Methods for preparing oil-in-water emulsions comprising cetalkonium chloride

Abstract

Compositions containing quaternary compounds in which the nitrogen atom is substituted by at least one alkyl group having at least 12 carbon atoms, and the composition includes at least 20% in weight by weight of the total composition, of ammonium halides in which the nitrogen atom is substituted by at least one alkyl group having at least 14 carbon atoms and more than 5%, preferably more than 7% in weight by weight of the total composition, of ammonium halides in which the nitrogen atom is substituted by at least one alkyl group having at least 16 carbon atoms. Also, ophthalmic oil-in-water emulsions containing such compositions, the ophthalmic emulsions being useful for eye care or for the treatment of eye conditions.

Claims

1. A method for preparing an oil-in-water emulsion; said oil-in-water emulsion comprising: an oil phase comprising mineral oil, castor oil or medium chain triglycerides, 0.0005% to 0.1% w/w of ammonium halide, cetalkonium chloride being the sole ammonium halide of the emulsion, and 0.1% to 1% w/w of a surfactant selected from tyloxapol, poloxamer 188, tocopherol polyethylene glycol succinate and a mixture thereof; and said method comprising the following steps: (i) providing said oil phase and an aqueous phase; (ii) optionally, heating said oil phase and said aqueous phase at the same temperature; (iii) dissolving said ammonium halide and said surfactant in said oil phase and/or in said aqueous phase; (iv) mixing said oil phase and said aqueous phase by magnetic stirring, so as to obtain a first coarse emulsion; (v) adjusting the droplet size of said first coarse emulsion by high shear mixing and/or high-pressure homogenization, so as to obtain said oil-in-water emulsion; and (vi) optionally, sterilizing said oil-in-water emulsion by heating.

2. The method according to claim 1, wherein said mineral oil phase is a mixture of heavy and light mineral oil.

3. The method according to claim 1, wherein said oil-in-water emulsion further comprises buffering agents and/or at least one tonicity agent.

4. The method according to claim 1, wherein said oil-in-water emulsion has a positive zeta potential.

5. The method according to claim 1, wherein said oil-in-water emulsion has a droplet size of 100 to 500 nm.

6. The method according to claim 1, wherein said oil-in-water emulsion comprises light and heavy mineral oil, tyloxapol, poloxamer 188, mannitol and cetalkonium chloride.

7. The method according to claim 1, wherein said oil-in-water emulsion comprises light and heavy mineral oil, tyloxapol, poloxamer 188, glycerol and cetalkonium chloride.

8. The method according to claim 1, wherein said oil-in-water emulsion comprises medium chain triglycerides, tyloxapol, poloxamer 188, glycerol and cetalkonium chloride.

9. The method according to claim 1, wherein said oil-in-water emulsion further comprises an active principle.

10. The method according to claim 9, wherein said active principle is cyclosporine.

11. A method for preparing an oil-in-water emulsion; said oil-in-water emulsion comprising: an oil phase comprising mineral oil, castor oil or medium chain triglycerides, 0.0005% to 0.1% w/w of ammonium halide, cetalkonium chloride being the sole ammonium halide of the emulsion, and 0.1% to 1% w/w of a surfactant selected from tyloxapol, poloxamer 188, tocopherol polyethylene glycol succinate and s rbitan m n lauratc and a mixture thereof; and said method comprising the following steps: (1) manufacturing a pre-concentrate of said oil-in-water emulsion, said pre-concentrate having a content in oil of at least 4% v/v, by emulsifying and/or mixing said oil phase and said surfactant with an aqueous phase, so as to obtain said pre-concentrate; and then (2) diluting one volume of said pre-concentrate with 2 to 50 volumes of water, so as to obtain said oil-in-water emulsion.

12. The method according to claim 11, wherein said pre-concentrate has a content in oil of at least 10% v/v.

13. The method according to claim 12, wherein said pre-concentrate has a content in oil of at least 20% v/v.

14. The method according to claim 13, wherein said pre-concentrate has a content in oil of at least 30% v/v.

15. The method according to claim 11, wherein said oil phase and/or said diluting water comprise(s) said cetalkonium chloride.

