Biomimetic synthesis and evaluation of histidine-derivative templated chiral mesoporous silica for improved oral delivery of the poorly water-soluble drug, nimodipine

Accepted Manuscript Biomimetic synthesis and evaluation of histidine-derivative templated chiral mesoporous silica for improved oral delivery of the poorly water-soluble drug, nimodipine

Heran Li, Haiting Li, Chen Wei, Jia Ke, Jing Li, Lu Xu, Hongzhuo Liu, Yangyang, Sanming Li, Mingshi Yang PII: DOI: Reference:

S0928-0987(18)30128-3 doi:10.1016/j.ejps.2018.03.013 PHASCI 4442

To appear in:

European Journal of Pharmaceutical Sciences

Received date: Revised date: Accepted date:

29 December 2017 6 March 2018 8 March 2018

Please cite this article as: Heran Li, Haiting Li, Chen Wei, Jia Ke, Jing Li, Lu Xu, Hongzhuo Liu, Yangyang, Sanming Li, Mingshi Yang , Biomimetic synthesis and evaluation of histidine-derivative templated chiral mesoporous silica for improved oral delivery of the poorly water-soluble drug, nimodipine. The address for the corresponding author was captured as affiliation for all authors. Please check if appropriate. Phasci(2017), doi:10.1016/j.ejps.2018.03.013

This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

ACCEPTED MANUSCRIPT Biomimetic synthesis and evaluation of histidine-derivative templated chiral mesoporous silica for improved oral delivery of the poorly water-soluble drug, nimodipine Heran Li1, 2, Haiting Li2, 3, Chen Wei2, Jia Ke2, Jing Li1, Lu Xu2，Hongzhuo Liu2, Yangyang2, Sanming Li2, 3*, Mingshi Yang1, 4*

1

NU

SC

RI

PT

Wuya College of innovation, Shenyang Pharmaceutical University, Shenyang 110016, China 2 School of Pharmacy, Shenyang Pharmaceutical University, Shenyang 110016, China 3 School of Traditional Chinese Medicine, Shenyang Pharmaceutical University, Shenyang 110016, China 4 Faculty of Health and Medical Science, University of Copenhagen, Copenhagen, Denmark

MA

*Correspondence: Sanming Li, School of Pharmacy, Shenyang Pharmaceutical University, Wenhua RD 103, 110016, Shenyang, China; E-mail: [email protected] ; Tel: 8613840020260

PT E

D

Mingshi Yang, Wuya College of innovation, Shenyang Pharmaceutical University, Wenhua RD 103, 110016, Shenyang, China; E-mail: [email protected]; Tel: +4535336141; Fax: +4535336061;

AC

CE

Author: Heran Li, [email protected] Haiting Li, [email protected] Chen Wei, [email protected] Jia Ke, [email protected] Jing Li, [email protected]

Lu Xu，[email protected] Hongzhuo Liu, [email protected] Yangyang, [email protected]

ACCEPTED MANUSCRIPT Abstract

RI

PT

In this study, spherical shaped chiral mesoporous silica nanoparticles (CMS) was biomimetic synthesized using histidine derivatives (C16-L-histidine) as template via the sol–gel reaction and employed as poorly water-soluble drug nimodipine (NMP) carrier. Characteristics of CMS and its application as drug carrier were intensively investigated and compared with MCM41. Then NMP was respectively loaded into CMS and MCM41 with the drug: carrier weight ratio of 2:1. Structural features of NMP before and after drug loading were systemically characterized. The results demonstrated that hydrogen bonds were formed between NMP and carriers during the drug loading process. After drug loading, crystalline state of NMP effectively

MA

NU

SC

converted into modification L and amorphous state, and the first form turned out to be easily removed by washing. On the other hand, drug dissolution rate was significantly improved after drug loading, and the best result came from NMP-C3 sample. It was able to release 17.83% of drug within 60 min, which was 6.8-fold higher than the release amount of pure NMP. Undoubtedly, NMP-C3 presented the highest relative bioavailability (386.22%), and the best therapeutic effect. Meanwhile, CMS improved the brain distribution of NMP in vivo.

D

Keywords: biomimetic synthesis, chiral mesoporous silica, nimodipine, oral bioavailability, brain distribution

PT E

Introduction

AC

CE

Nimodipine (NMP) is a dihydropyridine calcium channel blocker primarily used in the prevention and treatment of cerebral vasospasm and resultant ischemia caused by subarachnoid hemorrhage (Sandow et al., 2016; Riekes et al., 2015; Fu et al., 2012). It was known as a kind of drug existed in two polymorphic forms which can coexist for a long time under ambient conditions: H-nimodipine (H-NMP) refers to a racemic compound, while L-nimodipine (L-NMP) crystallizes as a conglomerate (Riekes et al., 2012; Zu et al., 2014). Like many traditional drugs, NMP has some shortcomings reducing the drug therapeutic effect in clinically. Firstly, as a poorly water-soluble drug classified as BCS II, NMP exhibits low bioavailability after oral administration (less than 13%) which demonstrates poor and erratic absorption (Zhang et al., 2015; Yu et al., 2009). Besides, an extensive first pass metabolism of NMP is verified by researchers, which results in unsatisfactory pharmacokinetics and brings a lot of troubles for the clinical therapy and patient compliance (Riekes et al., 2015; Fu et al., 2012). In order to improve the bioavailability of poorly water-soluble drug, effective methods have been adopted such as co-solvents, micronization, salification and complexation (Li et al., 2012; Wang et al., 2015; Li et al 2017a). Meanwhile, a variety of drug delivery systems have been developed involving solid dispersions, liposomes,

ACCEPTED MANUSCRIPT

AC

CE

PT E

D

MA

NU

SC

RI

PT

vesicles and micelles (Zhao et al., 2017; Li et al., 2012; Li et al., 2017a). To sum up, increasing the solubility to improve the dissolution and controlling the drug release to maintain stable blood concentrations are two key strategies to improve the oral bioavailability of poorly water-soluble drugs. Compared with traditional materials, mesoporous silica materials with unique structural and textural properties have integrated the advantages of both nanomaterials and silica-containing fabrications, including: (1) large surface area and pore volume are favorable for drug adsorption and loading; (2) controllable mesoporous (2 to 50 nm) structure and tunable pore size provide better control of the drug size and drug loading and release; (3) non-toxic nature and excellent biocompatibility enable the application in biomedical field and minimize the toxic side effects; (4) easily-modified exterior and interior surface could improve the functionalities of controlled and targeted drug delivery systems; (5) functionalization with magnetic and/or luminescent compounds allows in vivo real-time monitoring and simultaneous drug delivery (Wang et al., 2015; Zhao et al., 2017; Song et al., 2017; Atluri et al., 2013). Due to the distinctive properties described above, mesoporous silica nanoparticles are considered as excellent candidates in the field of drug delivery (Paris et al., 2017; Popat et al., 2012). Since a MCM41 based drug release systems was first reported in 2001, biocompatible and non-invasive mesoporous silica materials as efficient drug delivery systems have attracted increasing attention (Mehmood et al., 2017; Perez et al., 2017). Great advances in structure control have been received for biomedical applications. For example, conventional mesoporous silica carriers, MCM41 and SBA-15 were utilized as immediate drug delivery systems by Jesus and co-workers (Jesus et al., 2016). The study demonstrated that pore size and structure were two important factors to command the drug-loading/releasing behavior of mesoporous silica. Maleki et al. exploited the potential of mesocellular siliceous foam (MSF) as atorvastatin carrier (Maleki et al., 2016). The result indicated that MSF released 70% of its cargo within 5 min. The fast drug release behavior of MSF could be attributed to the highly accessible pore network. Hu et al studied the feasibility of SBA-16 with a 3D cage-like cubic mesoporous structure as carvedilol carrier and compared that with MCM41 (Hu et al., 2012). The results demonstrated that SBA-16 exhibited a faster dissolution rate than MCM41, because the interconnected pore structure of SAB-16 reduced the diffusion resistance and facilitate the transport of drug. Li and coworkers reported the successful application of hollow mesoporous silica nanoparticles (HMSNs) with tunable shell thicknesses and hollow cores as drug carrier, and confirmed that the shell thickness and hollow size had significant impacts on drug-release profile (Li et al., 2017b). Recently, our group discovered that shell–core structure mesoporous silica with curved pores enhanced the dissolution rate and bioavailability of diazepam (DZP) after oral administration (Li et al., 2017a). In this case, due to geometric confinement of the twisty nanospace, DZP existed in amorphous form. Besides, brain distribution of DZP was improved after loading into this kind of mesoporous silica, which was 62.31-fold higher than that of pure DZP. Here, a new kind of chiral mesoporous silica nanoparticle (CMS) was synthesized

