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Encapsulation of Antibiotic Levofloxacin in Biocompatible Microemulsion Formulation: Insights from Microstructure Analysis Muhammad Faizan Nazar, Muhammad Atif Saleem, Sana Nawaz Bajwa, Basit Yameen, Muhammad Ashfaq, Muhammad Nadeem Zafar, and Muhammad Zubair J. Phys. Chem. B, Just Accepted Manuscript • DOI: 10.1021/acs.jpcb.6b09326 • Publication Date (Web): 22 Dec 2016 Downloaded from http://pubs.acs.org on December 27, 2016

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The Journal of Physical Chemistry

Encapsulation of Antibiotic Levofloxacin in Biocompatible Microemulsion Formulation: Insights from Microstructure Analysis

Muhammad Faizan Nazar,a* Muhammad Atif Saleem,a Sana Nawaz Bajwa,a Basit Yameen,b Muhammad Ashfaq,a Muhammad Nadeem Zafar,a Muhammad Zubaira

a

b

Department of Chemistry, University of Gujrat, Gujrat 50700, Pakistan

Department of Chemistry, Syed Babar Ali School of Science and Engineering (SBASSE), Lahore University of Management Sciences (LUMS), Lahore 54790, Pakistan

* Corresponding author E-mail address: [email protected], [email protected]

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ABSTRACT: Microemulsions (µEs) are unique systems that offer exciting perspectives in biophysical research for mimicing biomembranes at the molecular level. In the present study, biocompatible µE formulation of a new oil-in-water (o/w) system comprising of clove oil/Tween 20/2-propanol/water was accomplished for encapsulating an antibiotic, Levofloxacin (LVF). The pseudo-ternary phase diagram was delineated at constant cosurfactant:surfactant (2:1) mixture to meet the economic feasibility. The gradual changes occurring in the microstructure of asformulated four-component µE was explored via multiple complementary characterization techniques. The results of electrical conductivity (σ), viscosity (η), and optical microscopic measurements suggested the existence of a percolation transition to a bicontinuous structure in microregions of as-formulated µE. LVF displayed high solubility (5.0 wt%) at pH 6.9 in an optimum µE formulation comprising of 2-propanol (36.4%), Tween 20 (18.2%), clove oil (20.7%), and water (24.7%). The LVF-loaded µE composition showed long term stability for over 6 months of storage. Fourier Transform Infrared (FTIR) analysis showed that LVF was stable inside the µE formulation indicating the absence of any possible aggregation of LVF. Dynamic light scattering (DLS) revealed that the average particle size of drug-free µE (64.5 ± 3.4 nm) increases to 129.7 ± 5.8 nm upon loading of LVF, suggesting the accumulation of LVF in the interfacial layers of the micelles. Moreover, the fluorescence measurements indicated that the LVF might be localized in the interfacial film of µE system, which may result in controlled release of drug.


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INTRODUCTION With the aim to improve human health and life expectancy of the global population, nowadays rapid advancements could be observed in numerous diverse areas of biophysical and pharmaceutical sciences. One such imperative and emerging area is the development of controlled drug delivery systems (CDDS), which can administer active pharmaceuticals efficiently to execute therapeutic benefits during medical treatment.1─6 Among numerous CDDS, microemulsions (µEs) have emerged as attractive platforms mainly due to their ease of formulation, tunable physiochemical attributes, and biocompatibility. µEs can be defined as homogeneous dispersions of two immiscible fluids stabilized by surfactants, .7─10 µEs play significant role in drug delivery and sustained release of active drug component for topical, oral, nasal, ocular, transdermal, and parenteral medicaments.11─16 µEs have great solubilizing capacity for amphiphilic molecules. As smart drug carrier systems, µEs are optically isotropic, transparent, and thermodynamically stable homogeneous dispersions of two immiscible fluids (typically oil and water), stabilized by surfactants and often formulated in combination with cosurfactants.7,8,17─19 The surfactant molecules diffuse and link the inter-fluid boundaries thereby acting as a barrier for destabilization of architecture. A co-surfactant is used to adjust the interfacial interactions and to facilitate the spontaneous emulsification by the formation of an interfacial layer with an appropriate curvature. As compared to conventional vehicles (emulsions, pure oils, aqueous solutions, etc.) the amphiphilic components of the µE formulations act as penetration enhancers to increase the cutaneous absorption of both lipophilic and hydrophilic pharmaceutical ingredients across the diffusional barriers.20,21 Since, µE consist of various microdomains i.e. water-in-oil (w/o), oil-in-water (o/w), and bicontinuous, it is



