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Montmorillonite-Alginate Nanocomposites as a Drug Delivery System: Intercalation and In Vitro Release of Vitamin B 1 and Vitamin B6 Bhavesh D. Kevadiya, Ghanshyam V. Joshi, Hasmukh A. Patel, Pravin G. Ingole, Haresh M. Mody and Hari C. Bajaj J Biomater Appl 2010 25: 161 originally published online 8 September 2009 DOI: 10.1177/0885328209344003 The online version of this article can be found at: http://jba.sagepub.com/content/25/2/161

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Montmorillonite-Alginate Nanocomposites as a Drug Delivery System: Intercalation and In Vitro Release of Vitamin B1 and Vitamin B6 BHAVESH D. KEVADIYA, GHANSHYAM V. JOSHI, HASMUKH A. PATEL, PRAVIN G. INGOLE, HARESH M. MODY AND HARI C. BAJAJ* Discipline of Inorganic Materials and Catalysis, Central Salt and Marine Chemicals Research Institute (Council of Scientific & Industrial Research CSIR), Gijubhai Badheka Marg, Bhavnagar 364 002, Gujarat, India

ABSTRACT: Sustained intestinal delivery of thiamine hydrochloride (Vitamin B1; VB1) and pyridoxine hydrochloride (Vitamin B6; VB6) seems to be a feasible alternative to existing therapy. The vitamins (VB1/VB6 ) intercalated in montmorillonite (MMT) and intercalated VB1/VB6-MMT hybrid is further used for synthesis of VB1/VB6-MMT-alginate nanocomposite beads by gelation method and in vitro release in the intestinal environment. The structure and surface morphology of the synthesized VB1/VB6-MMT hybrid, VB1/VB6-alginate and VB1/VB6-MMT-alginate nanocomposite beads were characterized by XRD, FT-IR, TGA and SEM. In vitro release experiments revealed that the VB1/VB6 releases suddenly from VB1/VB6-MMT hybrid and is pH dependent. The controlled release of VB1/VB6 from VB1/VB6-MMT-alginate nanocomposite beads was observed to be controlled as compared to their release from VB1/VB6-MMT hybrid and VB1/VB6-alginate beads. KEY WORDS: pyridoxine hydrochloride, thiamine hydrochloride, alginate, montmorillonite, nanocomposites.

*Author to whom correspondence should be addressed. E-mail: [email protected] Figure 4 appears in color online: http://jba.sagepub.com

JOURNAL OF BIOMATERIALS APPLICATIONS Volume 25 — August 2010 0885-3282/10/02 0161–17 $10.00/0 DOI: 10.1177/0885328209344003 The Author(s), 2010. Reprints and permissions: http://www.sagepub.co.uk/journalsPermissions.nav


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INTRODUCTION

F

or successful therapy, a drug should be delivered at appropriate sites for a long duration, so as to have the maximum pharmacological activity. Special attention has been paid to find an approach to control the rate of drug release by means of a carrier where the drug is dispersed or integrated in an inert matrix. Recently, stimuli-responsive hydrogels and clay minerals have attracted great interest due to their potential application in pharmaceutical sciences [1–4]. Vitamin B1 (thiamine hydrochloride; VB1) and Vitamin B6 (pyridoxine hydrochloride; VB6) are water-soluble B vitamins and they are required for the metabolism of carbohydrates, fats, and proteins. VB1 deficiency leads to beriberi, which is characterized by an accumulation of body fluids, tenderness, paralysis, and eventually death. Symptoms of VB6 deficiency include muscle weakness, nervousness, irritability, depression, difficulty concentrating and short-term memory [5,6]. The main perception in modified drug release technology is that any pharmaceutical dosage form should be designed to provide therapeutic levels of drug to the site of action and maintained throughout the treatment. One class of drug delivery vehicle that has received more attention in recent years is layered materials, which can accommodate therapeutic compounds between their layers [7–9]. Intercalation of drug molecules into layered materials provides a useful and convenient route to prepare organic-inorganic hybrids that contain properties of both the inorganic host and organic guest in a single matrix and can be used as drug carrier [10]. One promising inorganic material as a host compound is montmorillonite (MMT) belongs to smectite clay family. Smectite clays have a layered structure and layer is constructed from tetrahedrally coordinated silica atoms fused into an edge-shared octrahedral plane of aluminum. It is not only harmless but also very stable both in acidic and basic media [11]. Alginate has been extensively used as a vehicle for control release of therapeutic agents. The ability of alginate to form a gel in the presence of multivalent ions has been exploited to prepare multi-particulate systems, incorporating numerous drugs, proteins, cells, or enzymes. Alginate, a linear, naturally occurring polysaccharide extracted from brown sea algae contains D-mannuronic (M) and L-gulcuronic (G) acids which are arranged in homo polymeric MM or GG blocks separated by blocks with an alternating sequence. Alginate is a hydrophilic, biocompatible, mucoadhesive, nontoxic, and inexpensive polymer and can be used for controlled delivery of drugs [12]. Theoretically, alginate shrinks at low pH (gastric environment) and the encapsulated drugs