ACCEPTED MANUSCRIPT

PT

by biomimetic method and employed as NMP carrier. Characteristics of CMS and its application as nanocarrier were explored and compared with MCM41, which was a kind of mesoporous silica material that widely used in the field of drug delivery. NMP was respectively loaded into CMS and MCM41 using solvent dispersion method. XRD, DSC and FTIR studies were performed to give information on the crystalline conversion state of drug and the interactions between drug and carriers. Then the in vitro release behaviors and the in vivo pharmacokinetics of drug loaded silica carriers were investigated to evaluate the controlled release properties of CMS and MCM41. Finally, biodistribution and therapeutic effect studies were carried out to analyze the differences of their absorption and distribution.

RI

2. Materials and methods

SC

2.1 Materials

D

MA

NU

1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDCI), 1-hydroxybenzotriazole (HOBT), palmitic acid and 3-aminopropyltriethoxysilane (APTES) were purchased from Chengdu Xiya Chemical Technology Co. Ltd., (Chengdu, China). L-histidine methyl ester, tetraethyl orthosilicate (TEOS), 4-(dicyanomethylene)-2-methyl-6-(4-dimethylaminostyryl)-4H-pyran (DCM) and dimethylformamide (DMF) were obtained from Aladdin (Shanghai, China). MCM41 (average pore diameter, 3-4 nm; particle size, 200-300 nm) was obtained from Suzhou Jiedi Nano Technology Co., Ltd., (Suzhou, China). All other chemicals were

PT E

commercially available and used without any further purification. 2.2 Synthesis of C16-L-histidine

AC

CE

As described in Fig. 1A(a), C16-L-histidine methyl ester was synthesized by condensation of L-histidine methyl easer with palmitic acid through a one-pot synthetic procedure. Briefly, triethylamine and TMF were added to L-histidine methyl ester hydrochloride, and the achieved mixture was stirred for 20 min at room temperature. Then palmitic acid, EDCI and HOBT were introduced to the system. After stirring for 20 min, DCM was added dropwise, and the obtained mixture was transferred to an oil bath at 80℃. Progression of the reaction was monitored by thin layer chromatography (TLC). Afterwards, the obtained solution was filtered and successively washed with a saturated aqueous solution of NaHCO3, the organic solvent was removed under rotary evaporation, and the crude obtained was dried, crushed, weighted and recrystallized with n-hexane and ethyl alcohol (3:1, w/w). C16-L-histidine was synthesized by hydrolyzing C16-L-histidine methyl ester (Fig. 1A(b)). In detail, C16-L-histidine methyl ester was introduced to a mixture solution of NaOH (6.7 mol/L) and ethanol (3:7, w/w) with stirring on an oil bath at 50℃.

ACCEPTED MANUSCRIPT Afterwards, the system was moved to 0℃ water bath and stirred for another 6 h. The procedure was monitored by TLC. Then ethanol was removed using reduced pressure distillation. After adjusting pH to 2-3 by HCl (1 mol/L), the white precipitate obtained named C16-L-histidine was separated out, filtered, washed, dried and weighted. Figure 1

PT

2.3 Preparation of CMS Chiral mesoporous silica (CMS) was synthesized through sol-gel method. Briefly, a mixture of APTES and TEOS was dropwise added into a mixed solution of

SC

RI

C16-L-histidine, NaOH and deionized water under mild stirring at 25℃. The final gel molar composition was 1.1 C16-L-histidine: 7.0 TEOS: 1.0 APTES: 1.0 NaOH: 1111.1 H2O (pH 13). Stirring was stopped after 15 min, and then the system was remained statically for 24 h. Subsequently, the synthesized mixture was filtered, centrifuged,

NU

thoroughly washed, dried and finally calcined at 550℃ for 6 h.

PT E

2.5 NMP loading process

D

MA

2.4 Preparation of physical mixtures Nimodipine was selected and used as model drug. Physical mixtures of NMP with CMS and MCM41 were prepared through simple mixing of drug and carrier (2:1, w/w), and were denoted as NMP-C1 and NMP-M1, respectively.

NMP loaded to CMS was carried out by adding CMS to the methanol solution of NMP at the drug: carrier ratio of 2:1 (w/w). Then the mixture were stirred for 24 h

AC

CE

under ambient conditions, dried at 50℃ under vacuum, and labelled as NMP-C2. Afterwards, NMP-C2 was washed using ethanol to remove the drug adsorbed on the surface of CMS and dried under vacuum to get NMP-C3. Besides, drug loading sample composed of NMP and MCM41 was also prepared by solvent evaporation method at the drug: carried ratio of 2:1 (w/w) and the samples before and after washing were named as NMP-M2 and NMP-M3, respectively. Drug loading capacity (%) was ascertained by taking an accurately weighed quantity of both unwashed and washed products. Then the loaded NMP was extracted completely using ethanol under ultrasound. Finally the drug content was measured by ultraviolet (UV) spectroscopy (UV-1750, Shimadzu, Japan) at a wavelength of 238 nm. 2.6 Characterization The morphology and porous structure of MCM41 and the prepared CMS were

ACCEPTED MANUSCRIPT

MA

NU

SC

RI

PT

studied using a transmission electron microscope (TEM) instrument (Tecnai-G2-F30, FEI, Netherlands) and a scanning electronic microscope (SEM) instrument (JSM-6510A, JEOL, Japan). Small-angle X-ray diffractometry (SAXD) of silica samples was recorded on a XRD instrument (EMPYREAN, PANalytical, Netherlands) equipped with PIXcel-1D detector. The surface area and pore volume (Vt) of silica samples were analyzed by determining the nitrogen adsorption and desorption using a surface area and pore size analyzer (SA3100, Beckman Coulter, USA). Meanwhile, Brunauer–Emmett–Teller (BET) method and Barrett–Joyner–Halenda (BJH) method were used to calculate the specific surface area (SBET) and the pore size distribution (WBJH), respectively. Fourier transform infrared spectrums (FTIR, Spectrum 1000, Perkin Elmer, USA) of samples were collected over the spectral region 400–4000 cm-1 to study typical changes before and after drug loading. Besides, X-ray diffractometry (XRD) of samples were carried out at 30 mA and 30 kV with a Ni filtered CuKa line as the source of radiation using an X-ray diffractometer (X'pert PRO, PANalytical, Netherlands). Data was recorded from 5° to 45° (diffraction angle 2θ) at steps of 0.05°. Differential scanning calorimetry (DSC) tests were performed on a differential scanning calorimeter (Q1000, TA Instrument, USA) by heating the samples from 20 to 200℃ at a heating rate of 5℃/ min on an alumina holder under a nitrogen flow.