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significant to inspect the microstructure of µE domains before its application as drug delivery vehicle.22─24 The aim of this work was the development of rapid oil-in-water (o/w) biocompatible µE to load the amphiphilic antibiotic, levofloxacin. Levofloxacin (LVF; refer to Fig. S1 in the SI section) is an efficient antibacterial drug capable of performing against bacterial infections and bacterial conjunctivitis.25,26 In the present study, a phase diagram is used to elucidate the roles of clove oil (eugenol), Tween 20 (surfactant) and 2-propanol (cosurfactant) in the encapsulation and improved loading of LVF. The reason for the selection of clove oil was its essential component eugenol, which exerts powerful anti-inflammatory, antimicrobial, antifungal, antiseptic, antiviral, aphrodisiac and stimulating effects.27 Tween 20 (refer to Fig. S2 in the SI section) is a biocompatible nonionic emulsifier and widely used as inert carrier in many pharmaceutical formulations,28,29 whereas the cosurfactant (2-propanol) is used to overcome the need of any additional input of energy in critical packing parameter of Tween 20. To explore the gradual changes occurring in the microstructure of µE, electrical conductivity, viscosity and optical microscopic analyses were performed. Moreover, the drug loading capacity, the storage stability, optical texture, and microstructure of LVF loaded optimum µE were evaluated. The stability of LVF inside µE was examined via Fourier Transforms Infrared (FTIR) spectroscopy. The average particle sizes of drug-loaded and drug-free optimum µEs were measured by dynamic light scattering (DLS), whereas the confinement of LVF in microregions of optimal µE was studied by fluorescence measurements. The high loading capacity of LVF (5.0 wt%) under milder conditions highlight the interesting feature of the herein developed µEs for pharmaceuticals sciences.


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EXPERIMENTAL Materials & Chemicals. Tween 20 (Polyoxyethylenesorbitan monolaurate), 2-propanol (99.5%) and clove oil were purchased from Sigma-Aldrich. Levoﬂoxacin (M.W: 361.37g/mol, 99.5%) was obtained from Schazoo Zaka Lab (PVT), Lahore, Pakistan and used without further purification. Acetonitrile of analytical grade (HPLC grade) was purchased from Merck®, whereas 0.45µm nylon filter papers (Sartorius, Germany) were used for filtration of the mobile phase. To control the pH abuffer solution consisting of 0.05M dipotassium hydrogen phosphate was used. Deionized water was used for dilution and for other experimental purposes. Microemulsion Preparation and Drug Incorporation. The pseudo-ternary phase diagram was mapped (as shown in Fig. 1) using oil (clove oil), surfactant (Tween 20; HLB=16.7), cosurfactant (2-propanol), and water with constant cosurfactant:surfactant mass ratio (2:1). In order to achieve stable and transparent microemulsion (µE) formulations, clove oil was first mixed with cosurfactant:surfactant mixture, subsequently water was added to obtain the desired µE compositions. Under the continuous stirring, LVF (5.0 wt%) was dissolved in an optimal µE formulation (optimized after electrical conductivity and viscosity analyses) comprising 36.4% of 2-propanol, 18.2% of Tween 20, 20.7% of clove oil and 24.7% of water. Both of the samples (drug-free and drug-loaded µEs) were stable, remaining clear and transparent over 6 months of storage.

Microemulsion Characterization Thermodynamic stability, Optical Transparency and Microscopy Measurements. To reveal the thermodynamic stability of pure and drug-loaded µEs, formulations were centrifuged at 5500 rpm for 20 min. Moreover, the homogeneity and isotropic nature of as-formulated µEs were verified by a Polarimeter (ATAGO, AP-100 Automatic Polarimeter) and visual evaluation at Page 5 of 28 ACS Paragon Plus Environment


