FORMULATION AND CHARACTERIZATION OF SOLID LIPID NANOPARTICLES OF ATORVASTATIN CALCIUM 1
Mukesh Sharma1 , Atindra Shukla1 , Dinesh O. Shah1 , B. N. Suhagia2 , Tejal G Soni2 Shah-Schulman Center for Surface Science & Nanotechnology, Dharmsinh Desai University, Nadiad-387001 2 Faculty of Pharmacy, Dharmsinh Desai University, Nadiad– 387001, Gujarat 3 Department of Pharmaceutics and Pharmaceutical Technology, L. M. College of Pharmacy, Ahmedabad -380009 Email: Sharma.mukesh62@gmail.com
INTRODUCTION
Atorvastatin calcium is a selective, competitive inhibitor of 3-Hydroxy-3-methyl glutaryl coenzyme A reductase (HMG CoA) an enzyme catalyzing conversion of HMG CoA to mevalonate, the rate limiting step in cholesterol biosynthesis. Atorvastatin is used as an adjunct to dietary therapy to decrease elevated serum total cholesterol, LDL-cholesterol and elevated serum triglyceride concentrations. Solid lipid nanoparticles (SLNs) are a colloidal carrier system for controlled drug delivery. Nanostructures have the ability to protect drugs encapsulated within them from hydrolysis & environmental conditions, target the delivery of drug to various area of the body for prolonged release. As per several clinical research, SLN is found to be improve oral bioavailability of many drugs.
AIM OF WORK Atorvastatin calcium was selected for the SLNs due to following properties: Poorly water soluble Extensive first pass metabolism & high intestinal clearance Low oral bioavailability Light sensitive drug So the aim of present work was: To enhance drug absorption via improvement in dissolution & solubilization. To enhance drug absorption via bypass hepatic first pass metabolism. To increase the intestinal permeability. To protect the drug from environmental condition (light). To provide the controlled /sustained drug release.
ANALYTICAL METHODS 1. UV-VIS Spectroscopy: λmax for atorvastatin calcium was 243 nm in distilled water containing 0.5% Sodium lauryl stearate.
2. HPLC Method: Column: Octadecaylsilane RC 18bonded to porous silica (150×4.6 mm, 5µm) Mobile phase: 0.02 M Potassium hydrogen phosphate: methanol: acetonitrile (20:40:40) (pH 4.0) Internal standard solution (IS): Methaqualone (100µg/ ml) Volume of loop injector: 20µl Flow rate of mobile phase: 1 ml/min. Wavelength of detection: 246 nm.
RESULTS Formulation parameters optimizations; 1. Optimization of drug/ lipid ratio 2. Optimization of surfactant ratio 3. Optimization of stirring speed 4. Optimization of sonication time 5. Optimization of stirring time
Optimization using 32 full factorial design : Atorvastatin calcium = 50 mg X1 = Concentration of Glyceryl monostearate X2 = Concentration of Poloxamer 188 Batch
X1
(Mg) X2 (Mg) Particle size (d-nm)
PDI
Zeta potential % Entrapment % Drug release in (mV) Efficiency 24 hrs
F1
200
80
108.3±2.03
0.094±0.033
-27.2±4.98
69.81±1.52
72.49±1.23
F2
200
100
96.4±2.86
0.087±0.045
-28.1±4.57
68.10±2.01
75.30±2.05
F3
200
120
79.9±3.97
0.123±0.029
-34.1±4.78
65.16±1.76
78.21±1.60
F4
300
80
127.6±2.14
0.129±0.028
-26.1±4.68
76.36±2.04
65.58±2.84
F5
300
100
102.8±3.64
0.193±0.021
-28.4±3.87
74.90±1.65
68.15±1.38
F6
300
120
88.6±2.82
0.122±0.034
-33.7±4.56
70.53±1.56
69.91±2.71
F7
400
80
144.3±2.64
0.142±0.043
-25.2±4.32
81.30±1.21
59.18±1.69
F8
400
100
116.1±2.84
0.109±0.041
-31.0±4.61
79.63±1.72
61.52±0.94
F9
400
120
97.9±3.37
0.192±0.027
-33.9±4.56
73.42±1.83
64.57±1.32
Fig 2: Overlaid DSC thermogram:
Fig 4. Plasma concentration–time curve of ATC-SLNs & ATC soln
Fig 3: P-XRD pattern
Fig 5: Particle size of ATC-SLNs
Fig 6: In vitro release of ATC– SLNs
Cmax (ng/ml)
T max (h)
AUC (o-t) (ng/ml/h)
Relative bioavailability (%)
ATC solution
612±4.77
2±0.05
3292±25.62
100
Release mechanism and Selection of the best batch
ATC-SLNs
1023±6.16
2±1.68
9029±42.97
274.86
From the regression results the optimum formulation was found Batch F7 which contains poloxamer 188 (80 mg) glyceryl monostearate (400 mg). The % Entrapment Efficiency of batch F7 was found to be 81.30±1.21 % and % Drug release in 24 hrs 59.18±1.69%. So the batch F7 was selected for further characterization study.
