POLY (METH) ACRYLATES NANOSPHERES OF CENTRALLY ACTING ANALGESIC DRUG FOR POSTOPERATIVE PAIN: IN-VITRO AND IN-VIVO CHARACTERIZATION STUDY

POLY (METH) ACRYLATES NANOSPHERES OF CENTRALLY ACTING ANALGESIC DRUG FOR POSTOPERATIVE PAIN: IN-VITRO AND IN-VIVO CHARACTERIZATION STUDY

Sukhbir Singh ^{*1, 2}, Neelam Sharma ², Yash Paul Singla ³ Sandeep Arora ² and Jitender Madan ⁴

Department of Research ¹, Innovation and Consultancy, IKG Punjab Technical University, Jalandhar-Kapurthala Highway, Kapurthala 144603, Punjab, India.

Chitkara College of Pharmacy ², Chitkara University, Chandigarh Patiala National Highway (NH-64), Patiala-140401, Punjab, India.

Lord Shiva College of Pharmacy ³, Sirsa-125055, Haryana, India

Chandigarh College of Pharmacy ⁴, Landran, Kharar, Sahibzada Ajit Singh Nagar, 140307 Punjab, India

ABSTRACT: Postoperative pain management is prerequisite element for care of surgical patient. Objective of this study was to develop and evaluate nefopam hydrochloride loaded poly (meth) acrylates nanospheres (NFH-NS) for post-incisional pain treatment. Nefopam hydrochloride (NFH) is a centrally acting, non-opioid analgesic exclusive for treatment of post-incisional pain. NFH-NS were fabricated using eudragit RL 100: RS 100 by quasi solvent diffusion technique to provide sustained release of NFH. Entrapment efficiency (% EE), drug loading (% DL), z-average, polydispersity index and zeta potential (ζ) of NFH-NS was found 84.972 ± 1.23%, 21.41 ± 2.02 %, 648 ± 4.8 nm, 0.53 and + 4.48 mV, respectively. Scanning electron micrographs asserted smooth morphology and substantial spherical shape of nanospheres. In-vitro dissolution test revealed that NFH-NS followed fickian diffusion for drug transport. Mean dissolution time, dissolution efficiency and difference factor (f1) for NFH-NS was 5.68 h, 68.9 % and 31.83, respectively, which indicated dissimilarity in dissolution profiles as compared to NFH. % EE and % DL of NFH-NS at 25 ± 2°C/60 ± 5 % RH and 40 ± 2°C/75 ± 5 % RH revealed stability of formulation. Maximum % reversal of mechanical allodynia was achieved after peroral administration of NFH-NS (30 mg kg^–1) in rat model of post-incisional pain for 14 days. Minimum effective dose (MED) and ED₅₀ of NFH-NS was found 10 mg kg^–1 and 16.4 mg kg^–1, respectively

Postoperative pain,

Dissolution efficiency, Fickian diffusion, Mechanical allodynia

INTRODUCTION: Drug delivery is multifaceted and autonomous field of research ¹. Polymeric drug delivery systems have been extensively employed to provide captivating substitute for dynamic and long-term delivery of therapeutic agents. Polymeric dosage forms contribute many benefits such as reduced frequency of drug administration, site-specific and sustained delivery of drugs ^2-4

Polymeric nanosphere delivery systems have proficiency to enhance drug stability, upgrade its duration of therapeutic effect and diminish its degradation as well as metabolism 5-8.