D

2.7 In vitro release behavior

AC

CE

PT E

The dissolution behaviors of samples were determined using a USP II paddle method with a stirrer rotation speed of 50 rpm at 37℃ in a dissolution apparatus (ZRD6-B, Huanghai Medicament Test Instrument, China) in the dark room. Then accurately weighed NMP, NMP-C3 and NMP-M3 samples containing 10 mg NMP were respectively exposed to 250 ml of deionized water. At predetermined time intervals, 5 ml of each sample was withdrawn and replaced by equal volume of fresh medium immediately. Then samples were filtered through a 0.22 μm membrane filter and measured using high performance liquid chromatography (HPLC) system equipped with a UV detector set at 240 nm. The column used was a Thermo ODS-2 Hypersil C18 column (250mm×4.6 mm, 5 mm, UK). The mobile phase was consisted of methanol, acetonitrile and water (3:4:3, v/v/v), and was pumped at a flow rate of 1 ml/min. The NMP content was calculated by a calibration curve. All the in vitro dissolution tests were performed in triplicate. 2.8 In vivo pharmacokinetics All the experiments were approved by the Animal Ethics Committee of Shenyang Pharmaceutical University. Animals were maintained in accordance with the

ACCEPTED MANUSCRIPT

PT

guidelines for the Care and Use of Laboratory Animals. Male Sprague-Dawley rats weighing 180-200 g were randomly divided into 3 groups (n=4): NMP group, NMP-C3 group and NMP-M3 group, and fasted overnight before the experiment with free access to water. The NMP, NMP-C3 and NMP-M3 samples suspended in physiological saline at a dose equivalent to 10 mg NMP were oral administrated to rats. Blood samples were collected via the ocular vein (about 0.6 ml each mouse) at 0.25 h, 0.5 h, 1 h, 2 h, 3 h, 4 h, 8 h, 12 h, 24 h after administration, centrifuged at 8000 rpm for 10 min, and then plasmas were stored at -20℃.

RI

2.9 Biodistribution

MA

NU

SC

To study the biodistribution of NMP after oral administration, 54 SPF-grade Kunming male mice weighing 18–22 g were randomly allocated into 3 groups: NMP group, NMP-C3 group and NMP-M3 group. Prior to oral administration, the animals were fasted overnight. Then mice in each group were treated with aqueous suspensions at a dose equivalent to 2 mg NMP. At 0.5 h, 1 h and 2 h after oral administration, the eyeballs of the mice were extirpated and the blood samples were collected from the fundus vessels of the eyes. After collection of blood, animals were sacrificed, tissue samples including the liver, kidney, spleen, lungs and brain were rapidly removed, weighed, homogenized, extracted with cold saline, centrifuged to

PT E

D

get supernatant, and stored at -20℃ for future analyses. The NMP concentration in blood and tissues were determined by a validated high performance liquid chromatography method (HPLC). 2.10 Sample preparation and analysis

AC

CE

200 μl plasma or tissue sample was mixed with 20 μl of internal standard nitrendipine solution (20 μg/ml) and vortexed for 1 min. Afterwards, 100 μl NaOH (0.01 mol/L) was added and vortexed for another 1 min. Then the mixture was extracted with 2.5 ml diethyl ether. After centrifugation at 6000 rpm for 12 min, the above organic layer (2 ml) was transferred to a new tube, evaporated to dryness at room temperature under a gentle stream of N2 and reconstituted with 100 μl mobile phase. After vortex and centrifugation, the sample (20 μl) was subjected to HPLC analysis. Nimodipine concentration in plasma and tissues were detected by HPLC. Analyses were performed by a Hitachi 10-AT pump, and a UV detector set at 240 nm. The separation was performed on a Thermo ODS-2 Hypersil C18 column (250mm×4.6 mm, 5 mm, UK) equipped with JanuSep C18 pre-column (Liaoning, China) with optimized mobile phase composed of 30% methanol, 40% acetonitrile and 30% water (v/v/v) at a flow rate of 1 ml/min. The oven temperature was kept at 25℃.

ACCEPTED MANUSCRIPT The NMP content was calculated by internal standard method. The main pharmacokinetic parameters were estimated by non-compartmental methods using the software of DAS2.0. 2.11 Pharmacodynamic evaluation

NU

SC

RI

PT

To further probe the function of NMP-C3 and NMP-M3, the mouse models of cerebral anoxia were established. 24 SPF-grade Kunming male mice weighing 18–22 g were randomly divided into 4 groups (n=6): saline group, NMP group, NMP-C3 group and NMP-M3 group. Before the experiment, animals were fasted overnight. The saline group was given saline only, and other groups were treated with a dose equivalent to 0.8 mg NMP by gastric gavage. 90 min after oral administration, cerebral anoxia of mice was induced by intraperitoneal injection of 0.4 ml 0.2% NaNO2 (w/v). The survival time of mice was recorded to detect the influence of dosage forms on protective effect of cerebral anoxia in mice. 2.12 Statistical analysis

PT E

3. Results and discussion

D

MA

Data was expressed as means with standard deviations (SD). Statistical analysis was analyzed using one-way analysis of variance. Statistical significance was set as p < 0.05.

CE

3.1 Morphology and structure of CMS and MCM41 Figure 2

AC

As illustrated in Fig. 1A(c), the anionic surfactant C16-L-histidine could self-assemble to form micelles in distilled water. By mixing, APTES and TEOS were polymerized to form silica framework. Then the negatively charged hydrophilic part of the C16-L-histidine interacted electrostatically with the positively charged ammonium ion of the APTES (Fig. 1A(d)). The synthesis of CMS was ascribed to biomimetic synthesis because amines catalyzed the condensation of silica precursors (Cusack et al., 2016; Fitzer et al., 2016). The supramolecular chirality was induced by the dynamic behavior of C16-L-histidine which grew in helical forms and acted as templates to direct local curved structure in the sol–gel reaction system. The size and detailed morphology of CMS and MCM41 were mainly analyzed by TEM and SEM. As can be seen in the SEM images (Fig. 2B and Fig. 2D), CMS and MCM41 were similar in a SEM recording scale, which were well-formed spherical nanoparticles with a diameter of 200-300 nm. However, significant difference can be