Table 4: Pharmacokinetic parameters of ATC (SLNs and pure form) in rat plasma (n = 5) after oral administration (mean ± SD)
PDI = Polydispersity index
In present study, the effects of SLNs of atorvastatin calcium on serum lipid levels were evaluated in hyperlipidemic rat induced by poloxamer 407. Treatment of rat with SLNs of atorvastatin calcium significantly reduced the serum TG, TC, VLDL, LDL levels and significantly increased the serum HDL-C levels when compared to hyperlipidemic control rat . ATC-SLNs was found to be effective in significantly reducing serum TG and VLDL (p < 0.001) levels when compared to the P-407 induced hyperlipidemic control rat, after 12 and 24 h of treatment at a dose of 500 mg/ kg p.o. After 12 h of treatment, Atorvastatin calcium tablet form reduced serum TC and LDL-C levels but the results were not found to be significant but after 24 h of treatment, both the parameters were reduced significantly (p < 0.05).
2
DESIGN OF EXPERIMENT (3 FULL FACTORIAL)
SUMMARY AND CONCLUSION Fig 1: HPLC chromatogram of ATC loaded SLNs with IS from rat plasma.
METHOD
% Entrapment efficiency area:
Y1 = 75.1 + 5.5033X1 - 2.7683X2 - 1.34X1 2 1.754X22 - 1.29X1 X2 (R2 = 0.998325)
% Drug Release area:
Y2 = 67.74 - 6.969X1 + 2.4192X2 + 0.8357X1 2 + 0.1707X22 + 1.88X1 X2 (R2 = 0.99836)
STABILITY STUDY
Batch N o. = F7
Influence of storage condition and storage duration on particle size, PDI and residual drug content (%) of ATC-SLN.
Duration
Particle size d-nm.
(Days)
4±1°C
25±2°C
Initial
144.3±2.64
10
PDI
% Residual drug content
4±1°C
25±2°C
4±1°C
25±2°C
144.3±2.64
0.142±0.04
0.142±0.04
100
100
147.2±4.02
151.3±4.21
0.145±0.03
0.148±0.02
99.67±0.23
99.17±0.35
20
150.6±3.56
162.1±4.41
0.151±0.01
0.157±0.02
99.25±0.31
98.64±0.23
30
153.4±4.11
174.2±3.54
0.154±0.02
0.168±0.03
98.83±0.25
97.81±0.48
SLNs achieved higher drug incorporation and EE % of SLNs was more than 70 % and showed relative short-term stability as the leakage was very negligible after being stored for one month. Physicochemical characterization revealed that the prepared drug loaded SLNs were of spherical shape, homogenously distributed and amorphous. Solid lipid nanoparticles follows weibull model because good correlation coefficient obtained by this model around 0.9972. The result indicates no significant change in the % drug content after exposure of UVa light. Hence it is concluded that Atorvastatin calcium loaded SLNs were stable during the photostability stable. The pharmacokinetic parameters of optimized SLNs in wistar rats, obtained using graph pad software revealed 2.74 fold in bioavailability as compared to atorvastatin calcium (pure). SLNs are able to deliver bioactive molecules, protein, genes, anticancer gene therapy and vaccine for improving their bioavailability and prevent enzymatic degradation . Smart SLNs as the new generation offer much more flexibility in drug loading, modulation of release and improved performance in producing final dosage.
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