Development and study of mixed surfactant microemulsion of Ofloxacin

 

Shashikant Chandrakar *, Amit Roy, Ram Sahu

Columbia Institute of Pharmacy, Raipur (C.G.) 493111 India

 

Abstract:

Ofloxacin is fluoroquinolone derivative antibacterial agent and it is used for the treatment of bacterial infection into the eye as a eye drop. The eye drops are the most commonly used dosage form for the ocular route but it has several disadvantages like  rapid wash out and dilution of the formulation leading to a low permeability of drug. The purpose of this work is to develop microemulsion formulation of Ofloxacin for ocular drug delivery. Pseudoternary phase diagram was prepared by using Span 20, Tween 80 (ratio of 1:1 to 2:1) as mixed surfactant, oleic acid   as oil phase and double distilled water as aqueous media. The microemulsion OM1 to OM6 was prepared on the basis of microemulsion area present in pseudoternary phase diagram. The prepared microemulsion was evaluated for phase separation, globule size analysis, zeta potential polydispersity index, percentage drug content, viscosity, conductivity, %transmittance and pH.

 

 

1. INTRODUCTION:

The conventional ophthalmic drug delivery like eye drop has problem like low permeability due to ocular anatomical and physiological barrier present into the eye which causes the poor permeability of drug at the site of action. Due to blinking reflex and nasolacrimal drainage most of eye drop are washed out from the ocular surfaces .The cornea act as a barrier to the absorption of drug due to its tightness and loss of the instilled solution from the precorneal area. In conventional formulation Oflxacin is used as eye drop for treatment of bacterial infections. Oflxacin is fluoroquinolone derivative, is broad spectrum antibacterial agent. It is mostly used for treatment of ocular infection. The Oflxacin is antibacterial agents that target bacterial enzymes to inhibit enzymes involved in bacterial DNA synthesis. After instillation of an eye drop, typically less than 5-10 % of an applied dose reaches the intraocular tissues. This is due to restriction caused by corneal barrier and rapid loss of the instilled solution through nasolachrymal drainage^(1.2,3). Therefore small amount of drug is available for therapeutic response and it needs frequent dosing of therapeutics agent^{(3, 4)}. The ocular bioavailability of the topically administered drug can be increased by increasing the corneal permeability and by increasing   contact time of drug on the ocular surface^(5-8).  After the penetration of the drug through the  cornea or sclera, kinetics of drug transport dependents on  physicochemical properties of drug like  molecular  weight or molecular volume, and binding  characteristics of drug to the tissue binding sites, clearance  of the fluid  reservoirs present in the eye.  This approach of ocular drug delivery can possible when a non toxic and non irritating permeation enhancer is used in dosage form ^(9,10,11,12). This strategy is used in Microemulsion. Microemulsion is composed of surfactant, co-surfactant, oil phase and aqueous phase ⁽¹³⁾. The aim of this study was to formulate microemulsion by using nonionic surfactant, oleic acid and water. The effect of ratio of surfactant, on globule size, PDI, drug content viscosity, pH, conductivity, size etc. were observed. 

 

2. MATERIAL AND METHOD:

The nonionic surfactant, span 80, span 20,tween 20, tween 80, methanol, disodium hydrogen phosphate, potassium dehydrogenate phosphate ethanol, sodium hydroxide, potessium chloride were purchased from Loba chemi. The Ofloxacin was obtained as gift sample from Sun Pharmaceutical Limited, Dewas (MP),The aqueous media used for preparation of microemulsion was double distilled water prepared in Lab. The entire chemical used were analytical grade.

 

2.1 Selection of surfactant

 The surfactant was selected on basis of literature survey. The km ratio was determined on the basis mixing surfactant at different ratio with oil. When the clear phase was obtained, such type of ratio was selected for construction of pesudoternary phase diagram ^{(14, 15)}.