ACCEPTED MANUSCRIPT

AC

CE

PT E

D

MA

NU

SC

RI

PT

detected in TEM images. As can be seen in Fig. 2A, a large number of curved mesoscopic channels can be easily observed with wormlike arrangement. As shown in Fig. 2C, MCM41 (Fig. 2C) was nanospheres with distinguishable well-ordered lattice fringes. To sum up, CMS exhibited short and twisty disordered nanochannels, MCM41 possessed long and straight ordered channels. As can be seen Fig. 3A, in the SAXD pattern of CMS, the main peak was observed at 1.52° (2θ), and the second and third peaks were observed at 2.7° (2θ) and 3.1° (2θ), respectively, indicating the successful forming of mesostructure using C16-L-histidine as template. In the SAXD pattern of MCM41 (Fig. 3C), the three peaks appeared at 2.79° (2θ), 4.62° (2θ) and 5.26° (2θ), respectively. Judging from the SAXD patterns, the mesoporous silica samples all possessed 2D-hexagonal p6mm mesostructured (Goscianska et al., 2015). Meanwhile, the (100), (110) and (200) peaks in the diffractogram of CMS were located at smaller angles, which suggested larger lattice constant and d-spacing. It should be noticed that, based on the TEM result, although CMS possessed hexagonal porous structure, it should be identified as non-ordered mesoporous silica. The nitrogen adsorption/desorption isotherms and pore size distribution curves of CMS and MCM41 were displayed in Fig.3B and Fig.3D, respectively, and the calculated textural parameters of them were presented in Table 1. The nitrogen adsorption–desorption isotherms of the CMS (Fig. 3A) and MCM41 (Fig. 3B) after calcination exhibited type IV patterns with hysteresis loops, indicating the presence of uniform mesoporous. The BET surface area of CMS and MCM41 was found to be 732.16 m2/g and 997.54 m2/g, respectively, and the total pore volume of them was 0.94 cm3/g and 0.93 cm3/g, respectively. Besides, the BJH pore size of calcined CMS and MCM41 sample was calculated to be 3.98 nm and 3.36 nm, respectively. FT-IR analysis (Fig. 5) suggested that CMS was synthesized successfully consisting of bands at 3429 cm−1 (Si–OH antisymmetric stretching vibration) and 1091 cm−1 (Si– O–Si antisymmetric stretching vibration) (Li et al., 2014; Goscianska et al., 2015; Nanaki et al., 2017). It also indicated that MCM41 demonstrated a typical pattern for mesoporous silica materials, consisting of bands at 1081 cm-1 (antisymmetric stretching vibration）and 3431 cm-1（Si–OH antisymmetric stretching vibration). In addition, the XRD (Fig. 4A and Fig. 4B) and DSC results (Fig. 4C and Fig. 4D) revealed that CMS and MCM41 were amorphous material (Hu et al., 2015). The features mentioned above confirmed that CMS had great potential to be used as promising drug carriers. Figure 3 Table 1

ACCEPTED MANUSCRIPT 3.2 Drug loading capacity

RI

PT

Solvent deposition method was carried out to load NMP into mesoporous silica nanoparticles. The drug loading capacity (%) of CMS and MCM41 before and after washing was displayed in Table 2. For NMP loaded CMS, the drug loading content before and after washing was respectively measured to be 68.27% (NMP-C2) and 65.01% (NMP-C3), while for NMP loaded MCM41, that was respectively measured to be 66.80% (NMP-M2) and 59.29% (NMP-M3). The result demonstrated that CMS had the ability to load drug with high efficiency, and the drug loading capacity of CMS was higher than that of MCM41. It was worth noticing that only a small fraction (3.26%) of NMP was removed by washing in NMP loaded CMS, while 7.51% of

D

MA

NU

SC

NMP was removed in NMP loaded MCM41. The results demonstrated a stronger affinity between CMS and guest molecules. As displayed in Table 1, CMS and MCM41 had similar total pore volume, while the BET surface area of MCM41 was even higher than that of CMS. This difference in drug loading capacity between CMS and MCM41 was related to their differences in pore size and pore structure. As indicated by Perez et al., the mesopore size was one of the most important factors to influence the loading amount of drug molecules, because the pore diameter determined the size of the guest drugs that can be incorporated and the steric hindrance they would encounter during incorporation (Perez et al., 2017). In this case, the enlarged pore size of CMS provided smaller

AC

CE

PT E

steric hindrance for drug loading. On the other hand, assuming that NMP molecules (8.8 Å×7.6 Å×7.5 Å) were not sterically hindered in both mesoporous matrices, the fulfillment of CMS (mean pore size 3.98 nm) and MCM41 (mean pore size 3.36 nm) nanopores would imply formation of three layers and two layers of NMP molecules, respectively. In this respect, CMS had a higher storage capacity for NMP molecules. Meanwhile, as revealed by TEM images, the short twisty interconnected pore structure of CMS further reduced the diffusion hindrance and facilitated the mass transfer, while the long lattice fringes networks of MCM41 limited the accessibility of pores (Maleki et al., 2016; Gouze et al., 2014; Izquierdo-Barba. et al., 2005). Table 2 3.3 Drug crystalline state 3.3.1 XRD analysis XRD studies were performed to determine the crystalline state of samples before and after drug loading. The X-ray diffractograms of NMP, mesoporous silica carriers (MCM41 and CMS), physical mixtures (NMP-C1 and NMP-M1), unwashed drug

ACCEPTED MANUSCRIPT

AC

CE

PT E

D

MA

NU

SC

RI

PT

loaded samples (NMP-C2 and NMP-M2) and washed drug loaded samples (NMP-C3 and NMP-M3) were shown in Fig. 4A and Fig. 4B, respectively. It was well known that, nimodipine can exist in two polymorphic forms in nature at room temperature, which were H-NMP and L-NMP (Zu et al., 2014; Riekes et al., 2012; Babu et al., 2012). In both polymorphic forms, the unit-cell was built by four nimodipine molecules, and weak hydrogen-bondings were formed between the alkoxy groups and the dihydropyridine moieties in nimodipine units (Kiwilsza et al., 2015; Liu et al., 2016). The differences were mainly caused by the intermolecular van der Walls forces. Between them, H-NMP referred to a racemic mixture (Fig. 1B(b)), while the thermodynamically stable polymorphic form L-NMP crystallized as a conglomerate (Fig. 1B(c)). As shown in Fig. 4, CMS and MCM41 all showed a broad band between 5° and 45° (2θ), indicated the amorphous features of silica matrices. Pure NMP was highly crystallized and showed typical peaks of H-NMP including characteristic peaks at 20.7° (2θ), 18.75° (2θ) and 6.75° (2θ). Meanwhile, the physical mixtures (NMP-C1 and NMP-M1) showed the diffraction peaks of H-NMP with reduction in the intensity. However, the XRD patterns of NMP-C2 and NMP-M2 both showed different reflections from pure NMP sample with reduction in peak intensity. Peaks were observed at 15.3° (2θ), 20.5° (2θ) and 10.8° (2θ), respectively, which were found to be the characteristic partial diffraction peaks of L-NMP, indicating a phase transition during drug loading process (Zu et al., 2014; Urbanetz et al., 2005; Docoslis et al., 2007). Besides, NMP-C3 and NMP-M3 samples showed the amorphous broad peaks with very weak diffraction peaks belonging to a few amount of L-NMP. The results proved that, in the unwashed samples (NMP-C2 and NMP-M2), part of NMP molecules were located on the external surface of carriers, and the others were entered into the inside nanopores of mesoporous carriers. For washed samples (NMP-C3 and NMP-M3), most of the NMP molecules adsorbed on the surface of carriers can be easily removed by washing, and NMP left inside the pores did not have a crystalline form. Figure 4

3.3.2 DSC analysis DSC analysis was also carried out to check the physical state of samples before and after drug loading. Fig. 4C and Fig. 4D presented the DSC thermograms of samples. According to literatures, H-NMP possessed a melting peak at higher temperature (124.0℃) , while L-NMP showed a melting peak at lower temperature (114.4℃) (Zu et al., 2014; Barmpalexis et al., 2011). As can be seen in the DSC thermograms, the endothermic events of CMS and MCM41 samples were comprised of amorphous

