Oral administration is one of the most simple, noninvasive and acceptable medication routes for most patients. However, the oral bioavailabilities of poorly soluble and/or poorly permeable drugs have been extremely low, which limits their clinical use by oral administration. For example, nimodipine is a dihydropyridine calcium channel blocker that is clinically used in preventing a major complication of subarachnoid hemorrhage. However, its absolute bioavailability after oral administration is as low as about 13%, thus resulting in an extraordinarily high required dose of about 360 mg per day [1
]. Therefore, development of new formulations for improving oral absorption of poorly soluble and/or poorly permeable drugs has been a sustained focus of pharmaceutics. Many lipid formulations, such as solid lipid nanoparticles (SLN) [2
], nanostructured lipid carrier (NLC) [3
], and nanoemulsions [4
] have been developed to improve the oral bioavailability of nimodipine. Among various strategies, the self-microemulsifying drug delivery system (SMEDDS) has attracted much attention. SMEDDS is an isotropic mixture with drugs dissolved or suspended in a mixture of oils, surfactants, and hydrophilic co-solvents, which can form spontaneously oil-in-water microemulsion in aqueous media under mild digestive motility of the gastrointestinal tract (GIT) [5
]. It has been widely proven that SMEDDS is one of the most effective approaches to improve drug solubility and dissolution, and oral absorption of poorly water-soluble drugs [6
]. However, there are also some shortcomings for SMEDDS, such as the risk of GIT irritation caused by a relatively high proportion of hydrophilic surfactants (20%–50%) and co-solvents (20%–50%) in SMEDDS, physical destabilization of the in situ formed microemulsions, drug crystallization and precipitation in vivo which becomes unavailable for absorption due to dispersion of gastric liquid and/or lipolysis digestion of small intestine lipase [9
]. In addition, just like SLN, NLC and nanoemulsions, SMEDDS also is a liquid form and is inconvenient for transportation and clinical applications.
Solid dosage form is preferable because of its good physicochemical stability, convenience of manufacturing, patient compliance and cost-performance. Therefore, transforming SMEDDS into a solid dosage form became a promising approach to overcome its fundamental drawbacks while retaining its pharmacokinetic benefits [11
]. Various solid self-microemulsifying drug delivery systems (S-SMEDDS) have been investigated by adding solid carriers to solidify SMEDDS [13
], such as silica-based water-insoluble adsorbents (e.g., porous silica), cellulose-based hydrophilic diluents (e.g., microcrystalline cellulose, hydroxypropyl methyl cellulose) and saccharide-based water-soluble diluents (e.g., maltodextrin, lactose) [16
Good S-SMEDDS must keep all the inherent merits of liquid SMEDDS. Appropriate solid carriers for S-SMEDDS could be selected by comparing the pharmacokinetic properties in vivo between S-SMEDDS and SMEDDS [19
]. In our previous study, the oral bioavailability in rabbits demonstrated that S-SMEDDS loading nimodipine (S-SMEDDS-Ni) with dextran as the solid carrier could preserve an improved bioavailability with releasing microemulsion droplets from the formulation in vivo [20
]. However, the mechanism of such a property of the solid carrier is not clear. Determining the influences of solid carrier on in vitro properties of S-SMEDDS, especially the structural transitions of reconstructed microemulsions after redispersed in water and drug loading, as well as precipitation and dissolution of S-SMEDDS in simulated gastric fluid, is essential for the reasonable choice of solid carriers. In addition, pharmacokinetic study in vivo was labor-intensive and expensive. Since characteristics in vitro of S-SMEEDS were closely related to their pharmacokinetic properties in vivo [21
], it was considered a reasonable, economical and convenient method to select proper solid carriers by studying influences of solid carriers on properties of S-SMEDDS with in vitro experiments.
In this study, we tried to compare the influences of different hydrophilic carriers on in vitro properties of S-SMEDDS-Ni and thereby, set up good in vitro methods to optimize a suitable carrier for SMEDDS. Influences of various hydrophilic carriers on in vitro characteristics of SMEEDS, including microstructural transitions, droplet size, drug loading, dispersion and precipitation of SMEDDS in simulated gastric fluid, were systematically studied. The effects of hydrophilic carriers on in vitro properties of S-SMEEDS, such as micromorphology, reconstruction of microemulsion, physical state of nimodipine and dissolution, were also assessed.
