Advances in Pharmaceutical Research


Research Article [Artilce ID-6657176]

Formulation & Development of Self-Micro Emulsifying Drug Delivery System (SMEDDS) for Oral Bioavailability Enhancement of a Low Soluble Anti-Diabetic Drug: Gliclazide

Anand Sonia1, Gupta Rishikesh1, Prajapati Sunil 1
1Institute of Pharmacy, Bundelkhand University Jhansi, UP, India
 

Received: 05 Dec, 2018; Revised: 04 Feb, 2019; Accepted: 15 Mar, 2019; Published: 30 Mar, 2019

DOI - 10.36218/APR/6657176
DOI URL- http://doi.org/10.36218/APR/6657176

Copyright 2019 This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Correnspondence should be addressed to Anand Sonia: soniaanand002@gmail.com


Abstract

The purpose of the present study was to enhance the solubility and to improve bioavailability of low water soluble (lipophilic) drugs by self-micro emulsifying drug delivery systems (SMEDDS) of Gliclazide via oral route resulting in increasing their clinical efficacy. Gliclazide is used for the treatment of Hyperglycaemia. In the present work, SMEDDS were successfully developed using different ratios of surfactant co-surfactant and oil’’. SMEDDS were prepared using Sunflower oil, Tween 80 (surfactant) and Poly ethylene glycol (PEG-400) (co-surfactant) and water. The range of pH is 7.4. Conventional SMEDDS are mostly prepared in a liquid form. SMEDDS can be orally administered in soft or hard gelatine capsules and form fine relatively stable oil- in- water (O/W) emulsions. The results indicated that SMEDDS containing Gliclazide can be a promising vehicle for enhanced oral bioavailability of Gliclazide.


INTRODUCTION

Oral intake is the most convenient and generally employed route of drug delivery, due to ease of administration, high patient compliance, cost-effectiveness, and flexibility in the dosage form. As a result, many of the generic drug companies are inclined more to produce bioequivalent oral drug products [1]. However, the major challenges with the design of oral dosage forms lie with their poor bioavailability. The most frequent causes of low oral bioavailability are because of poor solubility and low permeability. Approximately 40% of new drug candidates show less solubility in water, resulting in poor oral bioavailability, high intra and inter-subject variability, and lack of dose proportionality.

Thus, for those drugs, the absorption rate from the lumen of the gastrointestinal tract is controlled by dissolution. Therefore, it is necessary to prepare suitable formulations for improving solubility and bioavailability of such medicines. One of the most popular and commercially viable formulation approaches for solving these problems is Self-Micro Emulsifying Drug Delivery System (SMEDDS) which have attracted considerable attention of pharmaceutical scientists, who want to increase the oral bioavailability of such bad water-soluble medicines [2].

SMEDDS are isotropic mixtures of oils, surfactants and co-surfactants, which form oil-in-water micro emulsion in aqueous media under gentle agitation. The finely divided oily droplets, with a droplet size less than 50 nm, provide a large surface area for drug release and absorption. The oily phase allows the drug to be present in its solubilised state, thereby avoiding the slow and rate-limiting dissolution process of a hydrophobic drug [3]. This makes SMEDDS a promising approach for enhancing the absorption and bioavailability of poorly water-soluble drugs.

SMEDDS formulations are encapsulated in hard or soft gelatin capsules and administered as solid oral dosage forms. Some studies have suggested that the use of SMEDDS may not only increase the GI adsorption but also rectal and vaginal adsorption of poorly water-soluble drugs. Enhanced drug absorption was observed when using long chain triglycerides (LCT) compared with medium chain triglycerides (MCT) in the SMEDDS formulations. SMEDDS offers many advantages such as thermodynamic stability, spontaneous formation, better bioavailability and feasibility for preparation. Enhanced solubility and improved bioavailability are amongst the main advantage of SMEDDS [4].

Gliclazide comes in the class of the second-generation sulfonyluria which is used in the treatment of non-insulin-dependent diabetes. It is one of the most prescribed long-acting anti-hyperglycemic agents. Gliclazide is classified as BCS class II drug, which means it has high permeability and poor water solubility [5]. The poor water solubility of Gliclazide is responsible for its poor dissolution rate, which leads to variable absorption of Gliclazide. The bioavailability and in-vivo performance of Gliclazide is dependent on its dissolution rate. SMEDDS of Gliclazide with globule size <150 nm is prepared. Gliclazide, is an anti-diabetic drug in a class of medications known as sulfonylureas, closely related to sulfa drugs. It was developed in 1966 in a cooperative study between Boehringer Mannheim (now part of Roche) and Hoechst (now part of Sanofi-Aventis) [6].

