FIELD OF INVENTION
This invention relates to a process for the preparation of poly DL-lactide-co-glycolide nanoparticles (PLG-NP) having two or more active substances encapsulated therein and which can be administered orally. Reference to active substances is intended to include therapeutic and/or bioactive substances. Thus, and as by way of example, the nanoparticles may be employed for encapsulation of antitubercular drugs (AID).
BACKGROUND OF INVENTION
The need to administer multiple ATD daily for 6-9 months is responsible for patient non-compliance as well as drug -related hepatotoxicity, which result in therapeutic failure. Another consequence of incomplete/irregular treatment is the emergence of drug resistance.
Nanotechnology based drug delivery systems employing biodegradable polymers is known in the art. Of the several procedures available to prepare nanoparticles such as double-emulsion-solvent-evaporation, solvent diffusion in oil, microemulsion, gas antisolvent precipitation, gelification of anionic polysaccharides etc., none is perfect in terms of particle size, drug encapsulation efficiency and drug release kinetics. Further, multidrug encapsulation in single formulation is not yet known. It is generally recognized that if the drug or active substance is not solubilized, it can not be encapsulated. Thus, the known processes utilize the property of solubilization for encapsulation. A problem associated with such a known method is that different drugs having different solubility properties.
OBJECTS OF THE INVENTION
An object of this invention is to propose a process for the preparation of PLG-NP having two or more different active substances with different or same solubility properties encapsulated therein and which obviate the disadvantages associated with the known art.
Another object of this invention is to propose a process for the preparation of PLG-NP having two or more different active substances with different or same solubility properties encapsulated therein which provides a prolonged and sustained drug (s) release.
Still another object of this invention is to propose a process for the preparation of PLG-NP having two or more different active substances with different or same solubility properties encapsulated therein capable of being modulated to entrap maximum drug.
Yet another object of this invention is to propose a process for the preparation of PLG-NP having two or more different active substances with different or same solubility properties encapsulated therein capable of distributing the drug(s)' evenly to different organs where tubercle bacteria reside.
A further object of this invention is to propose a process for the preparation of PLG-NP having two or more different active substances with different or same solubility properties encapsulated therein which can be lyophilized and reconstituted for use as an oral/aerosol formulation.
A still further object of this invention is to propose a process for the preparation of PLG-NP having two or more different active substances with different or same solubility properties encapsulated therein which does not exhibit hepatotoxicity.
BRIEF DESCRIPTION OF THE INVENTION
According to this invention, there is provided a process for the preparation of PLG-NP having two or more different active substances with different or same solubility properties encapsulated therein comprising:
1. preparing an aqueous solution of water soluble drugs
2. preparing polymer solution in an organic solvent along with the hydrophobia
drug.
3. mixing the aqueous and organic solution.
4. sonicating the mixture to make an emulsion under cold conditions.
5. pouring the primary emulsion in aqueous PVA and resonicating under cold
conditions to form a secondary emulsion.
6. stirring the said emulsion and centrifiiging the same.
7. Washing nanoparticles, lyohilizing and reconstituting the same.
In accordance with this invention, an aqueous solution of hydrophilic drug is prepared in DW/ normal saline /phosphate buffer sale in the ratio of 1:0.1-100 weight by volume. A solution of polymer is prepared in an organic solvent preferably dichloromethane (DCM) in the ratio 1:0.3-1 weight by volume, also containing the hydrophobic drug in the ratio 1:0.5-5 weight by volume. The aqueous solution is poured into the organic solution in the ratio 1:5-20 volume by volume and sonicated for 45-120 sec at 4°-15°C. The primary emulsion is poured into 0.8-2.5% PVA solution keeping DCM.PVA ratio at 1:0.5-1.5, sonicated for 2-5 min at 4°-15°C and stirred for 18-30hr. The stirred mixture is centrifuged at 8000-12000 rpm for 15-30min at 4°-20°C to obtain the pellet and washed 3-4 times with DW/NS/PBS, resuspended in same and lyophilized. The ratio of drug and polymer is kept at 1:1 w/w.