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room temperature. Whereas, the structural transitions in the microstructure of µE were observed under a biological microscope (LABOMED FLR Lx 400; Jenoptic, Germany) through 4x/0.10/0.17 magnification. The samples were placed on slide to minimize possible destruction. Electrical Conductivity and Viscosity Measurements. The effects of water content on the microstructure of µE were monitored quantitatively by measuring the electrical conductivity (σ) and viscosity (η) experiments. The electric conductivity was measured by means of a Conductivity Meter (4510, Jenway) whereas, viscosities were measured with calibrated Brookfield viscometer (LVDV–2T) at 100 rpm. Both these measurements were performed by diluting oil/surfactant/cosurfactant mixture with water (along the dilution line AB in Fig. 1). Spectroscopic Measurements. The infrared (IR) spectra of pure and drug-loaded µE (5 wt.% drug) were recorded on Bruker FTIR (Alpha series) from 600 to 4000 cm-1. Steady-state fluorescence (SSF) measurements were performed on a Thermo Scientific NanoDropTM 3300 Fluorospectrometer with linear silicon CCD array detector. The light source was LED and operating voltage was 5.0 Vdc. Good resolution of the bands was obtained at the slit width (ex. 5.0 nm, em. 3.0 nm) with 40.0 µM concentration of the LVF. The fluorescence spectra for LVF in water phase, individual oil phase, surfactant/cosurfactant (1:2) mixture, and in the optimum µE system were recorded in the range of 395–660 nm upon excitation at 290 nm. The sample size was 2.0 µL. Particle Size Analysis. The average droplet size and polydispersity index (PDI) of drugloaded and drug-free µE was measured using laser particle size analyzer (BT-90 NANO PSA Bettersize) at laser wavelength of 635nm.

RESULTS AND DISCUSSION


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Phase Studies. Figs. 1 and S3 manifest the pseudo-ternary phase diagrams and area of existence of a large single-phase µE region for Tween 20/2-propanol/clove oil/water, which formed spontaneously at ambient temperature when the components were brought in contact. Mapping the area of the monophasic region is a handy tool for the selection of suitable cosurfactant:surfactant mixture (Smix) for drug encapsulation.11

0.0 1.0

0.2

0.8

Cl ov

Av era ge dr 64 o .5 nm plet si (D LS ze )

eo

il

0.4

0.6

0.6

B

ter Wa

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


0.4

0.8

0.2

µE 1.0 0.0

0.2

0.4

0.6

A 0.8

0.0 1.0

2-Propanol:Tween 20 (2:1) Fig. 1. Pseudo-ternary phase diagram showing microemulsion (µE) region (blue shaded) of clove oil/water/2-propanol/Tween 20 with Smix 2:1. AB represents the dilution line, selected for further investigations. The highlighted mark on dilution line represents the optimal µE composition.

The phase study revealed that the area of isotropic region remained nearly the same by increasing the Smix from 1:1 to 2:1. Although, in both the formulations the amount of the oil component was almost same, however, the viscosity of the formulation was less for Smix (2:1). In



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order to avoid any potential toxicity of higher surfactant content and to achieve a less viscous (i.e., having better flow properties) and larger isotropic region, Smix (2:1) was selected.11,30 In order to study the progress of morphological transformation from an oil-rich (w/o) to a water rich (o/w) system upon aqueous dilution without visual phase separation, a water dilution line AB with constant ratio of oil to Smix was selected. Such a titration method offers the advantages of being rapid, accurate and economical. However, the different types of o/w, w/o and bicontinuous µEs within the monophasic region cannot be identified from the phase diagram constructed on the basis of titration.11,31─34 Therefore, to explore the gradual changes occurring in the microstructure of µE, the electrical conductivity (σ) viscosity (η) and optical microscopic analyses were employed as a function of weight fraction of aqueous component Φw (wt.%) for the oil/Smix mixture along the dilution line AB (shown in Fig. 1). Electrical Conductivity Measurements. To assess the structural transition in µE and to explain the existence of a conductive network channel (bicontinuous µE), conductometry is a useful tool.35 Electrical conductivity (σ) was used to observe the phase transition from w/o (water-in-oil) to o/w (oil-in-water) µE as a function of weight fraction of aqueous component Φw (% wt) for the oil/Smix mixture along the dilution line AB. The results of variation of σ versus Φw are shown in Fig. 2.


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500 45 400

30

300

15

200

0

100

-15

dσ /dΦ

σ (µ S/cm)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


-30

0 W/O

B.C. Φp

-100 0

10

20

O/W -45

Φb

30

40

50

Φw (%wt)

Fig. 2. Variation of electrical conductivity (σ) and the first derivative of the electric conductivity (dσ/dΦ) with Φw (wt%) along the dilution line AB (shown in Fig. 1).