MATERIAL AND METHODS

Materials

Gliclazide was received as gift sample from Lupin Research Park, Pune, India. Polyethylene Glycol (PEG) 4000, PEG 6000 and Polyvinyl Pyrrolidone (PVP K30), Transcutol P, PEG 300 Ethanol, Capryol-90, olive oil, castor oil, soybean oil, Sesame oil, oleic acid as oil, Tween 20, Span 80, span 20, Span 60, Cremophor-EL Propylene glycol, Brij-35, Labrasol, were purchased from Central Drug House, New Delhi, India. All other chemicals used in this study were of either pharmaceutical or analytical grade.

Screening of Excipients

  • Screening of Oil

  • Screening of Surfactant

  • Screening of Surfactant

  • Construction of Phase Diagram

  • Screening of Oil: To find out appropriate oil with good solubilizing capacity of API, the saturation solubility of API was investigated in some oils by shake flask method. An additional amount of API was added in the bottle containing 0.5 g of each solvent. After sealing, the mixture was vortexed using a cyclomixer for 10 min to facilitate proper mixing of API with the vehicles. The mixture was kept for 72 hours to achieve equilibrium at ambient temperature, and later, the mixtures were centrifuged at appropriate rpm for 15 minutes. The splitting of the supernatant was filtered through the membrane filter (0.45 μm) and was diluted with the mobile phase. Drug content was determined using high performance liquid chromatography (HPLC) technology.

  • Screening of Surfactant: To find appropriate surfactant with good solubilising capacity, after screening of oil emulsifying ability of different surfactants with the screened oil was investigated. 0.3g of surfactant and 0.3g of oil phase were weighed and vortexed for two minutes followed by warming at 40-45°C for 30 seconds, so we can obtain an isotropic mixture. 50 mg of isotropic mixture was taken and was already diluted with double distilled water filtered through the membrane filter (0.45 μm) in volumetric flask. The number of flask invasions was visually observed to form a clear emulsion. The resulting emulsion was allowed to stand for 2 hours after transmitting. The surfactant, which creates a clear emulsion with less number of inversions and more transmittance, was selected.

  • Screening of Co-Surfactant: To find appropriate co-surfactant with good solubilizing capacity, after screening of oil emulsifying ability of different co-surfactants with the screened oil was investigated. 0.2 g of co-surfactant and 0.3 g of oil phase were weighed and vortexed for two minutes followed by warming at 40-45°C for 30 seconds to obtain an isotropic mixture [7]. 50 mg of isotropic mixture was taken and was diluted with double distilled water filtered through the membrane filter (0.45 μm) in a volumetric flask. The number of flask invasions was visually observed to create a clear emulsion. The resulting emulsion was allowed to stand for 2 hours after observance of transmittance. The co-surfactant, which creates a clear emulsion with the lesser number of inversions and with more transmittance, was selected.

Preparation of SMEDDS

A series of SMEDDS was prepared different ratios of oil, surfactant, co-surfactant and the drug Gliclazide. The formulation was prepared by preparing the optimized ratio of the Smix, for which the surfactant and co-surfactant were weighed accurately and then vortexed for 5min. After that the Smix was placed in the oven at 50oC for 1 hour. After this, the oils of different proportions were added to the Smix, then these ingredients were put in the vortex for 5-10min and kept at 50oC in the oven for 1hr, so that it became an isotropic mixture. The drug was then finally loaded into these isotropic formulations and was vortexed until the formation of a clear solution [8].

UV spectrophotometric method

Gliclazide was analyzed by UV spectrophotometry Technique. Gliclazide was scanned at UV spectrophotometer and λmax was determined. The reported UV spectrophotometric method was verified and the partial validation for range and linearity, accuracy, accurate and analytical solution was determined up to 24 hours (Table 1).

Table 1: Optical spectrophotometric condition

Parameters

Setting Employed

Measurement mode

Absorbance

Solvent

Distilled water

λ max

226

Slit width

2.0nm

Calibration curve of gliclazide in methanol

Since gliclazide has solubility in methanol, the calibration curve was taken by solubilizing the drug in methanol. Gliclazide API showed main peak in methanol at 226 nm. As the gliclazide is insoluble in water, methanol was used to solubilize gliclazide and then the volume was made up with water. The UV spectrum of gliclazide in methanol is shown in Figure 1 and Table 2.