16. The method according to claim 11, wherein the droplet size or the distribution of the droplet size in said pre-concentrate is the same as the droplet size or the distribution of the droplet size in said oil-in-water emulsion.

17. The method according to claim 11, wherein said diluting water comprises additives selected from tonicity agents, viscosifying agents, buffering agents, preservatives, antioxidants, and colorants.

18. The method according to claim 11, wherein said mineral oil phase is a mixture of heavy and light mineral oil.

19. The method according to claim 11, wherein said oil-in-water emulsion further comprises buffering agents and/or at least one tonicity agent.

20. The method according to claim 11, wherein said oil-in-water emulsion has a positive zeta potential.

21. The method according to claim 11, wherein said oil-in-water emulsion has a droplet size of 100 to 500 nm.

22. The method according to claim 11, wherein said oil-in-water emulsion comprises light and heavy mineral oil, tyloxapol, poloxamer 188, mannitol and cetalkonium chloride.

23. The method according to claim 11, wherein said oil-in-water emulsion comprises light and heavy mineral oil, tyloxapol, poloxamer 188, glycerol and cetalkonium chloride.

24. The method according to claim 11, wherein said oil-in-water emulsion comprises medium chain triglycerides, tyloxapol, poloxamer 188, glycerol and cetalkonium chloride.

25. The method according to claim 11, wherein said oil phase further comprises an active principle.

26. The method according to claim 25, wherein said active principle is cyclosporine.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) FIG. 1 refers to the timing of appearance of redness in ocular surface.

(2) FIG. 2 refers to Draize Test evaluation.

(3) FIG. 3 refers to the zeta potential of emulsions of example 2, part 3.b) in function of BAK concentration.

EXAMPLES

(4) All concentrations in the emulsion formula are expressed in weight/weight of the entire formulation, unless stated differently.

Example 1: Reduced Toxicity of Quaternary Amines when Incorporated into Emulsions

(5) Materials and Methods

(6) Material

(7) 1. Solution at 0.02% BAK (BAK So)

(8) TABLE-US-00001 Excipients Z01SOL472 BAK US 0.02 NaCl 0.612 Tris Buffer 5 mM pH 7.1 0.069% Tris HCl 0.006% Tris Base Water Ad 100

(9) 2. Emulsion at 0.02% BAK (BAK Em)

(10) TABLE-US-00002 Excipients Z01EM471 Mineral oil heavy 0.500 Mineral oil light 0.500 Tyloxapol 0.300 BAK US 0.02 Tris Buffer 5 mM pH 7.1 0.069% Tris HCl 0.006% Tris Base Poloxamer 188 0.100 Glycerol 1.6 Water (up to 100) Ad 100

(11) 3. Solution at 0.002% CKC (CKC Sol)

(12) TABLE-US-00003 Excipients Z01SOL473 CKC 0.002 NaCl 0.626 Tris Buffer 5 mM pH 7.1 0.069% Tris HCl 0.006% Tris base Water Ad 100

(13) 4. Emulsion at 0.002% CKC (CKC Em)

(14) TABLE-US-00004 Excipients Z01EM264 Mineral oil heavy 0.500 Mineral oil light 0.500 Tyloxapol 0.300 CKC 0.002 Tris Buffer 5 mM pH 7.1 0.069% Tris HCl 0.006% Tris Base Poloxamer 188 0.100 Glycerol 1.6 Water (up to 100) Ad 100

(15) 5. PBS

(16) Methods:

(17) Albino rabbits were administrated with 1 drop (50 l) each 5 minutes, for 15 times.

(18) Results

(19) Evaluation of Toxicity by Time of Redness and DRAIZE Test Items Analyzed at H4 and D1.

(20) The time of the beginning of the redness in conjunctiva following the 15 times of instillations was evaluated (FIG. 1). PBS did not induce any redness during all instillation period (data not shown). BAK Sol induced conjunctival redness very fast, about 10 to 15 minutes after the first instillation (after 2-3 drops). BAK Em, CKC Sol-instilled groups showed redness at about 25-35 minutes after the first instillation of eye drops (after 5-7 drops). CKC Em presented a visible redness at almost the end of the experimentation: 60 to 65 minutes after the first instillation (after 12-13 drops).

