1. INTRODUCTION:

Drugs Cancer remedy approach needs to conquer physiological boundaries including vascular endothelial pores, heterogeneous blood deliver, heterogeneous structure [1], strongly relies upon on the method of delivery. In the past, many anticancer drugs had most effective confined achievement and had fundamental negative facet outcome [2,3]. Nanoparticles have attracted widespread attention in international level due to their unique functional characters along with small particle length, high stability, decrease in toxicity, tuneable hydrophilic-hydrophobic balance and the potential to to bear surface features for goal the localization, and many others. Thus, polymeric nanoparticles represent a flexible drug transport system [4].

The utility of biodegradable polymeric particles within the scale of micrometers and nanometers as a controlled release dosage form of anticancer drugs has generated giant interest. Compare to microspheres, the nanospherers ought to direct the drug to target tissues or cells through systemic circulation or throughout the mucous membrane because of their smaller dimension [5-8]. Nanoparticles had been efficaciously used for systemic, oral, pulmonary, transdermal and different administration routes for numerous purposes together with drug targeting, enhancement of drug bioavailability and safety of drug bioactivity and stability [9-11]. Also, nanoparticles can enhance the bioavailability of poorly absorbed medicine, accordingly enhancing oral transport [7,11]. Moreover, the nanoparticles are able to permeate cells for cell internalisation and connective tissue permeation and so deliver the drug successfully to the targeted tissue without clogging capillaries [11,12]. The prospective of nanoparticles is to enhance drug diffusion through biological boundaries is a regular gain for the delivery of anticancer agents.

The polymer based nanopaclitaxel is demonstrating improved pharmacokinetic profiles of the drug, for instance, increasing the half-life and tumour accumulation of paclitaxel also the surface of paclitaxel a nanoparticle gadget may be functionalized with active ligands for targeting function [13-17]. Yadav et al confirmed N-isopropylacrylamide/ vinyl pyrrolidone (NIPAAm/VP) nanoparticles viability of MCF-7 and B16F0 cells in contrast to free paclitaxel [18].

Paclitaxel encapsulated PLGA nanoparticels confirmed better in vitro cytotoxicity as compared to free Paclitaxel in numerous cases, such as glioma C6 cells [19], L. Mu et. al conclude that TPGS has the potential to act both as emulsifier or as matrix material blended with PLGA for the manufacture of nanoparticles for controlled launch of paclitaxel [20]. Fonseca et al validated that the in vitro anti-tumoral activity of Ptx-PLGA-Nanaoparticles on human small cellular lung most cancers mobile line (NCI-H69 SCLC) is greater as compared to the in vitro anti-tumoral interest of the commercial components [21]. Paclitaxel stacked PEGylated PLGA-based nanoparticels were fabricated by Danhier et al and it was verified that the tumour development restraint impact in vivo on TLT tumor of these nanoparticles is more prominent, contrasted and Taxol [22]. Kim et al formulated chitosan-covered Poly (D,L-lactide-co-glycolide) (PLGA) nanoparticels and hindered the in vitro medication discharge rate and essentially changed the zeta potential from negative (- 30.1+/-0.6mV) to positive (26+/ -1.2 mV) and furthermore the drug delivery rate from chitosancoated nanoparticles was somewhat slower than that of the uncoated nanoparticles [23]. Chakravarthi et al. [24] showed that the A 4-10 overlap increment in cell bond of paclitaxel was seen when chitosan was adsorbed or conjugated to the PLGA particles.