 

2.2 Solubility of Ofloxacin in different oil

The solubility Ofloxacin in different oil was determined. The oils used were seasame oil, Isopropyl myristate, and Oleic acid. Solubility studies were conducted by placing an excess amount of drug in  2 ml of vehicle. Then the mixture was vortexed and kept for 48 hrs at 25ºC in a Orbital shaker (Remi Ltd.) to facilitate the solubilization. The samples were centrifuged at 3000 rpm for 15 min to remove the undissolved drug. The supernatant was taken and the aliquots of supernatant were filtered through 0.45 μm membrane filters and the solubility of Ofloxacin was determined by analyzing the filtrate spectrophotometrically after dilution with methanol at 296 nm. The concentration of drug in each vehicle was quantified by U.V-spectrophotometer ^{(16, 17, 18)}.

 

2.3 Construction of pseudoternary phase diagram

For preparation of microemulsion, the concentration of S_mix, oil, and water in appropriate ratio is required. Pseudo-ternary phase diagrams were constructed by water titration method at room temperature. Oleic acid  oil was used as oil phase and span 20, tween 80 was used as surfactant. Distilled water was used as an aqueous phase. Three phase diagrams were prepared with the 1:1 and 2:1 weight ratios of S_mix respectively. For each phase diagram at a specific surfactant weight ratio, the ratios of oil to the mixture of surfactant were varied as 1:9 to 9:1.The mixtures of oil, surfactant at certain weight ratios were titrated by adding water drop wise with stirring in magnetic stirrer. If it was clear and showed flowability it was considered as microemulsion. If the prepared microemulsion was highly viscous and it did not show change in height of meniscus when it was tilted,it was considered as gel or liquid crystal. The prepared microemulsion was further evaluated for different parameter ^{(16, 20)}.

 

2.4 Preparation of microemulsion

The microemulsion was prepared according to the composition given in table 3. Drug was dissolved in oil phase, then surfactant mixture was added with steering in magnetic stirrer with at 25⁰C.The water was added drop wise. When the entire component was mixed properly, the prepared mixtures were visually observed for clear/transparent or translucent mixture and their flowability ^{(19, 20)}.

 

2.5 Phase separation study

The small amount of sample of the above prepared microemulsion was taken in centrifugation tube and centrifuged at 5000 rpm for 15 min in cooling centrifuge (Remi India Limited) for phase separation study of                  microemulsion ^(15,20).

 

2.6 Globule size, Zeta potential and polydispersity index (PDI) analysis and drug content The average globule size and PDI of micro emulsions were deter-mined by photon correlation spectroscopy. Measurements were made using Malvern Zetasizer .The zeta potentail is also measured by same equipment. It is measured for the stability of microemulsion. Drug content of microemulsions was analyzed by UV–visible spectrophotometer (Shimadzu 1800, Japan) at 295 nm ^{(21, 22, 23)}.

 

2.7Statistical Analysis

The values were expressed as mean ± SD.

 

 

 

3.  RESULT AND DISCUSSION:

3.1 Selection of surfactant

The km ratio was determined on the basis mixing surfactant at different ratio with oil.  The clear phase was obtained with surfactant ratio of 1:1 and 2:1 (Table 1) .such type of ratio was selected for construction of pesudoternary phase diagram.

 

Table 1.Selection of surfactant and Oil phase

Oil	Surfactant  Mixture	KM Ratio	Observation
Oleic acid	Span  20:Tween 20	1:1	Hazy
Oleic acid	Span  20:Tween 20	2:1	Hazy
Oleic acid	Span  20:Tween 20	3:1	Hazy
Oleic acid	Span  80:Tween 20	1:1	Hazy
Oleic acid	Span  80:Tween 20	2:1	Hazy
Oleic acid	Span  80:Tween 20	3:1	Hazy
Oleic acid	Span  20:Tween 80	1:1	Clear
Oleic acid	Span  20:Tween 80	2:1	Clear
Oleic acid	Span  20:Tween 80	3:1	Hazy
Oleic acid	Span  80:Tween 80	1:1	Hazy
Oleic acid	Span  80:Tween 80	2:1	Hazy
Oleic acid	Span  80:Tween 80	3:1	Hazy

 

3.2 Solubility of Ofloxacin

Solubility of Ofloxacin in different oil and surfactant was given in table 2 surfactant and water are shown in table 2. The Oleic exhibited highest solubility (27.2±0.06 mg/g) compared to other solvents.