ACCEPTED MANUSCRIPT broad peaks. Besides, the melting peaks of H-NMP can be easily detected in the DSC thermograms of pure NMP and the physical mixtures (NMP-C1 and NMP-M1)

MA

NU

SC

RI

PT

around 124.1 ℃ . Meanwhile the NMP-C2 and NMP-M2 samples respectively exhibited endothermic peaks at 114.4℃ and 114.7℃, suggesting a phase transition (Barmpalexis et al., 2011; Urbanetz et al., 2005). The DSC traces conducted on NMP-C3 and NMP-M3 were devoid of NMP peaks, suggesting that NMP existed in amorphous state. The amorphization of drug can be explained by the finite-size effect of mesopores, which prevented the drug molecules from rearranging themselves in a crystalline form. It’s worth mentioning that, the DSC findings were in good agreement with the XRD results. These results proved that in the washed samples (NMP-C3 and NMP-M3) drug was only located inside the mesopores in amorphous state. On the contrary, the unwashed samples (NMP-C2 and NMP-M2) should be interpreted as kinds of dual systems. NMP was partly left on the external surface of silica carriers in modification L, and partly entered inside the pores in amorphous state during the drug loading process. The fraction adsorbed on the outside turned out to be easily removed by washing. 3.4 Interactions between drug and carrier

AC

CE

PT E

D

FTIR analysis were carried out to illustrate the possible interactions between the NMP and silica carriers, and the FTIR spectra of NMP loaded CMS and MCM41 samples were presented in Fig. 5A and Fig. 5B, respectively. As shown in Fig. 5, NMP showed its characteristic peaks, including the peak at 3300 cm−1 belonging to the N-H stretching vibration, the peak at 1309 cm−1 attributed to the C-N stretching vibration, and the peak at 1693 cm−1 relative to the C=O group (Papageorgiou et al., 2009; Barmpalexis et al., 2011). The FTIR spectra of CMS and MCM41 were similar because of the similar composition of the two silica materials. Besides, the FTIR spectra of the physical mixtures (NMP-C1 and NMP-M1) were the overlap of the spectra of NMP and silica carriers, indicating the absence of polymorphic transitions or chemical interactions. In the spectra of the unwashed NMP loaded CMS and MCM41 samples (NMP-C2 and NMP-M2), the peaks corresponding to the –NH– groups were shifted from 3300 cm−1 to 3270 cm−1 and 3273 cm−1, respectively. Meanwhile, the peaks relative to Si-OH groups were also shifted to lower wavenumbers for both NMP-C2 (from 3429 cm-1 to 3424 cm-1) and NMP-M2 (from 3431 cm-1 to 3428 cm-1). The red shifts of–NH– group and Si-OH group respective bands indicated that hydrogen bonds were formed between silanol groups on the silica surface and the amine groups of NMP (Barmpalexis et al., 2011; Ewing et al., 2015; Gorajana et al., 2011). Besides, in the region of 1600-1700 cm-1, the sharpness of peak at 1693 cm−1 was decreased, and the relative intensity between peaks at 1648 cm−1 and 1621 cm−1 was pronounced changed, which strongly confirmed the existence of amorphous nimodipine (Barmpalexis et al., 2011; Kiwilsza et al., 2015; Riekes et al.,

ACCEPTED MANUSCRIPT

PT

2015). In the FTIR spectra of the washed samples (NMP-C3 and NMP-M3), most peaks of NMP can be hidden after loaded into carriers, demonstrating that NMP was loaded into the internal space of CMS and MCM41. In other words, NMP accumulated on the outside surface of silica nanoparticles was effectively removed by washing. The FTIR study illustrated that NMP can be successfully loaded into the mesopores of silica carriers. During encapsulation, hydrogen bonds were formed between silica materials and the NMP molecule.

RI

Figure 5 3.5 In vitro drug release study

D

MA

NU

SC

According to literatures, the aqueous equilibrium solubility of nimodipine was found to be 3.86 μg/ml (Kiwilsza et al., 2015; Zhang et al., 2015). So that, the drug release study was performed under supersaturating conditions (the mass concentration of NMP was about 10 times higher than the equilibrium solubility). The release behaviors of the washed drug loading samples (NMP-C3 and NMP-M3) were investigated and the results were presented in Fig. 6. For pure NMP, very poor and incomplete dissolution was observed, in which only 9.06% of NMP was released within 48 h. However, after loading into mesoporous silica, the dissolution rate of NMP was significantly improved. The cumulative drug release amount of NMP-C3

AC

CE

PT E

and NMP-M3 reached 25.76% (10.31 μg/ml) and 11.54% (4.62 μg/ml) within 48 h, which was 2.84-fold higher and 1.27-fold higher than NMP (3.62 μg/ml), respectively. Particularly, NMP-C3 and NMP-M3 were able to release 17.83% and 7.44% of NMP within 60 min, and the results were 6.8-fold higher and 2.8-fold higher than that of NMP (released 3.06% within 48 h), respectively. The dissolution-promoting effect of CMS and MCM41 for NMP was obvious. The supersaturated concentrations were maintained for at least 47.5 h and 40 h for CMS and MCM41, respectively. As one of the key strategies for dissolution improvement, the functionality of mesoporous silica materials can be explained by both the amorphization of the drug inside the nanopores and the increase of specific surface area after drug loading (Li et al., 2017b; Li et al., 2015). Comparing the dissolution profiles of NMP-C3 and NMP-M3, it was obvious that the dissolution of NMP from CMS was much faster than that from MCM41. Many previous studies had confirmed that, drug release from mesoporous materials was a mainly diffusion-controlled process controlled by the structure of mesoporous (Zhang et al., 2015; Fu et al., 2016; Li et al., 2016). TEM images revealed that, CMS possessed short and twisty disordered nanochannels with wormlike arrangement, while MCM41 exhibited long and straight ordered well-ordered lattice fringes. It had been verified

ACCEPTED MANUSCRIPT

NU

Figure 6

SC

RI

PT

that the pore diameter, combined with the pore geometry and connectivity, had a great impact on the drug release behavior of mesoporous silica (Wang et al., 2015; Paris et al., 2017). Both large size and high accessibility of pores can improve the drug release characteristic of mesoporous silica (Horcajada et al., 2004; Izquierdo-Barba. et al., 2005; Izquierdo-Barba. et al., 2009). In this case, the larger pore diameter of CMS (3.98 nm) provided smaller steric hindrance for drug release compared with MCM41 (3.36 nm). Besides, the outstanding dissolution performance of NMP-C3 can also be related to the pore morphology of CMS, the short and interconnected nanopores of CMS further reduced the diffusion hindrance and facilitated mass transport into the bulk solution (Izquierdo-Barba et al., 2009). On the contrary, MCM41 possessed long lattice fringes, resulting in longer diffusion distance and higher diffusion resistance for drug molecules to release in the dissolution medium. As a result of beneficial pore architectures, CMS could provide more favorable dissolution-promoting functionality.