Chitosan-conjugated PLGA microparticles were most cytotoxic with an IC(50) estimation of 0.77μM and furthermore chitosan-PLGA microparticles clung to the outside of 4T1 cells. Bhardwaj etal. [25] devised poly(lactide-co-glycolide) (PLGA) nanoparticles utilizing a quaternary ammonium salt didodecyldimethylammonium bromide (DMAB) and the MTT and LDH assays demonstrated the surfactant to be sheltered to in vitro cell cultures at concentration <33 microM. PLGA nanoparticles arranged utilizing this stabilizer were likewise observed to be non-dangerous to cell lines. A near report was done on a progression of polymers to PVA in a 2(2) full factorial structure [26]. The impact of the concentration of PVA and the polymers tried on molecule size and zeta potential esteem and found that the Zeta potential qualities were normally marginally negative. Poly (vinyl liquor) (PVA) is the most usually utilized emulsifier [27]. Kiran Kumar et al studied the estimation of Paclitaxel in Parenterals by RP-HPLC [28,29]. Meenashi Vanathi et al evaluated the Immuno-Stimulatory Property of PLGA/PBAE Microparticles [30]. Somasundaram et al. studied the formulation of PLGA Polymeric Nanosuspension containing Pramipexole Dihydrochloride for improved treatment of Parkinson’s Diseases [31].

The present work more seriously researched PVA utilized as aqueous phase and further sought after the likelihood of applying PLGA for the production of nanoparticles for clinical organization of paclitaxel. The formulation and characterisation of the nanoparticles of this novel samples were examined. The distinctive details with different proportions of the aqueous phase, the polymer material were assessed and improved. The outcomes exhibited that PLGA could be an incredible emulsifier for creation of polymeric nanoparticles, accomplishing excellent emulsifying impacts and high medication epitome effectiveness. The ideal size and size appropriation and its similarity can be acquired.

2. MATERIAL AND METHODS:

2.1. Materials:

Paclitaxel, purchased from Sigma Aldrich, India. PLGA (50:50 and other solvents and reagents used were of analytical grade.

2.2. Preparation of Paclitaxel Nanoparticles:

Paclitaxel loaded PLGA nanoparticles were prepared by using emulsification solvent evaporation method. Organic phase consisted that paclitaxel and PLGA was dissolved in methanol- acetone mixture and PVA solution was used as an aqueous phase. The organic phase was emulsified by adding thin stream to the aqueous phase using microtip probe sonicator (ultrasonics, Bombay) at an output of 50 w for 60 sec in ice bath. The organic solution was then rapidly evaporated under reduced pressure at 37⁰C leaving behind colloidal solution of drug loaded nanoparticles in water. Further the formed nanoparticles were freeze dried at -40⁰C under nitrogen flow for 48hrs using mannitol as cryoprotectant. The freeze dried nanoparticles were stored at -40⁰C until further use. Various batches of nanoparticles were prepared using different concentrations of PLGA, PVA and time of sonication.

Table.1. List of Nanoparticles prepared based on the concentrations

Nanoparticle formulations	Conc. of PLGA % w/v	Conc of PVA % w/v	Time of sonication (Sec)
NP1	20	1	60
NP2	40	1	60
NP3	20	2	60
NP4	40	2	60
NP5	40	2	90

3. Paclitaxel and Polymer Interaction Studies

3.1. Fourier Transform Infrared Spectroscopy (FT-IR) Studies:

FTIR spectra were taken on to investigate the possible drug-excipient interactions. PLGA, placebo and drug loaded nanoparticles (optimized) were recorded using Bruker alpha E ATR spectrophotometer in the wave number region of 400-4000 cm^-1.

Table.2. Comparison of IR frequencies of Paclitaxel and Polymer based Paclitaxel

FT-IR (Frequency in cm^-1)		Vibrational Assignment
Paclitaxel	Nanopaclitaxel	Vibrational Assignment
3305.74	3309.91	N-H/O-H stretching
2942.63	2940.96	CH₃/C-H stretching
1744.84	1734.79	C=O stretching
1645.99	1645.72	C-C stretching
1318.60-1368.70	1317.71-1369.18	CH3 deformation
1246.49	1245.47	C-N stretching
1069.61	1069.63	C-O stretching
808.69-897.76	807.30-897.79	C-H bending
705.75-767.68	705.26-766.54	C-H out-of-plane/C-C=O deformation

3.2. Differential Scanning Calorimetry (DSC) Studies:

DSC analysis was performed using Mettler Toledo 822E calorimeter to study the thermal behavior of drug in the presence of polymer and other excipients of the formulation. The samples were heated in a sealed aluminum pan scanning at a rate of 10^oC/min from 25⁰C to 350⁰C under nitrogen flow (40ml/min).