 

Table 2: Solubility of Ofloxacin in Different oil and surfactant

Values reported as mean ± SD; (n = 3)

 

3.3 Phase behavior study

For different ratio of surfactant, two phase diagram were constructed - comprising different S_mix ratio (Span 20: Tween 80) i.e., 1:1, 2:1. These pseudo-ternary phase diagrams (consisting oil, S_mix and water) demonstrated an extensive region of microemulsion formation (Figure. 1 and 2). In addition, phase diagrams also help in determination of concentration range of components used for formulation of microemulsion ^{(15, 16, 23)}.

 

 

Figure 1: Phase diagram for surfactant mixture (Span 20: tween 80) at 1:1

 

Figure 2: Phase diagram for surfactant mixture (Span 20: tween 80) at 2:1

Table 3: Composition of Microemulsion

Formulation	Oil  (%wt/ wt)	Water (%wt/ wt)	Smix1:1 (%wt/ wt)	Smix 2:1 (%wt /wt)	Drug  (% wt/wt)
OM1	10	45	45		0.3
OM2	15	40	45		0.3
OM3	20	35	45		0.3
OM4	20	45		35	0.3
OM5	25	40		35	0.3
OM6	30	35		35	0.3

 

3.4 Globule size, Zeta potential and polydispersity index (PDI) analysis and drug content

Globule size of microemulsions was found in the range 92.6 ± 0.07 to 187.4 ± 0.03 nm (Table 4). It was observed that globule size enhances on increasing the concentration of internal phase. The size reduces on enhancing surfactant concentration. The zeta potential values ranged between -24.67 ± 0.07 and -42.71 ± 0.06 mV. This variation may be due to diffusion of drug in interface. The zeta potential is determined to predict the physical stability of colloidal systems .Theoretically, the higher the zeta potential value stables the colloidal system. Here the zeta potential value is negative. The polydispersity index (PDI) of the prepared microemulsion was found between 0.191 ±0.03 to 0.384±0.16 which is below 0.4 that is showing narrow size distribution of globules. From the literature it was observed  that  narrow size distribution of globule, PDI ranges from 0.01 to 0.5.If the PDI value of dispersion is > 0.7 it has broad range size distribution^{(15,19,22.23,24)} The drug content of microemulsion varies from 98.38 ± 0.14% to 99.45 ± 0.18 % (Table 4).

 

Table 4: Globule size, Zeta potential, PDI, and Drug content of prepared microemulsion

Formulation	Phase separation	Globule size (nm)	Zeta Potential  (m V)	PDI	Drug Content
OM1	Not Obtained	92.6±0.07	-24.67±0.07	0.238±0.01	99.03± 0.11%
OM2	Not Obtained	123.3±0.04	-29.98±0.03	0.216±0.14	99.35± 0.31%
OM3	Not Obtained	134.1±0.06	-32.92±0.02	0.281±0.07	99.25± 0.22%
OM4	Not Obtained	156.7±0.08	-31.53±0.04	0.384±0.16	99.45± 0.18%
OM5	Not Obtained	162.2±0.07	-34.53 ±0.01	0.298±0.08	99.23± 0.26%
OM6	Not Obtained	187.4±0.03	-42.71±0.06	0.191±0.03	98.38± 0.14%

Values reported as mean ± SD; (n = 3)

 