The σ of the system comprising of clove oil/Tween 20/2-propanol slightly changed with Φw increment till critical Φw of ∼11.0%. Beyond this an abrupt change in σ was observed, known as percolation threshold (Φp).35 The value of σ below Φp suggests the distinctive reverse micelles, forming w/o µE, whereas beyond Φp (Φw > 11.0%), the steep rise in σ till Φb (Φw ∼30.0%) indicates a network of channels forming bicontinuous µE.36 On contrary, the σ showed a sharp dip in its values with more water content above Φb (Φw > 30.0%), which suggested the phase transition, forming o/w µE.36,37 These phase transitions in µE microdomains were further explored by means of dσ/dΦ with Φw increment as shown in Fig. 2. A maximum in the first



derivative (dσ/dΦ) of σ is observed at Φw of ∼24.0%; reflecting on the existence of bicontinuous microstructure in this region.38─41 Viscosity Measurements. To observe the occurrence of any phase inversion phenomenon in the µE system, viscosity measurement is a useful tool as it exclusively depends on the sizes of droplets.41-44 Viscosity (η) of oil/Smix mixture along the dilution line AB (Fig. 1) was measured as a function of Φw (Fig. 3).

24

0.40

21

0.32

18 0.24

dη /dΦ

η (cP)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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15 0.16 12 0.08 W/O 9 0

Φp

B.C.

10

20

Φb

30

O/W 40

50

Φw (%wt)

Fig. 3. Variation of viscosity (η) and its first derivative with water content (dη/dΦ) as function of Φw (wt%) along the dilution line AB (shown in Fig. 1).

The consistent increase in η with an increase in Φw is probably due to the change in the microstructure of µE that could be ascribed as either a change in the shape of droplets or a possibility of transition from w/o to bicontinuous µE. The resulted increase in η of µE system Page 10 of 28 ACS Paragon Plus Environment

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might be helpful in the slow diffusing of drug at infinite dilution.45 The phase transitions in µE microdomains and percolation threshold (Φp) can be affirmed by dη/dΦ plot with Φw increment, as shown in Fig. 3. A minimum of first derivative (dη/dΦ) at Φw of ∼24.0% is observed suggesting the existence of bicontinuous microstructure in accordance with electrical conductivity data.38─42 Optical Microscopic Studies. With the aim to explore the gradual changes occurring in the microstructure of as-formulated µE and to access the existence of bicontinuous structure, samples were examined under a biological microscope under the experimental conditions as reported in the previously published work.46 Fig. 4a and 4c represents the water in oil (w/o) and oil in water (o/w) µE respectively and reveal the spherical droplets of dispersed oil and water in their respective continuous phases. Fig. 4b represents the formation of network like structures instead the cluster of spherical droplets, indicating the appearance of bicontinuous structure. Similar kind of results were obtained and reported by other researchers.46



(a)

(c)

(b)

25µ µm

Oil-rich

Water-rich

25µ µm

25µ µm

Bicontinuous microemulsion

W/O microemulsion

O/W microemulsion

er at W

Water

~11.0 wt.%

Oil

~30.0 wt.%

Oil

Oil W at er

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Increase in aqueous phase concentrion

Fig. 4. Microscopic images along with suggested microstructure changes in µE with dilution.

The optical microscopic study was carried out to observe the effect of dilution on the microstructure of µE. The globules formed are found to be in the range of 3- to 9-µm. The suggested microstructure changes in µE with dilution are also sketched in Fig. 4. In the phase region deficient in aqueous content (11.0 wt.% and < 30.0 wt.%, a bicontinuous structure was assumed, which is in good agreement with previously published work.46 At a > 30.0 wt.% aqueous component, we again observed spherical droplets, which suggested the conversion of w/o µE to o/w µE passing through a bicontinuous structure, which clearly indicates the “Percolation phenomena”. Similar kind of results were found and reported by other researchers.35,39,47─50


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Selected optimal µE formulation, comprising of 2-propanol (36.4%), Tween 20 (18.2%), clove oil (20.7%), and water (24.7%) along with 5.0 wt% loading of LVF, was further analyzed by conductivity, viscosity, density, refractive index, FTIR, DLS and fluorescence measurements. The values of measured parameters have been presented in Table 1.