Fig: 2. d. DSC Thermogram of Paclitaxel nanoparticles

3.3. Particle size analysis:

The mean particle size and width of the particle size distribution of the prepared nanoparticles were measured by a laser diffraction technique using Haribo scientific, SZ-100. Nanoparticles were sonicated for 30 min prior to the size determination to reduce the inter particle aggregation. The average particle size was measured after performing the experiment for three times.

Fig.3. Size distribution graph of NP 5 Formulation

3.4. Particle charge (zeta-potential):

The values of the zeta potential of prepared nano suspensions were measured using Haribo scientific, SZ-100 at room temperature. The average value was calculated after measuring the potential for three times.

Fig. 4. e. Zeta potential graph of NP 5 Formulation

4. RESULTS AND DISCUSSION:

Paclitaxel nanoparticles were prepared by emulsification solvent evaporation method using a Bio-degradableandeco friendly polymer, Polylactic acid co-glycolic acid (PLGA) and employing poly vinyl acetate (PVA) as an emulsifying agent [32]. Initially compatibility studies were performed on palitaxel and polymers used in the preparation using FTIR and DSC studies. Figures 1-4 are the FT-IR Spectras of paclitaxel, PLGA, PVA and Paclitaxel nanoparticles. Fig 1 shows characteristic peaks corresponds to functional groups in paclitaxel structure confirms the taken drug sample was paclitaxel and identified the same. Fig 4-the FTIR spectra of paclitaxel nanoparticles revealed that there is no disappearance of the peaks corresponds to the functional groups in paclitaxel and shifting of the existed peaks confirms the compatibility of paclitaxel with the selected polymers in the study. The compatibility of drug with polymer was further confirmed by DSC studies. Fig 5-8 are the DSC thermograms of Paclitaxel, PLGA, PVA and Paclitaxel nanoparticles. Fig-5, The DSC of paclitaxel showed the endothermic peak at 221.0˚C, which is corresponding to its melting point confirms the sample taken was paclitaxel. Figure-8 shows the DSC thermogram of paclitaxel nanoparticles, indicates that there is no drug and polymer interaction as it shows an endothermic peak at 218.45˚C corresponding to the melting point of drug, 191.59˚C correspond to the melting point of PVA and 314.36˚C corresponds to the melting point of PLGA [33,34,35]. Hence it was proved that there is no change in the melting point of the drug in the presence of polymers revealed that no change in the chemical structure of the paclitaxel and confirmed the compatibility between drug and polymers. Fig F9 to F 13 shows the distribution graphs of nanoparticle formulations. The particle size of prepared five nanoparticle formulations, NP1, NP1NP2, NP3, NP4 and NP5 are ranges from 110-350, 100-350, 130-500, 90-500, 190-300 nm and the average particle sizes are 203.3, 187.0, 266.8, 191.4, 233.1 nm respectively. The NP5 formulation shoed the narrow size distribution compared to the remaining formulations. Fig F14 to F 18 shows the zeta potential graphs of nanoparticle formulations. The zeta potential values of NP1NP2, NP3, NP4 and NP5 are-50 to + 50, 80-170, -45 to + 20, -50 to + 30, -40 to + 30 mVs and the mean zeta potential values are -11.5, +121.9, -11.5, -6.8 and – 7.2mVs respectively. From the zetapotential values it was evident that NP5 formulation shows more macroscopic stability compared with remaining formulations as the value is with in ± 40 mVs [36].

5. ACKNOWLEDGEMENT:

I would like to take this opportunity to express my profound gratitude and deep regard to University Grand Commission (UGC) who offered me the financial assistance under the scheme of UGC-Minor project and I extend by gratitude towards Management of my Institution (SITAMS, Chittoor) for the Better supporting to do the Research activities

6. CONFLICT OF INTEREST:

The author declare no conflict of interest.