Figure 1: UV Spectrum of Gliclazide in Methanol

Table 2: Linearity of gliclazide in methanol at 226 nm wavelength

Conc. (μg/mL)

Absorbance (nm)

0

0

2.5

0.1089

5.0

0.2128

7.5

0.3183

10.0

0.4235

12.5

0.5345

15.0

0.623

17.5

0.7295

Equation

y= 0.0417x +0.0043

r2

0.9996

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

FTIR Spectra of Drug Gliclazide

The FTIR spectra of pure GLI (crystalline), amorphous GLI. The IR spectrum of GLIC (Figure 2) presents characteristic peaks at 3,273.57 and 3,192.58 cm−1 (N–H amide stretch), 3,112.55 cm−1 (C–H aromatic stretch), 2,949.59 cm−1 (C–H aliphatic stretch–asymmetric), 2,867.63 cm−1 (C–H aliphatic stretch–symmetric), 1,709.59 cm−1 (C=O, amide carbonyl stretch), 1,595.81 cm−1 (N–H amide bend), 1,590 and 1,473.35 cm−1 (C=C aromatic stretch), 1,348 cm−1 (S=O sulfonyl stretch), 1,240.97 cm−1 (C–N ring stretch, heterocyclic), and 811.885 cm−1 (p-phenyl group in fingerprint region) (figure 2).

Figure 2: FTIR Spectra of Drug Gliclazide

Dissolution Studies

The dissolution rate of pure gliclazide was studied using the basket method (USP type-II) at 37°C in 900 ml of 0.1N HCl (pH 1.2) at 100 rpm. Samples of 10 mg gliclazide samples were subject to test. To analyze the drug content using the UV spectrophotometer, the five ml samples of the dissolution medium were withdrawn and filtered at different time intervals. At each time of return, 5 ml of fresh PBS (pH 7.4) was added to the dissolution flask [9].

Drug Solubility

The excess amount of gliclazide was added to 2ml of each part stored in microtubes and the mixture was heated to 40°C on a water bath to dissolve the drug. The mixture was finally placed at the ambient temperature of 25°C for 48 hours to achieve equilibrium. The mixture was then centrifuged at 3000 rpm for 15 minutes. Aliquots of the supernatant were diluted with methanol, and the amount of drug was estimated using UV spectroscopic method. The solubility of the drug was determined by the calibration curve of Gliclazide in methane [10].

Thermodynamic Stability of SMEDDS

The purpose of thermodynamic stability is to evaluate the effect of phase separation and temperature variation on the SMEDDS formulations. SMEDDS were diluted with aqueous medium and centrifuged at 15,000 rpm for 15 minutes and formulation was observed visually for phase separation. Formulations were subjected to freeze thaw cycles (−20°C for 2 days followed by +40°C for 2 days). No change in the visual description of samples after freeze-thaw cycles. The formulations, which are thermodynamically stable, were selected for further characterization. The thermodynamic stability studies of liquid SMEDDS are the following:

  • Heating Cooling Cycle: Six cycles between refrigerator temperature -4oC and 45oC with storage at each temperature of not less than 48 h is studied. Those formulations, which were stable at this temperature, were subjected to centrifugation test.

  • Centrifugation Test: Passed formulations are centrifuged thaw cycle between 21oC and 25 oC with storage at each temperature for not less than 48 h is done at 3500 rpm for 30 min. Those formulations that did not show any phase separation, were taken for the freeze thaw stress test.

  • Freeze Thaw cycle: Freeze thawing was employed to evaluate the stability of formulations. The formulations were subjected to 3 to 4 freeze-thaw cycles, which included freezing at – 4°C for 24 hours followed by thawing at 40°C for 24 hours. Centrifugation was performed at 3000 rpm for 5 minutes. Those formulations passed this test showed good stability with no phase separation, creaming, or cracking. Only formulations that were stable to phase separation were selected for further studies [11].

  • Robustness to dilution: The robustness of Gliclazide SMEDDS at dilution was studied by diluting it 50, 100 and 1000 times with various dissolution media viz. water, SGF pH 1.2 and phosphate buffer pH 7.4. Diluted microemulsions were stored for 12 h and were observed for signs of phase separation or drug precipitation [12].