(21) Draize Test clearly showed that at four hours (H4) after the last instillation the ocular irritation was the most important in BAK Sol-instilled group, which was higher than BAK Em and CKC Sol groups (with no difference between these two groups). BAK Sol, BAK Em, CKC Sol all showed higher ocular irritation than CKC Em, which presented no difference with PBS-instilled group (FIG. 2).

(22) One day after the administrations (Dl), PBS, BAK Em, CKC Sol and CKC Em all returned to normal aspect without difference among them. But BAK Sol still induced more important ocular irritation than all other groups (P<0.0001).

Example 2: Stability of the Emulsions of the Invention

(23) 1. Emulsions Composition

(24) Some emulsions are described below:

(25) TABLE-US-00005 Composition Z01EM206 Z01EM209 MCT 2% 2% Tyloxapol 0.3% 0.3% BAK C16 (CKC) 0.02% 0.025 mM Poloxamer 188 0.1% 0.1% Glycerol 2.25% 2.25% Water qsp100 qsp100

(26) TABLE-US-00006 Z01EM Z01EM Composition Z01EM419 Z01EM264 Z01EM387 418(a) 418(b) Light mineral 0.5% 0.5% 0.5% 0.5% 0.5% oil Heavy mineral 0.5% 0.5% 0.5% 0.5% 0.5% oil Tyloxapol 0.3% 0.3% 0.3% 0.3% 0.3% Poloxamer 0.1% 0.1% 188 PG 0.19% PEG 300 0.19% PEG 400 0.19% Mannitol 3.3% 2% 2.5% 2.9% Glycerol 1.6% 0.19% 0.19% Cetalkonium 0.002% 0.002% 0.002% 0.002% 0.002% chloride (CKC) Tromethamine 0.006% 0.006% Tris HCl 0.071% 0.071% Water Up to 100% Up to 100% Up to 100% Up to 100% Up to 100%

(27) TABLE-US-00007 Composition Z01EM393-4 Z01EM395-6 MCT 1-2% 1-2% Tyloxapol 0.3% 0.3% PG 0.4% Mannitol 0.9% Glycerol 1% 1% Cetalkonium 0.01% 0.01% chloride (CKC) Water Up to 100% Up to 100%

(28) 2. Emulsions Preparation

(29) The oily and the water phases of the emulsion, which might contain or not an active principle, may be separately heated to an appropriate temperature. This temperature may be the same in both cases. Surfactants might be dissolved in the oil, water phase or in both. A first coarse emulsion is generated by magnetic stirring, and the droplet size is reduced by high shear mixing, high pressure homogenization, or both.

(30) The oil-in-water emulsions of the present invention can be sterilized after preparation using heat, for example, autoclave steam sterilization.

(31) 3. Impact of Chain Length on Emulsions Characteristics

(32) a) Emulsion Droplet Size

(33) The mean diameter of the oil droplets is determined by dynamic light scattering using a High Performance Particle Sizer type HPPS 5001 (Malvern Instruments, Worcestershire, UK). Measurements are performed at 25 C. following dilution of the emulsion in double distilled water.

(34) TABLE-US-00008 TABLE 1 Emulsions droplet size values (nm) 0.001% 0.0025% 0.005% 0.01% 0.02% 0.04% 0.1% BAK C12 198 263 230 225 180 BAK C14 204 190 190 155 238 185 BAK C16 220 210 148 180 155 188 183

(35) Emulsions of Table 1 and Table 2 contain 2% MCT, 0.3% Tyloxapol and 0.1%

(36) Poloxamer 188 and 2.25% glycerol and compositions of BAK; concentrations of BAK range from 0.001% to 0.1% in weight to the weight of the emulsion.

(37) b) Emulsion Zeta Potential

(38) The stability of the cationic emulsions of the invention may be evaluated by determining their ability to keep a positive and stable zeta potential overtime. Measurement of zeta potential is commonly used to assess the stability of colloidal systems. Indeed, zeta potential of the oil-in-water emulsion is representative of the electronic charges present at the surface of the oil droplets. When oil droplets are positively charged, droplets undergo electrostatic repulsions, avoiding their coalescence. Thus, retaining a stable positive zeta potential overtime for a cationic emulsion illustrates its stability.

