR J Pharm. 2012; 2(6): Lokhandwala H, Deshpande A, Deshpande SH. Kinetic modeling and dissolution profiles comparison: an overview. Int J Pharm Bio Sci. 2013; 4(1): Source of support: Nil; Conflict of interest: None Declared
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