3.6 Pharmacokinetic study

AC

CE

PT E

D

MA

Pharmacokinetic studies of NMP-C3 and NMP-M3 were carried out with pure NMP as a reference. The plasma drug concentration time profiles of NMP formulations (pure NMP, NMP-C3 and NMP-M3) after oral administration were presented in Fig. 7, and the pharmacokinetic parameters were summarized in Table 3. As shown in Fig. 7, triple peaks were observed in the profile of pure NMP at 0.5 h, 3 h and 8 h after oral administration, respectively, which strongly confirmed the existence of enterohepatic circulation. In the concentration versus time curves of NMP-M3, a small peak was observed at 0.5 h, then the plasma level of NMP rose quickly and the maximum concentration (Cmax) was reached at 3 h post-administration. Compared to pure NMP, NMP-M3 showed no significant difference in Tmax (3 h) and 2.68-fold higher in Cmax. In the case of NMP-C3, clear differences in the shape of the drug concentration time curves can be observed, in which only one peak was found. After loading into CMS, Tmax was reached later (8 h post-administration), and Cmax was improved. Interestingly, although the Cmax of NMP-C3 was lower than NMP-M3, the relative bioavailability of NMP-C3 was 2.74-fold higher than that of NMP-M3. As be seen in Fig .7, the plasma drug concentration of NMP-C3 was higher than NMP-M3 from 3 to 24 h, indicating that, NMP-C3 provided longer sustained plasma drug concentration and a sufficient drug exposure in vivo. As displayed in Table 3, the relative bioavailability of pure NMP, NMP-C3 and NMP-M3 were 100%, 386.22% and 140.82%, respectively. Compared to pure NMP, NMP-C3 and NMP-M3 showed 3.86-fold and 1.41-fold higher in AUC, respectively. The result revealed that the bioavailability of NMP was significantly improved after

ACCEPTED MANUSCRIPT

PT

loading into CMS and MCM41, which was in accord with the in vitro release behaviors. The improvement in oral absorption was mainly owing to the increased drug solubility caused by the formation of non-crystalline state and the controlled release effect of mesoporous silica carriers in the gastrointestinal tract. Besides, the high surface area of mesoporous materials made the nanoparticles more easily to adhere on the small intestine wall, which improved the opportunity of absorption (Wacher et al., 2001; Zhang et al., 2015). In addition, the internal space of the mesoporous materials enhanced the drug stability in vivo (Markel et al., 2011; Zhang et al., 2015).

RI

Figure 7

SC

Table 3 3.7 Biodistribution

AC

CE

PT E

D

MA

NU

To investigate the difference in drug distribution behavior between pure NMP, NMP-C3 and NMP-M3, the NMP contents in blood and organs were detected by HPLC at 30 min, 60 min and 120 min after oral administration, and the results were displayed in Fig. 8 A-C, respectively. Generally speaking, three NMP formulations showed different trends of distribution in vivo. NMP-C3 had a higher NMP content in plasma concentration and brain, and lower content in kidney at all time points. NMP-M3 had a higher NMP content in heart at all time points. At 30 min post-administration (Fig. 8A), NMP-C3 had a significantly improved drug distribution in brain (which was 1.81-fold higher and 1.70-fold higher than pure NMP and NMP-M3, respectively). It should be noticed that the fast distribution in brain might contribute to the clinical curative effect of NMP in the treatment of cerebral hemorrhage. At 120 min post-administration (Fig. 8C), observable amount of NMP-C3 had been trapped by spleen, which was 1.76-fold higher and 2.39-fold higher than pure NMP and NMP-M3 sample, respectively. In a word, CMS and MCM41 could alter the biodistribution of NMP. By loading into CMS, the brain distribution of NMP had been improved, and the mechanism need to be further studied. Figure 8 3.8 Therapeutic effect To evaluate the therapeutic effects of NMP-C3 and NMP-M3 on cerebral anoxia, the survival time of mice on cerebral anoxia induced by NaNO2 were investigated and compared with that of pure NMP (positive control). Meanwhile, normal saline was

ACCEPTED MANUSCRIPT

PT

given to mice and regarded as negative control. It can be seen from Fig. 8D, in saline group, the survival time of saline treated group was only 9.27 min. Meanwhile, the survival time of the NMP group underwent a prolongation to 10.43 min because NMP improved the oxygen supply in blood and promoted the oxygen utilization coefficient of brain cells. Meanwhile, NMP-M3 prolonged the survival time to 11.37 min. Noteworthy, after loading into CMS, the survival time of mice was significantly enhanced to 13.61 min as a result of the rapid brain distribution. The results proved that NMP-C3 exhibited high therapy efficacy, which provided convincing evidence for the improvement of oral absorption and brain distribution.

RI

4. Conclusion

PT E

D

MA

NU

SC

In summary, a novel drug release system established by CMS was successfully developed to provide effective strategy in improving the bioavailability of poorly water-soluble drug nimodipine. The results showed that CMS was spherical nanoparticles with curved disordered nanospace, while MCM41 was nanospheres with ordered lattice fringes. Benefiting from the pore architectures, CMS was able to load NMP with high capacity. During drug loading process, hydrogen bonds were formed and NMP effectively converted into its modification L and amorphous state, and the first form turned out to be easily removed by washing. As a result, dissolution, bioavailability and therapeutic effect of NMP were dramatically improved. Meanwhile, CMS improved the brain distribution of NMP in vivo. Undoubtedly, the novel CMS-based drug delivery system was effective in controlling the drug release rate and improving the oral bioavailability of poorly water-soluble drugs.

CE

Disclosure

The authors report no conflicts of interest in this work.

AC

Acknowledgements

This work was supported by the National Natural Science Foundation of China (No. 81773672, No. 81473161) and the China Postdoctoral Science Foundation (No. 2016M590235). References  Atluri, R., Iqbal, M.N., Bacsik, Z., Hedin, N., Villaescusa, L.A., Garcia-Bennett, A.E. 2013. Self-assembly mechanism of folate-templated mesoporous silica. Langmuir. 29 (38), 12003-12.  Babu, G.V.M.M., Kumar, N.R., Sankar, K.H., Ram, B.J., Kumar, N.K., Murthy,

ACCEPTED MANUSCRIPT













NU

MA

D

PT E



CE



AC



SC

RI



PT



K.V.R., 2002. In vivo evaluation of modified gum karaya as a carrier for improving the oral bioavailability of a poorly water-soluble drug, nimodipine. Aaps Pharmscitech. 3 (2), 55-63. Barmpalexis, P., Kachrimanis, K., Georgarakis, E., 2011. Solid dispersions in the development of a nimodipine floating tablet formulation and optimization by artificial neural networks and genetic programming. Eur. J. Pharm. Biopharma. 77, 122-131. Cusack, M., 2016. Biomineral electron backscatter diffraction for palaeontology. Palaeontology. 59, 171-179. Docoslis, A., Huszarik, K.L., Papageorgiou, G.Z., Bikiaris, D., Stergiou, A., Georgarakis, E., 2007. Characterization of the distribution, polymorphism, and stability of nimodipine in its solid dispersions in polyethylene glycol by micro-Raman spectroscopy and powder x-ray diffraction. Aaps J. 9 (3), E361-E370. Ewing, A.V., Wray, P.S., Clarke, G.S., Kazarian, S.G., 2015. Evaluating drug delivery with salt formation: drug disproportionation studied in situ by ATR-FTIR imaging and raman mapping. J. Pharmaceut. Biomed. 111, 248-256.. Fitzer, S.C., Chung, P., Maccherozzi, F., Dhesi, S.S., Kamenos, N.A., Phoenix, V.R., Cusack, M., 2016. Biomineral shell formation under ocean acidification: a shift from order to chaos. Sci. Rep. 6, 21076. Fu, Q., Kou, L.F., Gong, C., Li, M., Sun, J., Zhang, D., Liu, M.N., Sui, X.F., Liu, K., Wang, S.L., He, Z.G., 2012. Relationship between dissolution and bioavailability for nimodipine colloidal dispersions: the critical size in improving bioavailability. Int. J. Pharm. 427, 358-364. Fu, W.H., Guan, Y., Wang, Y.M., He, M.Y., 2016. A facile synthesis of monodispersed mesoporous silica nanospheres with Pm3n structure. Micropor. Mesopor. Mat. 220, 168-174. Gorajana, A., Rao, N.K., Nee, W.Y., 2011. Physicochemical characterization and in vitro evaluation of solid dispersions of nimodipine with PEG 8000. Asian J. Chem. 23, 2857-2859. Goscianska, J., Olejnik, A., Nowak, I., Marciniak, M., Pietrzak, R., 2015. Ordered mesoporous silica modified with lanthanum for ibuprofen loading and release behaviour. Eur. J. Pharm. Biopharm. 94, 550-558. Gouze, B., Cambedouzou, J., Parrès-Maynadié, S., Rébiscoul, D. 2014. How hexagonal mesoporous silica evolves in water on short and long term: role of pore size and silica wall porosity. Micropor. Mesopor. Mat. 183(1), 168-176. Horcajada, P., Rámila, A., Pérez-Pariente, J., Vallet-Regi. M. 2004. Influence of pore size of mcm-41 matrices on drug delivery rate. Micropor. Mesopor. Mat. 68(1), 105-109.