3.5 Viscosity, conductivity, % Transmittance and pH

Viscosity is important parameter to assess the ocular formulation; hence low viscosity causes fast drainage of formulation and reduces retention time. The viscosity value of above prepared microemulsions varied from 7.32 ± 0.23 to 13.18 ± 0.21 cp (Table No.4).The above range of viscosity for microemulsion is acceptable for ophthalmic formulation hence eye drops should have viscosity < 20 centipoises^(15,16). In the microemulsion variation in viscosity is due to variation in concentration of surfactant .In microemulsion OM3 surfactant concentration is higher as well as low amount of oil (internal phase) , therefore it showed higher viscosity. In microemulsion OM6, surfactant concentration is low as well as oil content is high therefore it was showing low viscosity value. The results from the conductivity analysis of the microemulsions showed conductivity values from 106.04 ± 0.19 to 118.04 ±0.16 µS/cm.(Table 4).The conductivity values were higher than 0.05 µS/cm which indicates that the prepared microemulsions were of the oil in water type ^(15,16).  The higher conductivity may be due to   presence of water in external phase which may contain diffused drug ^{(22, 25, 26)}.

 

The percentage transmittance of microemulsion varies from 98.2 ± 0.12% to 99.8 ± 0.16% % (Table 4).It showed acceptable value for ocular formulation. Low % transmittance value for OM1 is due to higher amount of surfactant. As surfactant concentration decreases the clear microemulsion was obtained. The pH values of microemulsions were varied from the range 6.55 ± 0.21 to 7.22 ± 0.15 (Table 4). The pH for ophthalmic solutions may range from pH 4.5 - 11.5 but to prevent corneal damage; the pH should be 6.6 to 7.8. The eye has limited buffering capacity the pH values of the prepared microemulsion should be considered as nonirritating to the eye ^{(15, 16, 22, 26)}.

 

Table 5: Viscosity, Conductivity, %Transmittance and pH of prepared microemulsion

Formulation	Viscosity(cp)	Conductivity(µS/cm)	% Transmittance	pH
OM1	9.14±0.14	118.04±0.16	98.2±0.12%	7.22±0.15
OM2	11.21±0.27	112.06±0.23	98.4±0.18%	7.13±0.12
OM3	13.18±0.21	106.04±0.19	98.7±0.11%	7.04±0.13
OM4	7.32±0.23	117.03±0.21	99.8±0.16%	6.74±0.16
OM5	10.22±0.17	114.02±0.14	99.6±0.13%	6.82±0.14
OM6	12.18±0.12	109.06±0.27	99.3±0.14%	6.55±0.21

Values reported as mean ± SD; (n = 3)

CONCLUSION:

Microemulsions of Ofloxacin using nonionic mixed surfactant were developed and characterized for different parameter. All the parameter were acceptable for ocular formulation accept globule size of OM7 to OM9.Narrow size range microemulsion (<200 nm) are better for ocular drug delivery. Therefore Microemulsion OM1 to OM6 was selected for further study of formulation.

 

REFERENCES:

1.     Goodman and Gilman’s, The pharmacological basis of Therapeutics, VIII edition, 9 (1); 1992:1027-1059.

2.     Wise R.  and Lockley M. R. The pharmacokinetics of ofloxacin and a review of its tissue penetration. Journal of Antimicrobial Chemotherapy, 22;1988: 59–64.

3.     Monk J.P., and Campoli-Richards D. M. Ofloxacin. A review of its antibacterial activity, pharmacokinetic properties and therapeutic use. Drugs.33 (4); 1987.346-91.

4.     Sanders W. E. Jr. Oral Ofloxacin: A Critical Review of the New Drug Application. Clinical Infectious Diseases. .14(2); 1992:539-54.

5.     Colo G Di. et al. Effect of chitosan on in vitro release and ocular delivery of ofloxacin from erodible inserts based on poly(ethylene oxide).   International Journal of Pharmaceutics. 2002;248(1–2):115-122.

6.     Mouton Y.  and Leroy 'O. Ofloxacin. International Journal of Antimicrobial Agents. 1991; 1 (2-3): 57-74.

7.     Patrick M. Hughes et al.Topical and systemic drug delivery to the posterior segments .Advanced Drug Delivery Reviews. 2005;57: 2010– 2032.

