Design, characterization and optimization of Rosuvastatin calcium nanosponges loaded transdermal patch

 

Satyalakshmi S, Karthik D*, Anusha J, Kamala Kumari PV, Srinivasa Rao Y, Rama Rao B

Department of Pharmaceutics, Vignan Institute of Pharmaceutical Technology,

Near VSEZ, Duvvada, Visakahapatnam-530049.

 

ABSTRACT:

Rosuvastatin calcium is a low solubility containing anti-lipidemic drug and provides only 20% of oral bioavailability. Following a different route of administration can overcome the first pass metabolism and may improve the bioavailability. Therefore the present study aimed to design and characterize a transdermal patch containing rosuvastatn calcium nanospoges (RST-NSP) to enhance the drug dissolution. Emulsion solvent diffusion method was used in the preparation of (RST-NSP) employing β-cyclodextrin and poly vinyl alcohol as solubility enhancers. Design Expert^® 13 was employed to design twenty formulations in which concentration of β-Cyclodextrin in mg (A), ethylcellulose in mg (B) and reaction time in hrs (C) were taken as independent factors, where as entrapment efficiency (%) and particle size were considered two responses. Triple fold increase in solubility, 86 % of entrapment efficiency, 200 nm particle size containing NSP were loaded in transdermal patch by solvent casting method, with hydroxypropyl methyl cellulose and cabopol as polymers in 3:1 ratio. SEM analysis showed the uniform distribution of nanosponges and its morphology. The transdermal patch loaded NSP released the drug up to 10 h.  Kinetic  modeling  on  release  data  showed  that  the  best  fitted model  was  Higuchi  model  and release  mechanism was by Fickian diffusion.

 

 

 

INTRODUCTION: 

Rosuvastatin is a synthetic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor for the treatment of dyslipidemia¹. Rosuvastatin calcium (RSC) selectively inhibits HMG-CoA reductase, which catalyzes the conversion of HMG-CoA to mevalonate, a precursor of cholesterol. The antilipidemic activity is resulted by decrease in hepatic cholesterol levels and increase in uptake of LDL cholesterol². Bioavailability of RSC after oral administration is about 20%. RSC as conventional oral dosage form has major shortcoming in the therapeutic application and efficacy due to its very low aqueous solubility and first-pass metabolism. Log P of RSC is 2.6³. To overcome poor aqueous solubility, hepatic first-pass metabolism and to enhance drug dissolution, nanocarriers are developed. These are gaining tremendous interest and have shown remarkable advantages over conventional drug delivery systems.

The nanocarriers can provide the reduced dosing frequency, increased selectivity, enhanced bioavailability and reduced side effects^4-6. Nanosponges are the novel class of encapsulating nanoparticles, exhibiting promising potential in controlled drug delivery especially topical formulations.^7,8 Among the topical delivery, transdermal delivery systems provide various distinct advantages.^9,10 In the recent years, TDDS has become one of the most innovative topics for the delivery of drugs.¹¹,¹² The nanosize containing nanosponge drug particles can overcome the permeability and solubility limitations. The present study aims to design NSP and formulate transdermal patch containing drug loaded nanosponges to improve solubility and to avoid first pass metabolism.

 

MATERIAL AND METHODS:

RSC was gifted from Aspire Chem Pvt Ltd Mumbai, India, β-cyclodextrin purchased from Jay Chem marketing Mumbai, India. All the remaining chemicals used in the fabrication of nanosponges and transdermal patch were purchased from Molychem, Mumbai, India.

Preparation of rosuvastatin calcium nanosponges (RSC-NS):

Nanosponges were prepared by emulsion solvent diffusion technology by following the formulations given in Table 1. The disperse phase consisting of 10 mg of RSC and specified quantity of ethylcellulose dissolved in 5 ml of dichloromethane and sonicated in a sonicator (Spinotech Pvt Ltd) for 10 min and was slowly added to different concentrations of β-cyclodextrin (β-CD) )in 10 ml of aqueous continuous phase. The mixture was stirred at 1000 rpm on a magnetic stirrer (Remi Equipments Pvt Ltd Mumbai, India) at different time intervals. RSC-NS were collected by vacuum filtration and dried in an oven at 40^oC for 24 h¹³.