Montmorillonite-Alginate Nanocomposites as a Drug Delivery System 163

cannot be released. It was reported that the biological activity of drugs can be retained in the calcium-cross-linked alginate encapsulation process. pH-sensitive hydrogels have attracted increasing attention due to their unique properties. Swelling of such hydrogels in the stomach is negligible and thus the drug release is also minimal. Due to increase in pH as the hydrogels pass down the intestinal tract, the degree of swelling increases [13,14]. Compared to other polymers, alginate offers additional advantages in terms of safety and biocompatibility, availability, and low cost and thus serves as an excellent candidate for the development of oral drug formulation [15] Since 1960s it was observed that oral absorption of several drugs was reduced by co-administration of clay-based intestinal adsorbents or by the presence of clay stabilizers in liquid formulations [16].Wang et al. synthesized biopolymer/MMT nanocompoistes as carrier for control release of Bovine serum albumin and observed that the nanocomposite nanoparticles exhibited higher drug-loaded capacity and better drug controlled release properties at certain MMT loading [17]. Joshi et al. studied intercalation of timolol maleate into the interlayer of MMT and observed that intercalation of timolol maleate into MMT depends on the pH of the interaction medium [18]. Dong et al. synthesized the poly (D, L-lactide-co-glycolide)-MMT nanoparticles through the emulsion/solvent evaporation method for oral delivery of paclitaxel and concluded that oral chemotherapy by PLGA-MMT nanoparticles seems to be feasible [19]. Bonina et al. illustrated the adsorption and release of Fe(III)salicylate complex on bentonite and its application as topical drug [20]. Lin et al. studied the intercalation of 5-fluorouracil with MMT as drug carrier [21]. Fejer et al. reported the interaction of monovalent cationic drugs promethazine chloride, benzalkonium chloride and buformin hydrochloride with MMT [22]. In the present study, we focused on intercalation of VB1 and VB6 into the interlayer of MMT under different reaction conditions, such as pH and initial concentration of VB1 and VB6, which are further compounded with alginate to obtain VB1/VB6-MMT-alginate nanocomposites beads. The release behavior of VB1 and VB6 from the composite beads was studied in phosphate buffer solution of pH 7.4. MATERIALS AND METHODS

Materials Thiamine hydrochloride, C12H17ClN4OS HCl, (VB1) Pyridoxine hydrochloride, C8H11NO3 HCl (VB6), Alginic acid sodium salt and


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cellulose acetate dialysis tube (MW: 07014) were purchased from SigmaAldrich, USA. CaCl2, (fused) HCl, KCl, KH2PO4, NaCl, and NaOH were purchased from S. D. Fine Chemicals, India and were used as received. The MMT rich bentonite clay was collected from Akli mines, Barmer district, Rajasthan, India. Deionized water was obtained from Milli-Q Gradient A10 water purification system. Methods Purification of MMT 300 g of raw bentonite was dispersed in 3 L of 0.1 M NaCl solution and stirred for 12 h. To obtain Na-MMT, the slurry was treated with NaCl for three times. Finally, the slurry was centrifuged and washed with MilliQ water until free from chloride ion as tested by AgNO3 solution [23]. NaMMT was purified by sedimentation technique as described earlier [24]. According to the Stokes law of sedimentation, the purified MMT was obtained by dispersing 150 g of Na-MMT in 10l Milli-Q water and collecting the supernatant dispersion of particles 52 mm after the precalculated time (10 h) and height (15 cm) at 308C. The MMT slurry was dried at 90–1008C and ground to pass through 200 mesh sieve (ASTM). The cation exchange capacity (CEC) of MMT was measured by the standard ammonium acetate method at pH 7 [23] and was 91 mequiv. /100 g dry weight of MMT (dried at 1108C).