  • Self-Emulsification Assessment: Various compositions were classified according to the speed of the emulsification, the clarity of the resulting emulsion and the clear stability. Visual assessment was performed by drop-wise addition of the pre-concentrate (SMEDDS) into 250 ml of distilled water. It was done in a glass beaker at room temperature, and the material was gently stirred by magnetic stirrer at100 rpm.

  • Turbidimetric Evaluation: Growth of emulsion can be monitored by evaluating nephalo turbidometrics. Certain quantities of self-micro emulsifiers were added to a certain amount of suitable medium (0.1N hydrochloric acid) under constant stirring (50 rpm) on magnetic stirrer at ambient temperature, and increase in turbidity was measured by using turbidmeter. However, since the time required for full emulsification was very low, it was not possible to monitor the rate of change of turbidity (rate of emulsification) [13].

  • Viscosity Instrument: The viscosity was measured using a Brookfield synchro-lectic viscosimeter. A disk-shaped spindle (LV3) was used in the measurements. The rotation speed was set at 12 rpm. The spindle factor for the LV 3 spindle is 100. The viscosity measurement was carried out after each titration series. The spindle was cleaned after each race.

In-vitro Dissolution Studies

In vitro drug release studies were obtained using the USP type II dissolution apparatus. 900 ml of pH 7.5 phosphate buffer was placed in the dissolution container and the SMEDDS formulation was placed in a hard gelatin capsule and placed in the dissolution medium and stirred at 50 rpm at 37°C. 5 ml of samples were extracted at predetermined time intervals of 5 minutes (up to 1 hour). And the concentration of the drug was determined by a UV spectrophotometer at a wavelength of 22 nm. The volume removed from samples was replaced by fresh solution each time. The accumulated amounts released were plotted against time.

Stability Study

After determining the drug content and the release studies, the optimized formulation was loaded for the accelerated stability studies according to the ICH guidelines (40 ± 2 °C and 75 ± 5% RH) for a period of 3 months in a stability chamber (Thermolab, Mumbai, India). The optimized formulations were placed in USP Type I flint vials and sealed with bromobutyl rubber stoppers and sealed with aluminum caps. The samples were extracted at 15, 30, 60 and 90 days and evaluated for drug content and drug release.

RESULT AND DISCUSSION

Formulation & Development

The formulation was prepared by preparing the optimized Smix ratio first, for the same surfactant and co-surfactant were accurately weighed and then stirred for 5 minutes after Smix was placed in the oven at 50 ° C for 1 hour. After that oil with different ratio was added to Smix then these formulations were vortexed for 5-10min and placed in oven at 50oC for 1hr, so that an isotropic mixture was formed. Then drug was loaded to prepared isotropic formulations at the end and vortexed by shaker until clear solution was formed.

Construction of Ternary Phase Diagrams

Phase diagrams were constructed to obtain the proportion of components that can result in maximum microemulsion existence area. These diagrams were constructed with oil, surfactant/co-surfactant and water using the titration method at room temperature. The procedure consisted of preparing solutions of different ratio of surfactant to co-surfactant by weight such as: 1:1, 2:1, 3:1 etc, those solutions then vortexed for 5 min and placed at 50°C for 1 h so that an isotropic mixture was obtained. Each of these solutions was then used to prepare a mixture containing oil and Smix (mixture of surfactant and co-surfactant) in the following proportions by weight: 1:9, 2:8, 3:7, 4:6, 5:5, 6:4,7:3, 8:2, 9:1 and after preparation vortexed for 5 min followed by placing in oven at 50°C for 1 hr.

All the mixtures were then placed at room temperature for 24 h. Water from 5% to 95% of the mixture was added at intervals of 10-15 minutes to each of the mixtures with stirring on a magnetic stirrer. After each addition, the appearance of the mixtures (turbid or transparent) was observed. The turbidity of the samples would indicate the formation of a coarse emulsion, while a clear isotropic solution would indicate the formation of a microemulsion. Percentage of oil, Smix and water at which clear mixture was formed were selected and the values were used to prepare ternary phase diagram (Figure 4).