ACCEPTED MANUSCRIPT

AC

CE

PT E

D

MA

NU

SC

RI

PT

 Hu, L., Sun, H.R., Zhao, Q.F., Han, N., Bai, L., Wang, Y., Jiang T.Y., Wang, S.L., 2015. Multilayer encapsulated mesoporous silica nanospheres as an oral sustained drug delivery system for the poorly water-soluble drug felodipine. Mat. Sci. Eng. C. 47, 313-324.  Hu, Y., Zhi, Z., Zhao, Q., Wu, C., Zhao, P., Jiang, H., et al. 2012. 3d cubic mesoporous silica microsphere as a carrier for poorly soluble drug carvedilol. Micropor. Mesopor. Mat. 147(1), 94-101.  Izquierdo-Barba, I., África Martinez, Doadrio, A.L., Pérez-Pariente, J., Vallet-Regí, M. 2005. Release evaluation of drugs from ordered three-dimensional silica structures. Eur. J. Pharm. Sci. 26(5), 365-373.  Izquierdo-Barba, I., Sousa, E., Doadrio, J. C., Doadrio, A.L., Pariente, J. P., & Martínez, A., et al. 2009. Influence of mesoporous structure type on the controlled delivery of drugs: release of ibuprofen from mcm-48, sba-15 and functionalized sba-15. J. Sol-Gel Sci. Techn. 50(3), 421-429.  Jesus, R.A., Rabelo, A.S., Figueiredo, R.T., Silva, L. C.C.D., Codentino, I.C., Fantini, M.C.A. 2016. Synthesis and application of the mcm-41 and sba-15 as matrices for invitro efavirenz release study. J. Drug Deliv. Sci. Tech. 31, 153-159.  Kiwilsza, A., Milanowski, B., Drużbicki, K., Coy, L.E., Grzeszkowiak, M., Jarek, M. 2015. Mesoporous drug carrier systems for enhanced delivery rate of poorly water-soluble drug: nimodipine. J. Porous Mat. 22(3), 817-829.  Li, H.R., Wang, J.X., Cong, J.L., Wei, C., Li, J., Liu, H., Li, S.M., Yang, M.S., 2017a. Biomimetic synthesis of proline-derivative templated mesoporous silica for increasing the brain distribution of diazepam and improving the pharmacodynamics of nimesulide. Drug Deliv. 24 (1), 1086-1098.  Li, J., Xu, L., Liu, H.Z., Wang, Y., Wang, Q.F., Chen, H.T., Pan, W.S., Li, S.M., 2014. Biomimetic synthesized nanoporous silica@poly(ethyleneimine)s xerogel as drug carrier: characteristics and controlled release effect. Int. J. Pharm. 467, 9-18.  Li, J., Xu, L., Yang, B.X., Bao, Z.H., Pan, W.S., Li, S.M., 2015. Biomimetic synthesized chiral mesoporous silica: structures and controlled release functions as drug carrier. Mat. Sci. Eng. C. 55, 367-372.  Li, J., Xu, L., Zheng, N., Wang, H.Y., Lu. F.Z., Li, S.M., 2016. Biomimetic synthesized bimodal nanoporous silica: bimodal mesostructure formation and application for ibuprofen delivery. Mat. Sci. Eng. C. 58, 1105.  Li, Y., Li, N., Pan, W., Yu, Z., Yang, L., Tang, B. 2017b. Hollow mesoporous silica nanoparticles with tunable structures for controlled drug delivery. Acs. Appl. Mater. Inter. 9(3).  Li, Z., Barnes, J. C., Bosoy, A., Stoddart, J. F., Zink, J. I. 2012. mesoporous silica nanoparticles in biomedical applications. Chem. Soc. Rev. 43(27), 2590-605.

ACCEPTED MANUSCRIPT

RI

PT

 Liu, W.L., Wang, X.N., Chen, R.L., Zhang, K.X., Li, Y., Li, Y., Si, D.Y., Gong, J.B., Yin, D.S., Wang, Y.L., Wei, Z.P., Yang, M.S., 2016. Effect of age on the pharmacokinetics of polymorphic nimodipine in rats after oral administration. Acta Pharm. Sin. B. 6, 468-474.  Maleki, A., Hamidi, M. 2016. Dissolution enhancement of a model poorly water-soluble drug, atorvastatin, with ordered mesoporous silica: comparison of msf with sba-15 as drug carriers. Expert Opin. Drug Deliv. 13(2), 171.  Markel, A.L., Achkasov, A.F., Alekhina, T.A., Prokudina, O.I., Ryazanova, M.A., Ukolova, T.N., Efimov, V.M., Boldyreva, E.V., Boldyrev, V.V., 2011. Effects of the alpha- and gamma-polymorphs of glycine on the behavior of catalepsy prone rats. Pharmacol. Biochem. 98, 234-240.