Intercalation of VB1 and VB6 into Interlayer of MMT at Different PH The experiments were carried out to determine the optimum pH value for intercalation of VB1/VB6 into the interlayer of MMT. These studies were performed by treating VB1/VB6 and MMT mixture at different pH and constant temperature, time and concentration. 100 mg of MMT was dispersed in 20 mL of Milli-Q water containing 105 mg or 26.3 mg of VB1 and VB6 respectively. The suspensions were shaken for 1 h at 508C (VB1) and at 358C (VB6). The reaction mixtures were filtered and the concentration of VB1/VB6 in the filtrate was determined by UV-Vis spectroscopy at max ¼ 242 nm and 291 nm for VB1 and VB6 respectively. The intercalation studies were performed in triplicate and the average values were used in data analysis. Intercalation of VB1/VB6 into Interlayer of MMT at Different Initial Concentration To obtain maximum intercalation of VB1/VB6 into the interlayer of MMT, experiments were carried out at different initial concentration



and constant time, pH and temperature. 100 mg of MMT was dispersed in 20 mL of Milli-Q water containing various amounts of VB1/VB6. The dispersions were shaken for 1 h at pH 7.0 and 508C (VB1) and pH 3.2 and 358C (VB6). The reaction mixtures were filtered and the concentration of VB1/VB6 in the filtrate was determined by UV-Vis spectroscopy.

Preparation of VB1/VB6-MMT-Alginate Nanocomposite Beads 750 mg of alginate was dissolved in 25 mL of Milli-Q water and the solution was warmed to 708C on a water bath. The desired quantity of VB1/VB6-MMT hybrid (200 meshes) were added into alginate solution and slowly heated on a water bath to 808C [25]. This emulsion was dropped using a peristaltic pump (Master flex L/S 7518-00, ColeParmer, USA) with the help of a 20-guage hypodermic needle fitted with a rubber tubing (falling distance 2 cm, pumping rate 1.5 mL/min) into 250 mL of 5% (w/v) calcium chloride solution, which was gently agitated with a magnetic stirrer [26]. The formed beads were allowed to stand in solution for 20 min to be cured and then collected by filtration and were washed thrice with Milli-Q water. A similar procedure was followed for the preparation of VB1/VB6-alginate beads. We have optimized the claydrug hybrid: alginate ratio in order to obtain stable beads, having a minimum amount of alginate and controlled release behavior. Encapsulation Efficiency For determining the encapsulation efficiency, 16.6 mg of VB1-MMT hybrid, 30 mg of VB1-alginate beads, and VB1-MMT-alginate nanocomposite beads were added into 50 mL of phosphate buffered of pH 7.4. The dissolution medium was stirred overnight to accomplish the dissolution of VB1. Solutions were filtered and VB1 content was estimated by UV-vis spectrophotometer. The VB1 encapsulation efficiency, EE (%) was determined as [26]. EE ¼ ðAQ=TQÞ 100 where AQ is the actual quantity of VB1 released from VB1-MMT hybrid, VB1-alginate beads, and VB1-MMT-alginate nanocomposite beads. TQ is quantity of VB1 present in VB1-MMT hybrid, VB1-MMT-alginate beads and VB1-MMT-alginate nanocomposite beads (amounts of initial loading). A similar method was followed to determine the encapsulation efficiency of VB6 from VB6-MMT hybrid, VB6-alginate beads, and VB6MMT-alginate nanocomposite beads.