2. Materials and Methods
2.1. Chemicals and Reagents
Nimodipine (purity > 99.5%) and nimodipine tablet were purchased from Kaifeng Pharmaceutical (Group) Co., Ltd. (Kaifeng, China). Ethyl oleate was purchased from Shanghai Chemical Reagent Factory (Shanghai, China). Labrasol® and Cremophor® RH 40 were purchased from Gattefossé Corp., Lyon, France and BASF Corp., Lampertheim, Germany, respectively. Dextran 40 of pharmaceutical grade (weight-average molecular weight of 40,000) was purchased from Shanghai Huamao Pharmaceutical Co., Ltd. (Shanghai, China). Maltodextrin of medicinal grade was purchased from Shanghai Yun Hong Chemical Co., Ltd. (Shanghai, China). PVP K30 of pharmaceutical grade was purchased from Shanghai Pharmaceutical Excipient Factory (Shanghai, China). Acacia of analytical grade was purchased from Sigma Chemical Co. (St. Louis, MO, USA). Other solvents and chemicals were of analytical grade.
2.2. Preparation of SMEDDS and Droplet Size Determination
SMEDDS-Ni was prepared based on our pre-experiment and literature [23
]. Briefly, 280 mg of Cremophor®
RH, 7 mg of Labrasol®
and 5 mg of nimodipine were mixed at 37 °C until nimodipine was dissolved completely. Then 600 mg of ethyl oleate was added and shaken slowly at 37 °C to obtain a transparent and homogeneous liquid. Blank SMEDDS was prepared using the same procedure as SMEDDS-Ni without nimodipine being added.
The droplet size determination was carried out as follows. SMEDDS of 50 μL was added to pure water of 10 mL and vortex-mixed for 30 s. After standing for 30 min at 25 °C, the droplet size of resultant microemulsion was measured by photon correlation spectroscopy (PCS) at a wavelength of 635.0 nm, a scattering angle of 90° and a temperature of 25 °C with a Nano series ZS instrument (Zetasizer Nano-ZS, Malvern Instruments, Malvern, UK).
2.3. Effects of Carriers on Microstructure of SMEDDS
A series of microemulsions with water content varying from 0 to 95% were obtained by adding different amount of water into blank SMEDDS or SMEDDS-Ni. The conductivities of resultant microemulsions were measured with a DDS-2A conductivity meter (Shanghai Second Analytical Instrument Factory, Shanghai, China) and the conductivity-water content curves were drawn. The viscosities of resultant microemulsions near percolation thresholds were also measured by NDJ-8S digital viscometer (Shanghai Jingtian Electronic Instrument Co., Ltd., Shanghai, China). In the same way, different hydrophilic carrier solutions (5%, w/v) were added to SMEDDS-Ni, respectively, and the conductivity-water content curves were measured using the aforementioned method.
2.4. Effects of Carriers on Drug Loading of SMEDDS
Excessive nimodipine was added into a series of SMEDDS with water content of 0–90%. The mixture was vortex-mixed for 1 min and then shaken in the dark at 37 °C for 72 h. Finally, the mixture was centrifugated at 6000 rpm for 5 min. Nimodipine concentrations in supernatants were determined by high performance liquid chromatography (HPLC) and solubilities of nimodipine in SMEDDS with different water content were calculated. The solubilities of nimodipine in mixtures of SMEDDS and hydrophilic carriers were also measured in the same way.
HPLC analysis of nimodipine was conducted in an Agilent 1100 system with a Lichrospher C18 column (4.6 mm × 250 mm, 5 μm particle size). The mobile phase consisted of 0.05 mol·L−1
ammonium acetate and acetonitrile (35: 65, v
). The flow rate was set to 1.0 mL·min−1
and column temperature was set to 30 °C. The detection wavelength was 237 nm [20
]. The HPLC method was verified according to the Chinese Pharmacopoeia (2015 edition). The retention time of nimodipine was 7.6 min and excipients in formulations did not affect determination of nimodipine. The linear range was 3.00–300.00 μg·mL−1
= 0.9999). The intra-day and inter-day precision were 1.52% and 2.30%, respectively. The RSD of the repeatability test was 2.83% and the accuracy was 98.72%.