(39) The zeta potential of emulsions comprising increasing concentrations of different quaternary ammonium compounds (BAK C12, BAK C14 and BAK C16) was thus measured.

(40) Zeta potential can be measured by a zetameter such as Zetasizer 2000, Malvern Instruments Ltd, UK. The zeta potential of the emulsion droplet surface is determined by electrophoretic mobility. Measurements are performed at 25 C. following dilution at 1:250 of the emulsion in double distilled water. The electrophoretic mobility is converted into zeta potential values through the Smoluchowsky equation. Results are presented in Table 2 below and graphically represented in FIG. 3.

(41) TABLE-US-00009 TABLE 2 Emulsions zeta potential values (mV) 0.001% 0.0025% 0.005% 0.01% 0.02% 0.04% 0.1% BAK C12 6.9 +4.2 +7.9 +16.8 +23.8 BAK C14 +11.4 +19.6 +22.9 +28.4 +39.3 +44.5 BAK C16 +16.2 +24.4 +31.4 +36.7 +44.1 +47.2 +48.9

(42) Results presented in Table 2 and FIG. 3 evidence that positive charges are attained at lower concentrations when the chain length of the quaternary ammonium compound is increased. They also suggest a preferential partition of more lipophilic chain (long alkyl chain) within the oil droplet surface.

(43) CKC (BAK C16) is thus proved to have a greater cationic power than a quaternary ammonium compound with smaller alkyl chain length. In other words, emulsions comprising CKC display a higher zeta potential than those comprising the same amount of BAK US. Therefore, BAK US and CKC are not interchangeable, which is confirmed by data of example 3.

(44) c) Emulsion Stability Overtime

(45) The stability of the emulsions can be evaluated by the evolution of their aspect, with a visual score with a visual score going from 13-best aspect to 1-total phase separation.

(46) It can be observed from the following table that, at equimolar concentration, longer (more lipophilic) chain length QA results in more stable emulsion.

(47) TABLE-US-00010 Following 3 months at Type and conc. of QA After preparation (T0) 40 C. 0.25 mM BAK C12 12 2 0.25 mM BAK C14 13 7 0.25 mM BAK C16 13 9 0.5 mM BAK C12 10 2 0.5 mM BAK C14 13 7 0.5 mM BAK C16 11 9

Example 3: Comparison of the Stability of Emulsions Comprising Either CKC or BAK

(48) The stability of emulsions containing 0.002% w/w of CKC and 0.002% w/w of BAK was compared. Compared emulsions have following compositions:

(49) TABLE-US-00011 0.002% CKC emulsion 0.002% BAK emulsion Light mineral oil 0.5% 0.5% Heavy mineral oil 0.5% 0.5% CKC 0.002% \ BAK US \ 0.002% Tyloxapol 0.3% 0.3% Poloxamer 188 0.1% 0.1% Glycerol 1.6% 1.6% Tris buffer 5 mM pH 0.071% Tris HCl 0.071% Tris HCl 7.1 0.006% Tris Base 0.006% Tris Base Water up to 100% up to 100%

(50) The zeta potential of the two emulsions was followed overtime using thermal stress test conditions. Thermal stress tests enable to accelerate commonly used stability tests. In the thermal stress test used therein, zeta potential of the emulsion was measured at T=0, i.e. as soon as the emulsion has been prepared. Glass vials of 10 mL effective capacity containing 5-10 mL of emulsion and sealed under nitrogen atmosphere were stored at 80 C. Then, at T=7 days and 14 days, the zeta potential was measured. Results of zeta potential measurement, in mV, are given in the table below:

(51) TABLE-US-00012 T0 T = 7 days T = 14 days 0.002% CKC emulsion +21.2 +22.6 0.002% BAK US emulsion +6.73 9.66 15.8

(52) It clearly appears that the emulsion containing 0.002% w/w of BAK does not keep a positive and stable zeta potential overtime, contrary to the emulsions of the invention containing 0.002% w/w of CKC.

(53) Considering the results reported above, it is clear that CKC and BAK US are not interchangeable.