AC

CE

PT E

D

MA

NU

SC

 Mehmood, A.，Ghafar, H.，Yaqoob, S.，Gohar, U.F. Ahmad, B. 2017. Mesoporous Silica Nanoparticles: A Review. J. Develop. Drugs. 6,2  Nanaki, S., Siafaka, P.I., Zaxariadou, D., Nerantzaki, M., Giliopoulos, D.J., Triantafyllidis, K.S., 2017. PlGA/SBA-15 mesoporous silica composite microparticles loaded with paclitaxel for local chemotherapy. Eur. J. Pharm. Sci. 99, 32-44.  Papageorgiou, G.Z., Docoslis, A., Georgarakis, M., Bikiaris, D., 2009. The effect of physical state on the drug dissolution rate. J. Therm. Anal. Calorim. 95, 903-915.  Paris, J.L., Colilla, M., Izquierdo-Barba, I., Manzano, M., Vallet-Regí, M. 2017. Tuning mesoporous silica dissolution in physiological environments: a review. J. Mat. Sci. 52(15), 8761-8771.  Perez, R.A., Singh, R.K., Kim, T. H., Kim, H.W. 2017. Silica-based multifunctional nanodelivery systems toward regenerative medicine. Mater. Horiz.  Popat, A., Liu, J., Hu, Q., Kennedy, M., Peters, B., Lu, G.Q. 2012. Adsorption and release of biocides with mesoporous silica nanoparticles. Nanoscale. 4(3), 970.  Riekes, M.K., Caon, T., Jr, J.D.S., Sordi, R., Kuminek, G., Bernardi, L.S., Rambo, C.R., de Campos, C.E.M., Fernandes, D., 2015. Enhanced hypotensive effect of nimodipine solid dispersions produced by supercritical CO2, drying. Powder Technol. 278, 204-210.  Riekes, M.K., Pereira, R.N., Rauber, G.S., Cuffini, S.L., de Campos, C.E., Silva, M.A., Stulzer, H.K., 2012. Polymorphism in nimodipine raw materials: development and validation of a quantitative method through differential scanning calorimetry. J. Pharmaceut. Biomed. 70, 188-193.  Sandow, N., Diesing, D., Sarrafzadeh, A., Vajkoczy, P., Wolf, S. 2016. Nimodipine dose reductions in the treatment of patients with aneurysmal

ACCEPTED MANUSCRIPT





PT

RI

SC

NU



MA



D



PT E



CE



AC



subarachnoid hemorrhage. Neurocrit. Care. 25, 29-39. Song, Y., Li, Y., Xu, Q., Liu, Z. 2017. Mesoporous silica nanoparticles for stimuli-responsive controlled drug delivery: advances, challenges, and outlook. Int. J. Nanomed. 12, 87-110. Urbanetz, N.A., Lippold, B.C., 2005. Solid dispersions of nimodipine and polyethylene glycol 2000: dissolution properties and physico-chemical characterisation. Eur. J. Pharm. Biopharm. 59, 107-118. Wacher, V.J., Salphati, L., Benet, L.Z., 2001. Active secretion and enterocytic drug metabolism barriers to drug absorption. Adv. Drug Deliv. Rev. 46, 89-102. Wang, Y., Han, N., Zhao, Q., Bai, L., Li, J., Jiang, T., 2015. Redox-responsive mesoporous silica as carriers for controlled drug delivery: a comparative study based on silica and peg gatekeepers. Eur. J. Pharm. ScI. 72, 12-20. Yu, H., Zhai, Q.Z., 2009. Mesoporous SBA-15 molecular sieve as a carrier for controlled release of nimodipine. Micropor. Mesopor. Mat. 123, 298-305. Zhao, Q., Lin, Y., Han, N., Li, X., Geng, H., Wang, X., et al. 2017. Mesoporous carbon nanomaterials in drug delivery and biomedical application. Drug Deliv. 24, 94. Zhang, Y., Zhao, Q., Zhu, W., Zhang, L., Han, J., Lin, Q., et al. 2015. Synthesis and evaluation of mesoporous carbon/lipid bilayer nanocomposites for improved oral delivery of the poorly water-soluble drug, nimodipine. Pharm. Res. 32(7), 2372. Zu, Y.G., Li, N., Zhao, X.H., Li, Y., Ge, Y.L., Wang, W. G., Wang, K. L., Liu, Y., 2014. In vitro dissolution enhancement of micronized l-nimodipine by antisolvent re-crystallization from its crystal form H. Int. J. Pharm. 464, 1-9.

ACCEPTED MANUSCRIPT Figure legend

PT

Fig. 1 A, Synthetic routes of C16-L-histidine and CMS. a, dehydration–condensation process; b, ester hydrolysis reaction; c, interaction between functional group of APTES and C16-L-histidine; d, cooperative self-assembly of silica source, CSDA and template; e, CMS obtained after calcination; B, a, 3D structure of nimodipine (with mean size of its molecule); b, unit cell of the H-nimodipine crystal; c, unit cell of the L-nimodipine crystal.

RI

Fig. 2 A, TEM images of CMS; B, SEM image of CMS; C, TEM images of MCM41; D, SEM image of MCM41.

NU

SC

Fig. 3 A, SAXD pattern of CMS; B, Nitrogen adsorption/desorption isotherm and pore size distribution curve of CMS; C, SAXD pattern of MCM41; D, Nitrogen adsorption/desorption isotherm and pore size distribution curve of MCM41.

D

MA

Fig. 4 A, XRD patterns of NMP, CMS, NMP-C1, NMP-C2 and NMP-C3; B, XRD patterns of NMP, MCM41, NMP-M1, NMP-M2 and NMP-M3; C, DSC thermograms of NMP, CMS, NMP-C1, NMP-C2 and NMP-C3; D, DSC thermograms of NMP, MCM41, NMP-M1, NMP-M2 and NMP-M3.

PT E

Fig. 5 A, FTIR spectra of NMP, NMP-C1, NMP-C2, NMP-C3 and CMS; B, FTIR spectra of NMP, NMP-M1, NMP-M2, NMP-M3 and MCM41.

CE

Fig. 6 In vitro release profiles of NMP, NMP-C3 and NMP-M3.

AC

Fig. 7 Plasma concentration–time profiles of NMP, NMP-C3 and NMP-M3. Fig. 8 A, NMP biodistribution at 30 min after oral administration; B, NMP biodistribution at 60 min after oral administration; C, NMP biodistribution at 90 min after oral administration; D, The survival time of mice on cerebral anoxia induced by NaNO2. Table 1 BET surface area, total pore volume and average pore diameter of CMS and MCM41. Table 2 Drug loading capacity (%) of NMP loaded CMS and NMP loaded MCM41 before and after washing. Table 3 Pharmacokinetic parameters obtained after oral administration of NMP, NMP-C3 and NMP-M3 in rats.

ACCEPTED MANUSCRIPT

Table 1 BET surface area, total pore volume, average pore diameter and NMP loading capacity of CMS and MCM41. SBET (m2/g)

Vt (cm3/g)

WBJH (nm)

CMS MCM41

732.16 997.54

0.94 0.93

3.98 3.36

AC

CE

PT E

D

MA

NU

SC

RI

PT

Sample

ACCEPTED MANUSCRIPT Table 2 Drug loading capacity (%) of NMP loaded CMS and NMP loaded MCM41 before and after washing.

Drug loading capacity (%)

NMP-C2 NMP-C3 NMP-M2 NMP-M3

68.27±2.28 65.01±1.12 66.80±3.34 56.29±1.34

AC

CE

PT E

D

MA

NU

SC

RI

PT

Drug loaded materials

ACCEPTED MANUSCRIPT

Table 3 Pharmacokinetic parameters obtained after oral administration of NMP, NMP-C3 and NMP-M3 in rats. Pharmacokinetic parameter

NMP

NMP-M3

NMP-C3

9.57±0.77

13.40±2.75

36.96±16.98

40.84±6.55

39.59±13.26

92.77±36.55

t1/2 (h)

24.55±4.04

28.408±13.55

61.98±37.77

3±0

3±0

8±0

0.29±0.01

0.80±0.11

0.438±0.06

100%

140.82%

386.22%

Cmax (mg/L) Relative bioavailability

AC

CE

PT E

D

MA

NU

SC

(%)

RI

Tmax (h)

PT

AUC0-∞ (mg/L*h) MRT0-∞ (h)

Graphics Abstract

Figure 1

Figure 2

Figure 3

Figure 4

Figure 5

Figure 6

Figure 7

Figure 8