Postoperative pain management is prerequisite element for care of surgical patient. Most of patients suffer from severe pain during the first few days post-surgery 9, 10. Therefore, post-incisional pain has great significance for investigation 11, 12. Nefopam hydrochloride (NFH) is a non-opioid analgesic that acts centrally by inhibiting reuptake of triple neurotransmitter i.e. dopamine (DA), noradrenalin (NA) and 5-hydroxytryptamine (5-HT) to maintain adequate monoamines in pre-synaptic cleft 11, 13. Other mode of action may be through blockage of NMDA mediated glutamatergic transmission of calcium influx or voltage-gated sodium channels which leads to anti-nociceptive effect 14-16. It has been reported in literature that plasma half-life, protein binding and oral bioavailability of NFH is 3-5 h, 75 % and 40 %, respectively 14. NFH undergoes substantial hepatic biotransformation to desmethylnefopam (which seems to be biologically active) and N-oxide-nefopam 17. Major route of elimination (87 %) is renal although a small part (8 %) is excreted in faeces 18. Nefopam 20 mg was equipotent to morphine 6-12 mg or to meperidine 50 mg 11, 19. NFH is contemplated to be safe and well tolerated because it’s reported adverse effects such as drowsiness, nausea, vomiting and sweating are minor 9, 20, 21. NFH neither affects platelet function like NSAIDS nor leads to addiction or tolerance like opioid. It has been exclusive drug for treatment of post-incisional and neuropathic pain. It can be used for relief of dental, musculo-skeletal, acute traumatic, acute wound and cancer pain 18, 22-24.

Objective of this research was to develop and evaluate nefopam hydrochloride loaded poly (meth) acrylates nanospheres (NFH-NS) for post-incisional pain management. In this study, NFH-NS were fabricated using eudragit RL 100: RS 100 by quasi solvent diffusion technique to provide sustained release of NFH and characterized for various parameters such as entrapment efficiency, drug loading, mean particle size, polydispersity index, zeta potential and scanning electron micrographs. In-vitro drug release characteristics and stability studies were also investigated. In addition, potency and efficacy of NFH-NS was investigated in a rat model of post-incisional pain.

MATERIAL AND METHODS:
Materials:
Nefopam hydrochloride (C17H20ClNO, 5-methyl-1-phenyl-1, 3, 4, 6-tetrahydro-2, 5-benzoxazocine hydrochloride, Mw 289.8 g mol–1, CAS NO-23327-57-3, 99.57% purity) was procured from Hangz Hou-Daying-Chem. Company Ltd. China. Eudragit RL 100 and RS 100 were received as gift sample supplied by Evonik Industries AG, Mumbai, India. Span 80 (sorbitan monooleate, HLB-4.3), magnesium stearate (magnesium octadecanoate, 591.27 g mol–1), sodium hydroxide, potassium dihydrogen phosphate and methanol were obtained from Loba Chemicals Private Limited, Mumbai, India. Acetone (2-propanone, C3H6O, Mw 58.08 g mol–1), heavy liquid paraffin and n-hexane (C6H14, Mw 86.18) were obtained from Merck Specialties Private Limited, Mumbai. Petroleum ether was purchased from Thomas Bakers Chemical Private Limited, Mumbai. Isofluorane was procured from Sigma, USA. All other chemicals utilized were of analytical grade.

Animals:
Animal experiments were carried out with prior permission from institutional animal ethics committee (IAEC/CCP/13/PR-010) and care of the animals was carried out as per the guidelines of committee for the purpose of control and supervision of experiments on animals (CPCSEA), ministry of environment and forest, government of India (Chitkara College of Pharmacy animal facility registration number: 1181/ab/08/CPCSEA). Rats (male wistar, 180-200 g) were used in all experiments and allowed free access to food and water. All the animals were maintained on 12 h light-dark cycle in temperature and humidity controlled rooms. Efforts were made to restraint affliction and to utilize minimum number of animals essential to achieve statistical significance.

Preparation of poly (meth) acrylates nanospheres:
Quasi solvent diffusion technique was successfully employed for fabrication of nefopam hydrochloride-loaded poly (meth) acrylates nanospheres (NFH-NS) which were composed of 1: 3 molar ratios of NFH and poly (meth) acrylates (eudragit RL 100: RS 100; 1: 2). Accurate quantity of NFH, eudragit RL 100 and RS 100 were dissolved in acetone-ethanol solvent mixture to form dispersed phase (DP). Continuous phase (CP) consist of heavy liquid paraffin containing n-hexane as hardening agent and sorbitan monooleate as an emulsifier. DP was extruded slowly through syringe # 20 to CP to form dispersion. After continuous stirring for 4 h on magnetic stirrer (Remi Instruments Division, India) at 38 ± 0.5o, dispersion was centrifuged and washed with petroleum ether. Nanospheres were accumulated by filtration utilizing 0.22 μm membrane filters followed by ultracentrifugation at 20,000 rpm for 30 minutes applying cooling centrifuge (RIS-24 BL, Remi Instruments Division, and India) and freeze-dried using lyophilizer (ISIC Make, India). Schematic representation for fabrication of NFH-NS is delineated in Fig. 1.