 

Table 1: Composition of rosuvastatin calcium nanosponges

Formulation	F1	F2	F3	F4	F5	F6
Drug (mg)	10	10	10	10	10	10
EC (mg)	100	150	200	100	150	200
β-CD (mg)	-	-	-	150	150	150
PVA (mg)	150	150	150	-	-	-
DCM (mL)	5	5	5	5	5	5
D W(mL)	10	10	10	10	10	10

 

Experimental design:

Design-Expert version 12 (Stat-ease Inc) was applied to optimize concentration of Beta- cyclodextrin (β-CD) (A), ethylcellulose (EC) (B), reaction time (RT) (C) considering as independent variables using 2³ full factorial design; the two responses entrapment efficiency (EE) (%), particle size (PS) (nm) were considered as dependent variables.

 

Characterization of RSC-NSP:

Percentage yield, entrapment efficiency, saturated solubility, scanning electron microscopy, were conducted by following the standard procedures given in the literature^14,15 to characterize the fabricated NSP. FTIR (Bruker FTIR, Invinio, Japan) spectra were done for the prepared formulations.

 

 

In vitro dissolution testing:

The USP dissolution test apparatus (model- DS 8000) type 2 (paddle) was employed for in vitro dissolution study¹⁶.

 

Preparation of transdermal patch-Solvent casting method and characterization:

In this method polymers like hydroxypropylmethyl cellulose (HPMC), EC, and carbopol were taken in different ratios as shown in Table 2. These polymers were dissolved in an organic solvent, water in 1:1 ratio. RC-NSP loaded in this mixture. Polyethylene glycol (PEG) was added to the polymer solution, stirred for 30 min by adding few drops of glycerin into it. Now the prepared solution was casted into petridish and dried at room temperature for 48 h. The formulated nanosponges were evaluated for physical appearance, thickness of the patch, uniformity of weight, flatness study, folding endurance, percentage moisture absorption, percentage moisture content, drug content and in-vitro drug release studies. The in-vitro drug release data was fitted to Higuchi, first-order, zero-order kinetics equations and to general exponential function: Mt/M∞= ktn, where Mt/M∞ represents solvent fractional uptake (or solute release) normalized regarding to conditions of equilibrium.¹⁷

 

RESULTS AND DISCUSSION:

The % of EE was in the range of 34 % - 88 %. Equation developed for response 1 (EE) was given below.

 

Entrapment efficiency (%) = +84.05-0.5458 A-1.27 B -0.2163 C-8.50 AB-7.25 AC+3.25 BC -16.27 A² -2.83 B² -10.79 C².

 

It was evident that increase in the polymer and cross-linker concentration increased the EE (%) of RSC-NSP. The model terms AB, AC, A² and C² were significant with P-value <0.005. β-CD and EC; β-CD and reaction time; has positive influence on EE (%) (figure.1).

 

Table 2: Composition of transdermal patch

Formulations	S1	S2	S3	S4	S5	S6	S7	S8
Pure RSC (mg)		10
RSC-NS (mg)	120	-	120	120	120	120	120	120
HPMC (g)	0.3	0.3	0.35	0.4	0.45	0.3	0.3	0.3
Carbopol (g)	0.1	0.15	0.2	0.25	0.3	-	-	-
EC (g)	-	-	-	-	-	0.5	1.0	1.5
Ethanol (mL)	10	10	10	10	10	10	-	-
PEG (mL)	4	4	4	4	4	4	4	4
Glycerin (mL)	0.1	0.1	0.1	0.1	0.1	0.1	0.1	0.1
Butanol (mL)	-	-	-	-	-	-	10	-
Isopropyl alcohol (mL)	-	-	-	-	-	-	-	10
Distilled water (mL)	10	10	10	10	10	10	10	10

 

 

 

 

Figure 1: Contour plots (A), response surface curve (B) shows the impact of β-CD and RT on % of EE; Contour plots (C), response surface curve (D) shows the impact of β-CD and EC on % of EE

 

Table 3: Describes the ANOVA of response 1 (% of EE)