2.5. Effects of Carriers on Dispersion and Precipitation of SMEDDS in Simulated Gastric Fluid
SMEDDS-Ni of 2g was added into simulated gastric fluid (0.1 mol·L−1 HCl) of 200 g and stirred at 100 rpm, 25 °C in the dark. Samples were withdrawn at 0, 5,15 min, 24, 48, 72, 96, 120 and 144 h and centrifugated at 6000 rpm for 5 min. Nimodipine concentrations in supernatants were determined by HPLC mentioned above and amounts of dissolved nimodipine were calculated. In the same way, the amounts of nimodipine dissolved in simulated gastric fluid containing hydrophilic carriers (1%, w/v) were also measured.
2.6. Preparation of S-SMEDDS
S-SMEDDS was prepared based on preliminary experiments. Hydrophilic carrier of 10.0 g was dispersed in pure water of 100 mL and stirred until dissolved completely. Subsequently, SMEDDS-Ni of 10.0 g was added and stirred for 10 min. The resultant mixture was spray-dried using a B-191 Mini Spray-dryer (Büchi, Flawil, Switzerland), employing a flow rate of 5 mL·min−1, dry air flow rate of 500 NL·h−1, inlet temperature of 120 °C, which resulted in an outlet temperature of 70 °C.
2.7. Morphological Analysis of S-SMEDDS
The morphologies of S-SMEDDS were assessed by scanning electron microscopy (SEM). Samples were placed on a double-side electro-conductive adhesive tape which was fixed on an aluminum stub, and then sputter-coated with gold under argon atmosphere. SEM micrographs were taken using a FEI Sirion-200 SEM (Thermo Fisher Scientific Inc., Bleiswijk, The Netherlands).
2.8. Reconstitution Properties of S-SMEDDS
SMEDDS-Ni of 50 μL and S-SMEDDS-Ni of 100 mg prepared with different carriers were respectively diluted with 10 mL pure water and then were shaken vigorously for 30 s. After setting quietly for 30 min, droplet sizes of resultant microemulsions were measured.
2.9. Characterization of Inner Physical Structure of S-SMEDDS
Nimodipine raw material, S-SMEDDS-Ni and mixtures of nimodipine with different carriers were analyzed by differential scanning calorimetry (DSC) and powder X-ray diffraction (PXRD). Accurately weighted samples of 5 mg were placed in open aluminum pan. DSC was performed on a diamond differential scanning calorimeter (PerkinElmer, Waltham, MA, USA) at 5 °C·min−1 in the range of 10–150 °C under a nitrogen purge gas flow of 40 mL·min−1. PXRD was carried out with an X’Pert PRO diffractometer (PANalytical Inc., Almelo, Netherland). Cu Ka radiation at 40 mA and 40 kV with a step of 0.02° and a speed of 2° (2θ)·min−1 were used, covering a 2θ range of 10–40°.
2.10. In Vitro Dissolution Studies of S-SMEDDS
The dissolution of S-SMEDDS-Ni and nimodipine tablets were studied using Chinese Pharmacopoeia II apparatus with paddles [24
]. Acetate buffer of 900 mL with pH of 4.5 containing sodium lauryl sulfate (0.05%, w
) was used as the dissolution medium. Equivalent amounts of S-SMEDDS-Ni and nimodipine tablets (containing 10 mg of nimodipine) were put into the dissolution medium of 37 °C and stirred at 75 rpm, respectively. Samples of 2 mL were collected at designed intervals and equivalent fresh media were added. The collected samples were filtered through a millipore filter of 0.22 μm and drug concentrations were quantified by the HPLC method mentioned above.
2.11. Statistical Analyses
All data were expressed as mean ± standard deviation (S.D.). One-way ANOVA was used to test the differences between groups and P < 0.05 or P < 0.01 was considered to be a significant difference.