7. REFERENCES:

1. Rowinsky EK, Cazenave LA, Donehower RC, Taxol: a novel investigational antimicrotubule agent. J Natl Cancer Inst.1990: 82(15); 1247-59

2. Sathyanarayana DN .Vibrational Spectroscopy Theory And Applications, 1st Edition New age .2004

3. Gowri S. A study on various computer-aided drug design methodologies. J of Chem. and Pharm.Research.2015: 7(3); 262-267.

4. http://shodhganga.inflibnet.ac.in

5. Yeh MK, Davis SS, Coombes AGA. Improving protein delivery from microparticles using blends of poly (D,L lactide-co-glycolide) and poly (ethylene oxide)-poly (pro- pylene oxide) copolymers. Pharm. Res. 1996: 13;1693- 1698.

6. Jalil R, Nixon JR. Biodegradable poly (lactic acid) and poly (lactide-co-glycolide) microcapsules: problems associated with preparative techniques and release properties. J. Microencapsul.1990: 7; 297-325.

7. Brannon-Peppas L. Recent advances in the use of bio-degradable microparticles and nanoparticles in controlled drug delivery. Int. J. Pharm.1995: 116;1-9.

8. Yogesh L. Jadhav, Bharat Parashar, Pankaj P. Ostwal, Manu S. Jain. Solid Dispersion: Solubility Enhancement for Poorly Water Soluble Drug. Research J. Pharm. and Tech. 5(2): Feb. 2012; Page 190-197.

9. Maincent P, Le Verge R, Sado P, Couvreur P, Devissaguet JP , Deposition kinetics and oral bioavailability of vincamin-loaded poly-alkyl cyanoacrylate nonparticles. J. Pharm. Sci. 1986: 75;1955-958.

10. Cappel MJ, Kreuter J. Effect of nanoparticles on transder- mal drug delivery. J. Microencapsul.1991: 8 ;369-374.

11. Couvreur P, Tulkens P, Roland M, Trouet A, Speiser P. Nanocapsules: a new type of lysosomotropic carrier. FEBS Lett. 1977:84 (2); 323-326.

12. Astier A, Doat B, Ferrer MJ, Benoit G, Fleury J, Rolland A, Leverge R. Enhancement of adriamycin antitumor activity by its binding with an intracellular sustained-release form, polymethacrylate nanospheres, in U-937 cells. Cancer Res.1988: 48;1835-1841.

13. Alexis F, Pridgen EM, Langer R, Farokhzad OC. Nanoparticle technologies for cancer therapy. Hand book Exp Pharmacol.2010:197; 55-86.

14. Wang J, Sui M, Fan W. Nanoparticles for tumor targeted therapies and their pharmacokinetics .Curr Drug Metab.2010:11; 129-141.

15. Haley B, Frenkel E. Nanoparticles for drug delivery in cancer treatment. UrolOncol . 2008: 26; 57-64.

16. Suri SS, Fenniri H, Singh B. Nanotechnology-based drug delivery systems . J Occup Med Toxicol . 2007: 2; 16.

17. Jain RK, Stylianopoulos T, Delivering nanomedicine to solid tumors. Nat Rev Clin Oncol.2010: 7;653-664.

18. Yadav D, Anwar MF, Garg V, Kardam H, Beg MN, Suri S, Gaur S, Asif M. Development of polymeric nanopaclitaxel and comparison with free paclitaxel for effects on cell proliferation of MCF-7 and B16F0 carcinoma cells. Asian Pac J Cancer Prev.2014:15(5);2335-40.

19. Dong Y, Feng SS, Poly(D,L-lactide-co-glycolide) (PLGA) nanoparticles prepared by high pressure homogenization for paclitaxel chemotherapy. Int J Pharm.2007: 342; 208-214.

20. Mu L, FengSS, Journal of Controlled Release.2003: 33–48.

21. Fonseca C, Simões S, Gaspar R. Paclitaxel-loaded PLGA nanoparticles: preparation, physicochemical characterization and in vitro anti-tumoral activity. J Control Release.2002: 83; 273-286.