A process for the preparation of PLG-NP having ante tuberculosis drug encapsulated is explained by the following example.
EXAMPLE
lOmg INK and lOmg PZA were dissolved in ImL DW. lOmg RIF and 30mg PLG were suspended in lOmL DCM. The aqueous solution was added to the DCM solution, sonicated at 4°C for 1 min and poured into 1% PVA solution (8mL) followed by
sonication at 4°C for 3 min. The emulsion was stirred for 18hr and centrifuged at 10,000rpm for 20 min. The pellet was washed 3 times with DW and then resuspended in the same for lyophilization.
Lyophilized particles were suspended in NS and administered orally to guinea pigs and the results are given in Table 1.
Table 1-Colony forming units (CPUs) of M. tuberculosis in lungs of guinea pigs after drug treatment.

(Table Removed)
*free drugs (REF+INH+PZA) were administered orally daily for 46 days.
**PLG-NP were administered orally / aerosol every 10 days for 6weeks (a total of 5
shots).
Determination of drug content in PLG-NP
The drug encapsulation efficiency for PLG-NP were as under:
RTF - 56.99±2.72%
INH - 66.31±5.83%
PZA - 68.02±5.58%
When singularly encapsulated the drug encapsulation efficiencies is the same.
The PLG-NP did not induce any hepatotoxicity as assessed by plasma bilirubin, alanine
transaminase and alkaline phosphatase

WE CLAIM
1. A process for the preparation of nanoparticles of poly-DL-lactide-co-glycolide (PLG-NP) having two different active substances selected from hydrophobic and hydrophilic with different or same solubility properties encapsulated therein comprising steps of:
(i) preparing an aqueous solution of hydrophilic drug Isoniazid and Pyrazinamide in a solvent selected from distilled water (DW), normal saline (NS) and phosphate buffer saline (PBS) in a ratio of 1:0.1-100 weight by volume (w/v);
(ii) preparing polymer solution in an organic solvent alongwith hydrophobic drug Rifampicin wherein the ratio between poly DL-Lactide-co-glycolide and the solvent is 1:0.3-1 weight by volume (w/v) and the ratio between the drug and solvent is 1:0.5-5 weight by volume (w/v);
(iii) mixing the aqueous and polymer solution of step (i) and (ii) in a ratio of 1:5-20 volume by volume (v/v) to obtain a mixture;
(iv) sonicating the mixture of step (iii) for 45-120 seconds at 4-15°C to make a primary emulsion under cold conditions;
(v) pouring the primary emulsion of step (iv) in polyvinyl alcohol (PVA) wherein the ratio between the organic solvent of step (ii) and polyvinyl alcohol is 1:0.5-1.5 and
resonicating for 2-5 minutes at 4-15°C under cold conditions to form a secondary emulsion;
(vi) stirring the said emulsion of step (v) and centrifuging the same at 8000-12000 rpm for 15-30 min at 4-20°C to obtain pellet;
(vii) washing the pellet of step (vi) 3-4 times with a solvent selected from the said DW/NS/PBS (pH 7.2-7.4) followed by lyophilization and suspension in the solvent so as to obtain the nanoparticles of poly DL-lactide-co-glycolide.
2. A process as claimed in claim 1 wherein said organic solvent is
dichloromethane .
3. A process as claimed in claim 1 or 2 wherein said hydrophobic
drug and poly DL-Lactide co-glycolide are preferably taken in a
ratio of 1: 1 weight by weight (wt/wt)
4. A process as claimed in claim 1 wherein said polyvinyl alcohol is
having strength of 0.8-2.5%.
5. A process for the preparation of poly DL-lactide-co-glycolide
nanoparticles substantially as herein described and illustrated in
the example.