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Characterization Powder X-ray diffraction (XRD) analysis was carried out with a Phillips powder diffractometer X’Pert MPD using PW3123/00 curved ˚ ) radiation with a slow scan of 0.38/s in Cu-filtered Ni-Ka ( ¼ 1.54056 A 2 range of 2–108. Fourier transform infrared spectra (FT-IR) were measured with Perkin-Elmer, GX-FTIR as KBr pellet over the wavelength range 4000–400 cm1. Thermogravimetric analysis was carried out within 30–8008C at the rate 108C/min in the nitrogen flow of 60 mL/min using Mettler -Toledo, TGA/SDTA 851e. UV-Vis absorbance of VB1 and VB6 solutions were measured at a characteristic max ¼ 242 nm and 291 nm respectively by UV-Vis spectrophotometer (Cary 500, Varian) equipped with a quartz cell having a path length of 1 cm. The morphology of nanocomposite was observed under scanning electron microscope (SEM), LEO-1430VP, UK. In Vitro Release Studies In vitro release studies were performed in USP six stage dissolution rate test apparatus (Thermonik, Campbell electronics, India) by using dialysis bag technique [27]. Phosphate buffer solution of pH 7.4 was prepared by mixing 1000 mL of 0.1 M KH2PO4 and 782 mL of 0.1 M NaOH. The dialysis sac was equilibrated with the dissolution medium for few hours prior to experiments. 300 mg of VB1-MMT hybrid, 200 mg of VB1-alginate beads, and VB1-MMT-alginate nanocomposite beads were suspended in dialysis bags containing 5 mL of phosphate buffer solution, which was sealed at both ends. The dialysis bag was set into baskets and rotated at 100 rpm and 37 0.58C in 900 mL phosphate buffer solution of pH 7.4. At 30 min of time intervals, 5 mL of the dissolution media were taken and the VB1 concentration was determined by UV-Vis spectroscopy. A similar procedure was used for VB6 release from VB6-MMT hybrid, VB6-alginate beads, and VB6-MMT-alginate nanocomposite beads. These studies were performed in triplicate for each sample and the average values were used in data analysis. RESULTS AND DISCUSSION

Intercalation of VB1/VB6 into Interlayer MMT at Different pH Values The intercalation of VB1 in MMT increases with an increase in the pH of the interaction medium from 2 to 7, while intercalation of VB6 in MMT decreases with increase in the pH of the interaction medium (Table 1). This phenomenon can be explained by the effect of pH on the


Montmorillonite-Alginate Nanocomposites as a Drug Delivery System 167 Table 1. Effect of pH on intercalation of VB1 and VB6 into MMT. Sl. No.

pH

VB1 intercalated into MMTa (%)

VB6 intercalated into MMTb (%)

1 2 3 4 5 6 7 8 9

2.0 3.2 4.0 5.0 6.0 7.0 8.0 9.0 10.0

10.2 0.51 10.5 0.48 11.1 0.44 11.6 0.54 13.5 0.52 23.5 1.04 12.8 0.61 11.7 0.51 1.11 0.55

9.2 0.46 13.9 0.2 12.0 0.6 7.4 0.31 1.5 0.07 1.4 0.7 -

a MMT ¼ 100 mg; VB1 ¼ 105 mg; reaction volume 20 mL; reaction temperature ¼ 508C and reaction time ¼ 1 h b MMT ¼ 100 mg; VB6 ¼ 26.3 mg; reaction volume 20 mL; reaction temperature ¼ 358C and reaction time ¼ 1 h

structure of VB1 as mentioned earlier [28]. We have selected pH 7 for further study because, at this pH, the intercalation was found to be maximum and also due to fact that denaturation of thiamine in alkaline medium can take place [29]. The amount of VB1 intercalated into MMT is less at low pH due to the formation of divalent cationic species. The intercalation of VB6 into MMT increases with a decrease in the pH of the interaction medium due to increase in the protonated form of VB6. Effect of VB1/VB6 Concentration on Intercalation Effect of initial concentration of VB1 and VB6 on intercalation into MMT shows that the intercalation of VB1/VB6 increased with increase in the initial concentration of VB1 and VB6, may be due to greater concentration gradient at the initial stage. The maximum amount of VB1 and VB6 intercalated into MMT is 235 mg/g and 139 mg/g of MMT respectively (Table 2). XRD and FT-IR Analysis The XRD pattern of MMT, VB1/VB6-MMT hybrid, VB1/VB6-MMTalginate nanocomposite is shown in Figure 1. The basal spacing of MMT is 1.18 nm (2 ¼ 7.48), which increases to 1.55 nm (2 ¼ 5.78) on the intercalation of VB1. The shifting of [001] plane to the lower angle suggests that the VB1 is intercalated into interlayer of MMT. Interaction of VB1MMT and alginate resulted in tactoid formation as observed from broadening of the [001] peak and decrease in basal spacing (1.33 nm at 2 ¼ 6.648).


