Preparation of FMM of Nimodipine
Melt granulation technique was adopted for preparing the solid dispersion of Nimodipine and lipophilic carrier such as COM, G43/01, G39/01 and PRE. The lipophilic carrier was heated on a water bath in a temperature range of 50 to 60 °C with continuous stirring. To the molten lipid dispersion, Nimodipine powder was added and stirred for 10 min. The SYL in 1:1 ratio with lipophilic carrier was added to the dispersion with continued mixing for 10 min in order to solidify the molten mass. Then, solid dispersion was sieved through mesh no. # 40. The resultant granules were compressed into 3-mm circular minitablets and were filled into a gelatine capsule of size ‘0’ [17].
Compatibility studies using differential scanning calorimeter
The pure drug Nimodipine and its solid dispersions with carriers were examined for compatibility study employing DSC (Shimazu Ltd., Japan). All the accurately weighed samples were placed in sealed aluminum pans before placing the pan on sample holder at a scanning rate of 10 °C/min from room temperature up to 220 °C. An empty aluminum pan was used as a reference. The DSC curve was plotted over a temperature range in order to predict the thermodynamic compatibility [18].
Micromeritic properties of granules
The pure drug Nimodipine and solid dispersion granules of all the prepared formulations were characterised for various micromeritic properties. The bulk density, tap density, angle of repose, Carr’s Index, and Hausner’s ratio of all the formulations were determined as per standard procedure [19].
In vitro buoyancy studies
Measurement of floating lag time
The FLT was determined by conducting in vitro buoyancy studies for each of the formulation. The required number of FMM was placed in a beaker containing 100 mL of 0.1 N HCl. FLT may be defined as the time required for each of the minitablets to start floating on the surface of the dissolution media [20].
Measurement of floating time
FMM tablets were placed in USP dissolution apparatus-II containing 900 mL of 0.1 N HCl to determine FT of each formulation. FT is the total time or duration of floating when a dosage form is placed in a dissolution media [21].
Evaluation of post-compression parameters
The compressed minitablets were subjected to various quality control tests such as weight variation, hardness, friability, disintegration and drug content as per the standard procedure [22]. Weight variation was determined by weighing 20 minitablets individually, hardness was determined by taking 6 tablets from each formulation using a digital hardness tester (Electro lab Ltd., India), friability was determined taking minitablets equivalent to 6.5 g in a Roche® friabilator (Electrolab Pvt. Ltd., India), which was rotated for 4 min at 25 rpm. The disintegration test was performed for 1 h in 0.1 N HCl by taking 6 minitablets. Drug content was determined by taking ten weighed minitablets from each formulation and finely powdered. The powder equivalent to 25 mg of Nimodipine was weighed and taken in a 25-ml volumetric flask, extracted with 0.1 N HCl. The mixture was then filtered and 1 ml of the filtrate was suitably diluted and analysed at 350 nm spectrophotometrically.
In vitro dissolution studies
The in vitro dissolution studies of different FMM were conducted in a USP dissolution apparatus-II (paddle method with a paddle speed of 50 rpm) taking 900 mL 0.1 N HCl as dissolution medium maintained at 37 ± 0.5 °C. A total of 5 mL sample was withdrawn at regular time intervals up to 12 h (0.5, 1, 1.5, 2, 4, 6, 8, 10 and 12 h) and were analysed using UV-spectrophotometer at 354 nm. The kinetics of drug release were analysed from the dissolution data by using zero-order, first order, Higuchi, and Korsmeyer-Peppas model [23, 24].
Zero order equation is Q = K0t.
First order equation is ln (100 − Q) = ln 100 − K1t
Higuchi Equation is Q = K2 √ t
Korsmeyer Peppa’s equation is \( \log \frac{M_t}{M_{\infty }}=\log K+n\log t \)
where Q is the amount of drug released at time t, K0 zero order release rate constant, K1 is first order release rate constant, the time, K2 diffusion rate constant, is the amount of drug released at time t, Mt is the amount of drug released at time t, M∞ is the amount of drug released at infinite time, initial amount of the drug in solution, K release rate constant and n is diffusional exponent.
Stability study
Stability study of the prepared FMM of Nimodipine was conducted for a period of 6 months as prescribed in ICH guidelines. The formulations were examined at the time points of 0, 1, 3 and 6 months for the different parameters [25].
In vivo pharmacokinetic study
Twelve number of male albino rabbits of body weight 1.5 kg were selected from the stock of animal house. They were divided into 2 groups having 6 animals in each group for test (minitablet administration) and standard (aqueous suspension of nimodipine). They were treated with hygienic food and fresh water twice daily. All animals were maintained for a wash-out period of 15 days prior to the study. The dose of nimodipine was calculated as 4.2 mg for each rabbit of weight approx. 1.5 kg.
The dose for rabbit was calculated as follows
$$ {\displaystyle \begin{array}{l} Total\ dose\ \left( in\ humans\right)\times 0.07\left( factor\ for\ each\ 1.5\ kg\ weight\ of\ rabbit\right)\\ {}=\left(60\times 0.07\times 1.5\right)/1.5=4.2\ mg\ for\ 1.5\ kg\ rabbit\end{array}} $$
The optimised formulation (minitablet) and aqueous dispersion of Nimodipine were administered in the form of dispersion to six male rabbits each in two groups using Ryle’s tube [26]. At different time points (0.5, 2, 4, 8, 12 and 24 h), blood sample of 0.5 mL was withdrawn from the marginal ear vein of the rabbit. Each blood samples was centrifuged for 10 min at 3000 rpm and the supernatant layer, i.e. serum was collected using micropipette. All the samples were analysed using reverse-phase ultrafast liquid chromatographic (UFLC) method. The various pharmacokinetic parameters like elimination rate constant (K), half-life (t1/2), peak plasma concentration (Cmax), time to attain the peak plasma concentration (Tmax), area under the curve (AUC) and area under the first moment curve (AUMC) were calculated. The animal experiments complied with the ARRIVE guidelines and carried out in accordance with the U.K. Animals (Scientific Procedures) Act, 1986 and associated guidelines, EU Directive 2010/63/EU for animal experiments. The pharmacokinetic study was approved by Institutional Animals Ethics committee [926/PO/Re/5/06/CPCSEA, Approval No.97]. Anaesthetic agents are not used since the procedures associated with routine blood withdrawal. Euthanasia method is not applicable for this study since the animals were rehabilitated after the study period as per the standard procedure. The animals were procured from Saha Enterprise, Kolkata, India (CPCSEA Regd no: 1828/PO/Bt/S/15/CPCSEA).