Source	Sum of Squares	df	Mean Square	F-value	p-value
Model	6148.76	9	683.20	19.76	< 0.0001	Significant
A-β-cyclodextrin	4.07	1	4.07	0.1177	0.7387
B-Ethylcellulose	22.08	1	22.08	0.6384	0.4428
C-Reaction time	0.6392	1	0.6392	0.0185	0.8945
AB	578.00	1	578.00	16.72	0.0022
AC	420.50	1	420.50	12.16	0.0059
BC	84.50	1	84.50	2.44	0.1491
A²	3812.77	1	3812.77	110.26	< 0.0001
B²	115.46	1	115.46	3.34	0.0976
C²	1676.42	1	1676.42	48.48	< 0.0001
Residual	345.79	10	34.58
Lack of Fit	257.79	5	51.56	2.93	0.1316	not significant
Pure Error	88.00	5	17.60
Cor Total	6494.55	19

 

The impact of process parameters were demonstrated by analysis of variance (ANOVA), regression coefficient-P and F values were shown in Table 3.

 

Smaller particle size enhances the drug release because of its larger interfacial area for diffusion of the drug. Particle size of all suggested trial runs was assessed using size analyzer and obtained data were fed to DoE. Numerical and graphical analysis of the data was done via software. The derived equation from the best fit mathematical model that can be related to the response R2 and factors (A, B, and C) was given below

 

Particle size = +205.49 +5.77 A-0.4027 B+14.79 C +23.13 AB+4.13 AC+3.63 BC+20.29 A² +16.76 B²-2.33 C². It was clear from the polynomial equation, that the concentration of polymers and cross-linkers had direct influence on NSP particle size (R2) whereas, on the other side the rotation time has less impact on particle size. The particle size of the RSC-NSP was in the range of 186-287 nm this was because of the increased concentration of polymer and cross-linkers in the formulation. ANOVA of equation has shown model F value 14.13 and model terms AB, A², B² were significant model terms with P-value <0.005; indicating the significance of the model (Table 4). The Lack of Fit F-value of 2.77 implies the Lack of Fit is not significant relative to the pure error. There is a 14.4 % chance that a Lack of Fit F-value this large could occur due to noise. Non-significant lack of fit is good. The predicted R² 0.8713 was close to the adjusted R² 0.8988.

 

Table 4: Describes the ANOVA of response 2 (Particle size in nm)

Source	Sum of Squares	df	Mean Square	F-value	p-value
Model	17535.90	9	1948.43	14.13	0.0001	significant
A-β-cyclodextrin	455.41	1	455.41	3.30	0.0093
B-Ethylcellulose	2.21	1	2.21	0.0161	0.0017
C-Reaction time	2986.43	1	2986.43	21.65	0.1029
AB	4278.13	1	4278.13	31.02	0.0002
AC	136.13	1	136.13	0.9869	0.3439
BC	105.13	1	105.13	0.7622	0.4031
A²	5935.26	1	5935.26	43.03	< 0.0001
B²	4047.37	1	4047.37	29.34	0.0003
C²	78.46	1	78.46	0.5689	0.4681
Residual	1379.30	10	137.93
Lack of Fit	1013.30	5	202.66	2.77	0.1440	not significant
Pure Error	366.00	5	73.20
Cor Total	18915.20	19

 

Percentage yield values of nanosponges were within the range of 62.47-90.32% and best for F4. The entrapment efficiency (%) was best for F4 formulation. Low entrapment has found due to the sticky nature of ethyl cellulose polymer, and its low solubility in the aqueous phase.¹⁸ Reham et al., 2020 was optimized nanosponges formulation to get optimized EE and PS in the formulation of hydrogels loaded NSP.¹⁹ The solubility of prepared nanosponges was improved when compared to pure drug (2.10 ±0.2 g/100 mL). Three fold of enhanced solubility was obtained with F4 formulation (6.30±0.33 g/100 mL)²⁰. The spherical shaped numerous RSC-NS were identified by SEM analysis and the characteristic porous nature of the nanosponges was shown in figure. 2.