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The basal spacing for VB6-MMT hybrid and VB6-MMT-alginate nanocomposite is 1.3 nm (2 ¼ 6.98) and 1.26 nm (2 ¼ 7.028), respectively. From the XRD study it may be concluded that in both nanocomposite beads, the alginate is not intercalated into the MMT but it interacted with the surface hydroxyl groups. Alginate and MMT formed electrostatic and intermolecular hydrogen bonds, which brought about numerous points of Table 2. Effect of initial concentration of VB1 and VB6 on the intercalation of VB1 and VB6 into MMT. Sl. No.

Initial VB1 (mg)a

Intercalated VB1 (mg)

Initial VB6 (mg)b

Intercalated VB6 (mg)

7.0 21.0 35.0 70.0 84.0 105.0 153.0

6.4 15.7 17.3 18.6 20.2 20.8 23.5

3 6 9 12 24 36 60

1.7 4.7 7.4 9.6 12.5 13.1 13.9

1 2 3 4 5 6 7 a

MMT ¼ 100 mg; reaction volume 20 mL; reaction temperature ¼ 508C and reaction time ¼ 1 h MMT ¼ 100 mg; reaction volume 20 mL; reaction temperature ¼ 358C and reaction time ¼ 1 h

b

(b) 500

400

400 MMT

Intensity (cps)

Intensity (cps)

(a) 500

300

200

MMT 300

200 VB6–MMT

VB1–MMT 100

100 VB1–MMT–alginate

0 2

3

4

5

6

7

8

VB6–MMT–alginate

0 9 10

2

3

4

5

6

7

2(θ)

8

9 10 2(θ )

Figure 1. XRD pattern of (a) MMT, VB1-MMT and VB1-MMT-Alginate nanocomposite beads and (b) MMT, VB6-MMT and VB6-MMT-Alginate nanocomposite.



contact to create a three-dimensional network, and alginate chains could also serve as a bridge between neighboring silicate layers when a higher content of MMT was incorporated. FT-IR spectrum of VB1, MMT, alginate, and VB1/VB6 -alginate, VB1/ VB6-MMT-alginate nanocomposites beads are shown in Figure 2. Asymmetric and symmetric stretching vibrations at 1617 and 1417 cm1 are due to carboxyl anions, and 1030 cm1 for cyclic ether bridge oxygen stretching of alginate. MMT shows that bands at 3620 and 3698 cm1 are due to –OH band stretch for Al–OH and Si–OH. The shoulders and broadness of the structural –OH band are mainly due to contributions of several structural –OH groups occurring in the clay [30]. The overlaid absorption peaks in the region of 1640 cm1 in the FT-IR spectrum of VB –MMT–alginate 6

VB –MMT–alginate 1

VB –alginate 6

VB –alginate 1

%T

VB –MMT 6

VB –MMT 1

MMT–alginate

MMT

4000.0

3000

2000

1500

1000

400.0

cm-1 Figure 2. FT-IR spectra of MMT, MMT-alginate, VB1/VB6-alginate, and VB1/VB6-MMT, VB1/VB6-MMT-alginate nanocomposites.