Self-Emulsification time

The evaluation of self-microemulsification wasdone through visual evaluation. 100 ml of water and 0.1 N HCl solution were taken as medium, which was stirred with a magnetic stirrer at 100 rpm at 37 ± 0.5 ° C and 1 ml of formulated SMEDDS was poured into the medium and the contents mixed gently at 100 rpm and the time required to form the microemulsion was determined after dilution of SMEDDS with water (Table 3).

Fig. 4: Pseudo ternary phase diagram of sunflower oil with tween 80: transcutol A (1:1), (1:2), (2:1)

Table 3: Self emulsification times of SMEDDS formulation

Formulation

Code

F5

F6

F7

F8

F9

F10

0.12 N HCl

Emulsification time sec

23

28

29

45

67

26

Tendency for emulsification

Good

Good

Good

Good

Good

Good

Phosphate

Emulsification time sec

20

31

34

50

61

22

Buffer (pH 6.4)

Tendency for emulsification

Good

Good

Good

Good

Good

Good

 

 

 

 

 

 

 

 

Characterization of SMEDDS Formulation

  • Visual Observation, phase separation of Self Micro Emulsification:

200ml of distilled water was taken in a beaker and each Gliclazide loaded SMEDDS formulation was dropped into water and this is maintained at 37°C, and the diluted preparation was vortexed for 1min. This preparation was stored for 24 hours and then phase separation and precipitation can be observed visually.

Mixtures exhibiting a negligible phase separation during the 2hr period were used for subsequent studies. It gave the information about stability and viability of the formed microemulsion (Table 4).

  • Droplet size analysis:

Gliclazide SMEDDS was diluted with distilled water and stirred slowly using magnetic stirrer to mix thoroughly. Zeta Sizer was used to determine the droplet size of resulting microemulsion. Size of droplet is measured by photon correlation using Malvern Zeta Sizer (Table 5 and Figure 5).

  • Zeta potential measurements:

The stability of the emulsion is directly related to the magnitude of the surface charge. The zeta potential of the diluted SMEDDS formulation was measured using a (Malvern Nano Zetasizer instrument). The SMEDDS were diluted with a ratio of 1:20 v/v with distilled water and mixed for 1 m using a magnetic stirrer (Table 5 and Figure 6).

  • Particle Size Measurement: The particle size distribution was determined using Dynamic Light Scattering (DLS) technique. DLS technique, also known as Photon Correlation Spectroscopy, is one of the most widely used methods to measure the size of micro particles. This technique assumes that all the particles are in Brownian motion in the solution and that all the particles are very small and spherical.

Table 4: Optical clarity values of SMEDDS formulation.

Formulations

F1

F4

F7

F8

F9

F10

0 Hours (absorbance)

0.756

0.234

0.456

0.467

0.045

0.225

24Hours (absorbance)

0.786

0.245

0.467

0.471

0.047

0.220


 

 

 

 

Table 5: Droplet size, zeta potential and zeta sizer values of SMEDDS.

Formulations

Particle Size(mm)

Zeta Potential(V)

PDI

FG7

55

-15.2

0.2400

FG8

19.43

-19.8

0.332

FG9

17.61

-22.3

0.222

FG10

62.32

-19.4

0.176

 

 

 

 

 

 

 

 

Table 6: Spectroscopic Analysis of Drug

Solvent

Partition coefficient

N-octanol/ distilled water

1.82

N-octanol/ SGF

1.56

 

 

 

 

 

Scattering of light (normally a laser) takes place when particles are hit by light. The particle size can be determined based on the physical properties of the scattered light: the angular distribution, frequency shift, the polarization and the intensity of the light.

Figure 5: Droplet size distribution of the nanoemulsion

Figure 6: Zeta potential distribution of nanoemulsion obtained from Liquid SMEDDS

  • Refractive Index:

Refractive index of the placebo SMEDS and drug-loaded SMEDDS was determined with an Abbe-type thermo stated refractometer.

  • Transmittance:

Transmittance of the Gliclazide SMEDS were measured against distilled water with a UV–visible Spectrophotometer.

  • Cloud Point Measurement:

The formulated SMEDDS was diluted with 50ml water in a beaker which was placed on a water bath with gradually increasing temperature until the diluted formulation turned cloudy. It mainly insists about the stability of microemulsion at the temperature of body.

  • Microscopy:

Gliclazide was found to have crystalline structure with rectangular crystals. Even though drug was micronized with PSD approx. 7 microns, the clear rectangle crystals were seen under microscope (Figure 8).

Gliclazide (Dry condition under 10X)