 

 

Figure 2: SEM photograph of RSC-NS

 

 

 

 

Figure 3: FTIR spectra A: RSC B: Optimized formulation (F4)

FTIR spectra of pure RSC demonstrated the characteristic absorption peaks of 2900 cm for O-H stretching, at 1370 cm for aromatic N-O stretching, 1200 cm for C-O stretching, 800 cm for C=C stretching and 510 cm for C=I stretching, showed almost similar absorption peaks indicate good compatibility with polymers (figure. 3A, 3B). F4 was selected as the optimized formulation based on evaluated parametrs and formulated into transdermal patch.Formulated transdermal patches were evaluated for thickness, uniformity of weight, flatness, folding endurance, % moisture absorption and % moisture content. All the evaluated parameters were with in the limit. In vitro release studies of RSC-NS in transdermal patch formulations were carried out in triplicate. After 10 h, the release was found to be 98.4±0.22, 80.45±0.14, 87.64±0.16, 84.98±0.24, 83.74±0.22, 87.14±0.11, 84.18±0.45, 79.91±0.61 for the formulations S1, S2, S3, S4, S5, S6, S7, and S8 respectively. In S1-S8 (except S2), 120 mg of nanosponges equivalent to 10 mg of drug and in S2, 10 mg of RSC pure drug was used and remaining excipients were same in all the formulations. S1 released 98.4% at the end of 10^th h but S2 released only 80.45% (even after addition of 1% SLS in the dissolution medium to solubilise the released drug). RSC in nanosponges have increased solubility than the pure drug because of the soluble complex formation with the cross linker β-CD. ²¹This may be the reason in the enhancement of dissolution. Obtained in vitro release data was fitted in various kinetic models such as zero order, first order, Higuchi model and Korsmeyer-Peppas model. All the formulations except S3, S8 followed first order release. S1 &S2 drug release mechanism was higuchi while other formulations were korsemeyer peppa’s so; the release mechanism for most of the formulations was erosion than diffusion. All ‘n’ values were in between 0.5 and 1 indicates the diffusion process in controlling release kinetics (non fickian anomalous or first order release). The prepared transdermal patches with different polymer concentrations were smooth, opaque, flexible and uniform. The thickness of the films varied from 0.230 to 0.834 mm and highest thickness was found to be 0.834 mm for S8, and lowest was of S1. From these values, it was observed that the thickness of the polymer depends on the solubility and concentration of the polymer. As the solubility decreases and concentration increases would increase the thickness of the patch. It infers that usage of the competent polymer is the prerequisite step to prepare a patch of optimum thickness, which can retard the release of drug from the patch. Weight of the formulated transdermal patches varies from 247 mg to 366 mg. Flatness of the all prepared transdermal patches was 100%. The folding endurance values from 145 to 169 revealed that the prepared patches were having good mechanical pressure and good flexibility. The folding endurance values of all the patches were found satisfactory. The moisture content in the patches ranged from 3.02±0.01 to 4.82±0.02%. The moisture content in the formulations was found to be increased by increase in the concentration of HPMC and EC. The moisture uptake in the patches ranged from 5.65±0.01 to 7.78±0.01%. The drug content ranged from 66.4 to 81.98%. All formulations were acceptable with regard to RSC content. The in vitro release profile of RSC transdermal patches (S1) could be best expressed by zero order kinetic model, as the plot showed the highest linearity (R²=0.98). The release exponent (n) value 0.62 indicates that the release from transdermal patch followed fickian release i.e., release always associated with diffusion mechanism.

 

CONCLUSION:

Rosuvastatin calcium nanosponges exhibited excellent solubility compared to pure drug. All the formulated nanosponges released more than 76% of the drug at the end of 30 min. 2³ full factorial design was employed where as β-Cyclodextrin, ethylcellulose and reaction time in hrs were taken as independent factors, entrapment efficiency (%) and particle size were considered two responses. 150 mg of β-CD, 375 mg of EC, and 1.5 hrs of RT were employed to achieve 86 % of EE and 200 nm of PS. Extension of drug release for 10 h reduces the dose; avoid the frequency of administration and systemic side effects. Hence the development of transdermal patch containing nanosponges of RSC is considered ideal and effective way of treatment in the management of cardiovascular diseases caused by lipidemia.

 

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Accepted on 27.12.2023           © RJPT All right reserved

Research J. Pharm. and Tech 2024; 17(4):1753-1757.