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MMT is attributed to –OH bending mode of adsorbed water. The characteristic peak at 1115 cm1 is due to Si–O stretching (out-of-plane) for MMT (Patel et al. 2007). The bands at 1350 and 1443 cm1 is attributed to C-H (-CH3 pyrimidine ring) and C-H (-CH3). The bands at 1532, 1550, and 1629 cm1 are due to stretching of pyrimidine ring. The band at 1662 cm1 is attributed to bending of NH (NH2) and N1H protonated. The band at 2928 cm1 is due to stretching of CH (CH2CH2). The band at 1385 cm1, 1280 cm1 and 1230 cm1 is due to C¼C, C¼N stretching, and C-O-H and N-H bending, respectively. Thaned et al. explained the change in the wave number of COO while alginate interacts with MMT. Incorporation of MMT into alginate caused a shift to a higher wavenumber and the intensity of COO stretching peaks of alginate decreases [31]. The negative charge of the carboxyl groups may have an electrostatic interaction with the positively charged sites at the edges of MMT. The OH stretching peak of the silanol group (SiOH) at 3698 cm1 disappeared in the spectra of MMT-alginate nanocomposites, and the OH stretching peak of alginate shifted to a higher wave number, which is an evidence of the intermolecular hydrogen bonding and electrostatic forces between alginate and MMT as confirmed by FT-IR studies. Thermal Analysis and SEM The TGA curves of MMT and VB1/VB6-MMT hybrid, VB1/VB6alginate, and VB1orVB6-MMT-alginate nanocomposites are shown in Figure 3(a) and (b). The TGA curves of MMT shows two distinct steps, the first one at 51508C is due to the free water evaporation and second weight loss at 550–7008C is due to the release of structural OH group from MMT VB1/VB6-MMT hybrid, VB1/VB6-alginate and VB1/VB6MMT-alginate nanocomposites are started to degrade from 2008C. Three step weight loss patterns is observed in these samples, mainly due to loss of adsorbed water (51508C), decomposition of alginate and/or vitamins (200–5008C) and removal of hydroxyl group from edges of MMT (550–7008C). The surface and internal morphology of VB1-alginate beads and VB1MMT-alginate nanocomposites beads are depicted in scanning electron micrograph images Figure 4. The VB1-alginate beads displayed a smooth surface while the surface of VB1-MMT-alginate nanocomposite beads are rough. This may be due to the reinforcement effect of MMT in the alginate matrix. It can be seen that both VB1-alginate and VB1-MMTalginate nanocomposite beads had a porous structure as shown in


Montmorillonite-Alginate Nanocomposites as a Drug Delivery System 171 (a) 100 MMT VB1–MMT

Weight loss (%)

80

60 VB1–MMT–alginate 40 VB1–alginate

20

0 25 (b)

225

625 425 Temperature (°C)

825

100 MMT

Weight loss (%)

80

VB6–MMT

60 VB6–MMT–alginate 40

VB6–alginate

20

0

25

225

425 Temperature (°C)

625

825

Figure 3. Thermogravometric analysis of (a) MMT, VB1-alginate, VB1-MMT, VB1-MMTalginate nanocomposites and (b) MMT, VB6-alginate, VB6-MMT, VB6-MMT-alginate nanocomposites.

Figure 4 (aIII) and (bIII). The morphology of VB1-alginate beads also changes from homogeneous and smooth structure to heterogeneous and coarse structure after interaction of MMT in VB1-MMT-alginate nanocomposite beads. This effect is believed to play an important role in control release of VB1 from VB1-MMT-alginate nanocomposite beads. A similar morphology was observed for VB6-alginate and VB6-MMTalginate nanocomposite beads.


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aI

bI

EHT=20.00kv WD=15 mm Chamber=5.40e–001Pa 100 µm* Mag=100X

EHT=20.00kv WD=15 mm Chamber=5.40e–001Pa 100 µm* Mag=100X

aII

bII

1 µm*

EHT=20.00kv WD=15 mm Chamber=5.40e–001Pa Mag=6.29KX

1 µm*

EHT=20.00kv WD=15 mm Chamber=5.40e–001Pa Mag=6.29KX

bIII

aIII

30 µm*

EHT=20.00kv WD=15 mm Chamber=5.40e–001Pa Mag=120X

30 µm*

EHT=20.00kv WD=15 mm Chamber=5.40e–001Pa Mag=120X

Figure 4. SEM images of (aI) VB1-alginate bead and (bI) VB1-MMT-alginate nanocomposite bead scale: 100 mm; (aII) surface images of VB1-alginate bead and (bII) VB1-MMTalginate nanocomposite bead, scale: 1 mm; (aIII) cross-section of VB1-alginate bead (bIII) VB1-MMT-alginate nanocomposite bead, scale: 30 mm.