Table 1. Physical Parameters of Optimal LVF-free and LVF-loaded µE (LVF = 5.0 wt%) at pH = 6.9 Physical Property Refractive index Conductivity (µS/cm) Viscosity (cP) Density (g/L) Particle size (nm) Stability FTIR Fluorescence

Clove oil (20.7%), Water (24.7%), 2-Propanol (36.4%), Tween 20 (18.2%) LVF-free LVF-loaded 1.44143 ± 0.00015 1.44083 ± 0.00012 205.7 ± 2.9 198.8 ± 2.5 18.2 ± 1.4 20.4 ± 1.6 0.94769 ± 0.002 0.95982 ± 0.002 64.5 ± 3.4 129.7 ± 5.8 Over 6 months storage No intermolecular interaction among LVF and µE ingredients LVF may reside in the interfacial film of µE

Infrared Study. To confirm the compatibility of LVF in µE formulation, Fourier Transforms Infrared (FTIR) anlaysis was conducted. FTIR spectra of pure LVF powder, drug-loaded and drug-free optimum µEs are shown in Fig. 5. The IR spectrum of LVF loaded µE is entirely different from LVF powder, while it closely resembles the spectrum of plain drug-free µE.


3080

2933

3264

C−−N str.

C−−F str.

3200

4000

2400

1600

3200

Drug free µE Drug loaded µE

801

C=Cstr.

1289

C=Ο Ο str.

1087

1721

ArH str. O−−H str. C−−H str.

1619

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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Transmittance (a.u)


ArH bend.

-1

800 cm

2400

1600

800

-1

Wavenumber (cm ) Fig. 5. Infrared (IR) spectra of plain microemulsion (µE) and drug loaded microemulsion. Inset: IR spectrum of pure powder LVF.

FTIR of LVF showed the following characteristic peaks: the major peak at 3264 cm-1 is observed due to carboxylic group (O-H stretching) whereas the peak at 2933 cm-1 assigned to CH3 group (C-H stretching). The peaks at 3080 cm-1 and 801 cm-1 assigned to C-H stretching and C-H bending due to aromatic group, respectively. The peaks at 1721 cm-1 and 1619 cm-1 are observed due to stretching of carbonyl group (C=O stretching) and aromatic rings (C=C stretching), respectively. Two peaks at 1289 cm-1 and 1087 cm-1 designate for stretching of amines (C-N stretching) and the presence of halogen group (C-F stretching), respectively. The IR spectrum of drug-loaded µE is similar to that of drug-free µE, indicating the high compatibility of drug with µE excipients. The absence of any additional peak in spectrum of drug-loaded µE


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signifies the complete solubilization and absence of larger aggregates of the drug within the formulation matrix. Similar kind of results were obtained and reported by other researchers.51─55 Dynamic Light Scattering Study. The average particle sizes of drug-loaded and drug-free µEs were measured by dynamic light scattering (DLS). The effect of water dilution on the size distributions of the oily droplets was measured as a function of Φw (inset in Fig. 6). The droplet diameters were decreased from 93.6 ± 4.6 to 63.2 ± 2.9 nm as the aqueous content of the systems increased from 2 to 34% w/w. In other words, as less oily phase and surfactant molecules were present in the system, the size of the dispersed droplets and also the polydispersity were decreased. This observation could be ascribed as when the surfactant concentration decreases upon aqueous dilution there will also be a decrease of interdroplet interactions in a monotonic manner and hence in the water-rich region the particle size is relatively small. Similar behavior has also been reported by other researchers for biocompatible U-type µEs.34,56─58 Fig. 6 shows the particle size distribution along with the values of polydispersity index (PDI) of both, drug loaded and drug free optimum µEs.



48

Drug-free µΕ

91

d (nm) PDI 64.5 0.112

Size (d.nm)

84

40

77

Drug-loaded µΕ

70

d (nm) PDI 129.7 0.247

63

Volume (%)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

32

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0

7

14

21

28

35

Φw (%wt)

24 16 8 0

0

40

80

120

160

200

Size (d.nm) Fig. 6. Particle size distributions of drug-loaded and drug-free optimum µE analyzed by DLS. Inset: size distributions of drug-free µE along the dilution line AB (shown in Fig. 1).