8.     Tangri P., Khurana S. Basics of Ocular Drug Delivery Systems. International Journal of Research in Pharmaceutical and Biomedical Sciences.2 (4); 2011:1541-1552.

9.     Urttia Arto et al. Ocular absorption following topical delivery. Advanced Drug Delivery Reviews.16; 1995:3-19.

10.   Lang John C. Ocular drug delivery conventional ocular formulations. Advanced Drug Delivery Reviews 16; 1995:39-43.

11.   Wilson Clive G. Topical drug delivery in the eye. Experimental Eye Research. 2004; 78:737–743.

12.   Jiao Jim. Polyoxyethylated nonionic surfactants and their applications in topical ocular drug delivery. Advanced Drug Delivery Reviews. 2008; 60:1663–1673.

13.   Suruse P. and Bodhake A. Ophthalmic Microemulsion: A Comprehensive Review. Int. J. Pharm Bio Sci. 2012; 3(3):1-13.

14.   Kantarci G, Karasulu HY, Ozguney I, Guneri T. Phase behavior of microemulsion systems containing short chain alcohols as co-surfactant. Acta Pharmaceutical Sciencia. 2002; 44:77-85.

15.   Shanni K. and Karthikeyan K. Phase-transition W/O Microemulsions for Ocular Delivery: Evaluation of Antibacterial Activity in the Treatment of Bacterial Keratitis. Ocular Immunology and Inflammation. 2017; 25(7): 1–12.

16.   G.N. El-Hadidy et al.  Microemulsions as vehicles for topical administration of voriconazole: formulation and in vitro evaluation, Drug Dev. Ind. Pharm. 2012; 38:64–72.

17.   Pawar P.K. and Majumdar D.K. Effect of formulation factors on in-vitro permeation of moxifloxacin from aqueous drops through excised goat, sheep and buffalo corneas, AAPS Pharm. Sci. Tech. 200 6;7:E89–E94.

18.   H. Zhu, A. Chauhan. Effect of viscosity on tear drainage and ocular residence time, Optom. Vis. Sci. 2008; 85:E715–E725.

19.   S.P. Moulik, A.K. Rakshit, Physiochemistry and applications of microemulsions, J. Surface. Sci. Technol. 2006; 22:159–186.

20.   Benita S, Levy M.Y. Submicron emulsions as colloidal drug carriers for intravenous administration: comprehensive physicochemical characterization. J Pharm Sci.1993;82: 1069–79.

21.   R.G. et al.  W/O microemulsions for ocular delivery: Evaluation of ocular irritation and precorneal retention Journal of Controlled Release. 2006; 111: 145–152.

22.   Gasco M. R. et al. Preparation and evaluation in vitro of solutions and o/w microemulsions containing Levobunolol as ion-pair. International Journal of Pharmaceutics. 1993; 100:219–225.

23.   Nair R.  et al. Development and Characterization of Ofloxacin Microemulsions for Ocular Application International Journal of Chemical and Pharmaceutical Sciences. 2011;2(2):43-49.

24.   Habe A et al. Development and characterization of microemulsions for ocular application. European Journal of Pharmaceutics and Biopharmaceutics. 1997; 43: 179–183.

25.   Fialho S. L. and. da Silva-Cunha A. New vehicle based on a microemulsion for topical ocular administration of dexamethasone,” Clinical and Experimental Ophthalmology.2004; 32:626–632.

26.   Chan J., et al. Phase transition water-in-oil microemulsions as ocular drug delivery systems: in vitro and in vivo evaluation,” International Journal of Pharmaceutics. 2007; 328:65–71.

 

 

 

Received on 01.10.2017       Modified on 20.12.2017 Accepted on 11.01.2018      ©A&V Publications All right reserved Research J. Science and Tech. 2018; 10(2):93-97. DOI: 10.5958/2349-2988.2018.00013.X