The VB1/VB6 Release Studied In vitro release profiles of VB1/VB6 from VB1/VB6-MMT hybrid, VB1/VB6-alginate beads and VB1/VB6-MMT–alginate nanocomposite beads were evaluated in phosphate buffer of pH 7.4 at 37 0.58C


Montmorillonite-Alginate Nanocomposites as a Drug Delivery System 173 (a)

80

VB1 release (%)

VB1–MMT 60 VB1–MMT–alginate 40

20 VB1–alginate 0 0

2

4

6

8

Time (h)

(b) 100

VB6–MMT

VB6 release (%)

80 60 VB6–MMT–alginate 40

20

0

VB6–alginate

0

2

4 Time (h)

6

8

Figure 5. Release profile of (a) VB1 from the VB1-MMT hybrid, VB1-Alginate beads and VB1-MMT-Alginate nanocomposite beads and (b) VB6 from the VB6-MMT hybrid, VB6Alginate beads and VB6-MMT Alginate nanocomposite beads in simulated intestinal fluid (pH 7.4) at 37 0.58C.

(Figure 5(a) and (b)). Initially, VB1 release was rapid up to 3 h and 56% of the intercalated VB1 was released. 64% of VB1 was released in 6.5 h from VB1–MMT hybrid. While in the case of VB6, the release rate was very fast as within 1 h, 83% of VB6 was released from VB6-MMT hybrid. Total release of VB6 is 87% from VB6-MMT hybrid. Up to 6% of VB1/VB6 was released from VB1/VB6-alginate beads within 1.5 h and 7.2% and 10.8% VB1 andVB6 was released after 5 h, which remained constant up to 8 h. In the case of VB1/VB6-MMT-alginate nanocomposite beads, the release of VB1/VB6 was 44-48% in 5.5 to 8 h. These results indicate that the release rate is faster in VB1/VB6-MMT


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Table 3. Formulation codes and different characteristic of VB1/VB6/MMT/alginate nanocomposite beadsa. Formulation code

MMT (mg)

VB1/VB6 (mg)

Alginate (mg)

Diameter beads (mm)

Encapsulation efficiency (%)

VB1-MMT VB1-MMT-alginate VB1-alginate VB6-MMT VB6-MMT-alginate VB6-alginate

100 71.6 100 76.6 -

18.8 13.4 140.6 11 8.4 82.5

750 750 750 750

0.84 0.71 0.83 0.73

64.6 82.5 23.9 87.2 89.7 28.2

a

Total volume for formulation was 25 mL.

hybrid and VB1/VB6–alginate beads as compared to VB1/VB6-MMTalginate nanocomposite beads. Controlled release of VB1/VB6 was observed in the case of VB1/VB6-MMT-alginate nanocomposite beads due to synergic effect of alginate and MMT in nanocomposite beads. The encapsulation efficiency of VB1/VB6-MMT–alginate nanocomposite beads is also higher as compared to VB1/VB6-alginate beads (Table 3). This may be due to strong hydrogen bonding between VB1/VB6 and surface hydroxyl group of the MMT and intercalation of VB1/VB6 along with alginate. The release process of VB1/VB6 from VB1/VB6-MMT hybrid may be interpreted on the basis of the ion-exchange process between the intercalated cations and cations present in the buffer solutions. The release experiments clearly demonstrate that VB1/VB6 release rate is highly dependant on the presence of alginate and MMT. CONCLUSIONS

The VB1/VB6 was effectively intercalated into MMT and was ionically bound into MMT. The maximum amount of VB1/VB6 intercalated into MMT is 235 and 139 mg/g respectively. VB1/VB6–alginate and VB1/VB6MMT–alginate nanocomposite beads were successfully synthesized by gelation method. XRD study reveals formation of intercalated nanocomposites and results in stiff and micro-porous nanocomposite beads. In vitro release profiles of VB1 and VB6 from VB1/VB6-MMT hybrid, VB1/VB6-alginate and VB1/VB6-MMT–alginate nanocomposite beads are carried out in phosphate buffer of pH 7.4 showed that VB1 (64%) and VB6 (87%) was released from VB1/VB6-MMT hybrid. 7.2% of VB1 and 10.8% of VB6 was released from VB1/VB6-alginate beads. In case of VB1/ VB6-MMT–alginate nanocomposites beads, the rate of release was controlled as compared to VB1/VB6-MMT hybrid and 48% of VB1 and



44% of VB6 was observed to be released in 8 h. From these studies it is clear that MMT and alginate can be used as the controlled release carrier for VB1 and VB6. ACKNOWLEDGEMENTS

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