22. Danhier F, Lecouturier N, Vroman B, Jérôme C, Marchand-Brynaert Jet al. Paclitaxel-loaded PEGylated PLGA-based nanoparticles: in vitro and in vivo evaluation. J Control Release.2009:133; 11-17.

23. Kim BS, Kim CS, Lee KM. The intracellular uptake ability of chitosan-coated Poly (D,L-lactide-co-glycolide) nanoparticles. Arch Pharm Res.2008: 31; 1050-4.

24. Chakravarthi SS, Robinson DH. Enhanced cellular association of paclitaxel delivered in chitosan- PLGA particles. Int J Pharm. 2011:409; 111-120.

25. Vandervoort J, Ludwig A. Biocompatible stabilizers in the preparation of PLGA nanoparticles: a factorial design study. Int J Pharm . 2002:238; 77-92.

26. Mu L, Feng SS. A novel controlled release formulation for the anticancer drug paclitaxel (Taxol): PLGA nanoparticles containing vitamin E TPGS.J Control Release.2003:86; 33-48.

27. Feng SS, Mu L, Win KY, Huang G. Nanoparticles of biodegradable polymers for clinical administration of paclitaxel. Curr Med Chem .2004:11; 413-424.

28. V Kiran Kumar, N Appala Raju, Namratha Rani, JVLN Seshagiri Rao, T Satyanarayana. The Estimation of Paclitaxel in Parenterals by RP-HPLC. Asian J. Research Chem. 2(1): Jan.-March, 2009;Page 90-92.

29. Muddukrishna B.S. Krishnamurthy Bhat, Gautham G. Shenoy. Preparation and Solid State Characterization of Paclitaxel Cocrystals. Research J. Pharm. and Tech. 7(1): Jan. 2014; Page 64-69.

30. B. Meenashi Vanathi, N. Sivagurunathan, K. Narayanan, J. Venkata Rao, N. Udupa. Evaluation of Immuno-Stimulatory Property of PLGA/PBAE Microparticles. Research J. Pharm. and Tech. 7(12): Dec. 2014; Page 1365-1367.

31. I. Somasundaram, B.V. Nagarjuna Yadav, S. Sathesh Kumar. Formulation of PLGA Polymeric Nanosuspension containing Pramipexole Dihydrochloride for improved treatment of Parkinson’s Diseases. Research J. Pharm. and Tech. 2016; 9(7):810-816 .

32. Chetan M Patel, Manan A Patel, Nikhil P Patel, PH Prajapati, CN Patel. Poly Lactic Glycolic Acid (PLGA) As Biodegradable Polymer. Research J. Pharm. and Tech. 3(2): April- June 2010kj; Page 353-360.

33. S Rajarajan, R Chandramouli. Preparation, Numerical Optimization and Evaluation of Ciprofloxacin PLGA and PLA Nanoparticles by Solvent Displacement Technique. Research J. Pharm. and Tech. 2(1): Jan.-Mar. 2009; Page 186-190.

34. Shruti Panwar, Sangeeta Loonker. Synthesis of Novel Film of Poly Vinyl Alcohol Modified Guar Gum with Tamarind seed Kernel Powder and its Characterization. Asian J. Research Chem. 2017; 10(5): 616-620.

35. G.H. Murhakar, A.R. Raut. Thermal Study of Modified Polyvinyl Alcohol Conjugates and Doped Modified Polyvinyl Alcohol Conjugates. Asian J. Research Chem. 7(11): November, 2014; Page 925-928.

36. Geetali S. Ingawale, Anita S. Goswami-Giri. Zeta Potential of Lantadene Post Alcoholic Reflux Method. Asian J. Research Chem. 6(12): December 2013; Page 1137-1139.

Received on 03.05.2019 Modified on 18.07.2019

Research J. Pharm. and Tech. 2020; 13(1): 265-269

DOI: 10.5958/0974-360X.2020.00054.2