The PDI value of both samples, drug loaded and drug free µEs, were found to be 0.247 and 0.112 respectively, suggesting the higher uniformity of the droplet size within the formulation.44,53,59 Moreover, the drug free µE showed average particle size of 64.5 ± 3.4 nm in diameter and upon loading of LVF (5.0 wt.%) the particle size increases to 129.7 ± 5.8 nm, indicating the presence of drug in the droplet of µE. The increase in the particle size in drugloaded µE is probably be due to the accumulation of LVF in the droplets’ interfacial layers.44,60,61 Fluorescence Measurements. Since, the fluorescence of the amphiphilic fluorophores mainly depends on the polarity of the medium, so to locate the solubilized drug in microregions of µE system the steady-state fluorescence measurements have been extensively used.23,41,59,62─66


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The fluorescence spectra for LVF in aqueous phase, oil phase, Smix (2:1), and in the optimum µE system are shown in Fig. 7.

2000 454nm

Fluorescence Intensity

1600

er at W

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


479nm O

Oil

O

HO

F N O

O

O

OH F

N

N N

N

1200

N O

800

Water Oil S/CoS µE

400

0 420

480

540

600

660

Wavelength (nm) Fig. 7. The fluorescence spectra of LVF in aqueous phase, individual oil phase, in S/CoS, and in the optimum µE.

The emission maximum (λem) of LVF in aqueous phase is 454 nm (characteristic of LVF), whereas for oil phase the λem is 479 nm and was found to be situated at 482 nm for S/CoS system. The fluorescence spectra of LVF in pure water and oil are constituted by a number of peaks of different intensities. The original pattern of LVF fluorescence signals observed in pure water and oil are also preserved in the optimum µE and S/CoS, however, the λem in case of optimum µE system and in S/CoS is observed to shift to a higher wavelength of 482 nm (Fig. 7). While comparing the fluorescence of LVF in pure water, oil, S/CoS and optimum µE, the highest Page 17 of 28 ACS Paragon Plus Environment


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quenching of LVF fluorescence was observed in the optimized µE system. This observation indicates the close confinement of LVF molecules while in µE, which leads to the self-quenching and hence the lower intensity of the fluorescence signal of LVF.

44,67

Moreover, an increase in

hydrophobicity in the core interior merely drags the LVF molecules more deeply into the interfacial film and ceases their movement towards the aqueous bulk domain. In our study, the high solubility of LVF (∼5.0 wt%) in µE probably due to the solubilization of LVF at the interface. It is therefore suggested that the LVF may reside in the interfacial film of µE system.

CONCLUSIONS We formulated a new four-component microemulsion (µE), comprising of clove oil/Tween 20/2propanol/water, for encapsulation of Levofloxacin (LVF). To explore the phase transition in the microstructure of µE, electrical conductivity, viscosity and optical microscopic analyses were employed. Moreover, the stability of LVF inside µE matrix was examined via FTIR, whereas the confinement of LVF in microregions of µE was assessed by steady-state fluorescence measurements while changing in average particle size of µE on LVF loading was measured by DLS. Our results demonstrate that as-formulated µE remained stable and clear for over 6 months upon loading of LVF (5.0 wt.%). It is, therefore, anticipated that the solubilized LVF may reside in the interfacial film of µE system which may lead to controlled release of LVF. However, to characterize the microstructure of µE an extensive amount of fundamental work is still required before its execution as adaptable drug delivery vehicle.


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Supporting Information Description: Supplementary data associated with this article shows basic molecular structures of Levofloxacin (drug) and Tween 20 (surfactant) and also shows pseudo-ternary phase diagram of clove oil/water/2-propanol/Tween 20 with Smix 1:1.

Acknowledgement Authors express gratitude to the Department of Chemistry, University of Gujrat, Pakistan for the provision of lab facility. The authors also extend their sincere appreciations to Higher Education Commission of Pakistan for providing financial support through NRPU Project No. 4557.



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Biocompatible Microemulsion Formulation

40

Clove oil (20.7%), Water (24.7%), Tween 20 (18.2 %), 2-Propanol (36.4%)

Stable over 6 months r

er at W

32

e at W O

Oil

Oil

O

HO

F N O

Volume (%)

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O

O

OH F

N

N N

N

N O

24 drug-free µΜ

drug-loaded µΜ

d (nm) PDI 64.5 0.112

d (nm) PDI 129.7 0.247

σ (µS/cm) η (cP)

σ (µS/cm) η (cP)

16 198.8

18.2

205.7

20.4

8

0

0

40

80

120

160

200

Size (d.nm)


Encapsulation of Antibiotic Levofloxacin in ... - ACS Publications

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