FIG. 1: SCHEMATIC REPRESENTATION OF EXPERIMENTAL PROCEDURE FOR FABRICATION OF NFH-NS

Characterization of NFH-NS:
Entrapment efficiency and drug loading determination:
50 mg of NFH-NS was accurately weighed and extracted with phosphate buffer, pH 7.4 for 24 h and centrifuged for 10 min at 3500 rpm. Supernatant was withdrawn and analyzed spectrophotometrically at 266 nm (Systronics AU2701). NFH concentration in the sample was determined using calibration curve regression equation. Entrapment efficiency (% EE, w/w) and drug loading (% DL, w/w) of NFH-NS was calculated by following equations 25, 26:

Eq. (1)
Eq. (2)

Particle size distribution by dynamic light scattering (DLS):
Z-average and polydispersity index (PDI) of NFH-NS was determined by dynamic light scattering (DLS) using zetasizer ver. 7.03 (Nano ZS, Malvern Instruments Ltd., UK). Analysis was performed at 25oC using disposable size cuvette at count rate of 165.6 kcps. Double distilled water with refractive index and viscosity of 1.330 and 0.8872 cP, respectively, was utilized for dilution of sample (500 ×).

Zeta potential (ζ) analysis:
NFH-NS was characterized by zeta potential (ζ) analysis using zetasizer ver. 7.03 (Nano ZS, Malvern Instruments Ltd., UK). Double distilled water with refractive index, dielectric constant and viscosity values 1.330, 78.5 and 0.8872, respectively, was used for diluting sample to appropriate concentration. Sample was placed in clear disposable zeta cell and analyzed at count rate of 29.3 kcps.

Surface morphology of nanospheres:
Surface morphology of NFH-NS was investigated using scanning electron microscopy (SEM). Nanospheres were coated with gold palladium for 150 seconds to apply 20 nm films under an atmosphere of air (Coater Polaron, 18mA current at 1.4 kV) and examined with a variable pressure scanning electron microscope (Hitachi S3400N).

In-vitro dissolution test:
In-vitro drug release profile study:
Dialysis bag diffusion technique was used for in-vitro drug release profile study of NFH-NS 27, 28. Diffusion membrane with molecular weight cut-off (MWCO) of 12,000-14,000 Da (Himedia, India) was soaked in phosphate buffer, pH 7.4 for 12 h before being used for experiment 29. NFH or NFH-NS was placed in dialysis bag which was sealed at both ends. Dialysis bag was immersed in 50 mL of phosphate buffer, pH 7.4, which was stirred at 200 rpm on magnetic stirrer (Remi, India) and maintained at 37 ± 0.5 ºC. Two mL sample was withdrawn at regular time intervals (0.5, 1, 2, 4, 6, 8, 12, 16, 24 h) and reloaded with equivalent volume of fresh release medium.

Concentration of NFH in sample was analyzed by UV spectrophotometry at 266 nm using double beam UV spectrophotometer (Systronics AU2701, India). All the experiments were performed in triplicate (n = 3), and the average values were taken. Cumulative drug released (%) vs. time (h) were plotted to observe in-vitro drug release pattern from NFH-NS and free NFH 6, 30.

In-vitro drug release kinetics analysis by mathematical-models:
In order to investigate the release mechanism of drug from NFH-NS, in-vitro drug release data was analyzed using various mathematical models i.e. zero-order kinetics, first-order kinetics, Hixson-crowell cube root model, higuchi model, Korsmeyer-peppas model, and weibull model employing Eqs. (3) - (8), respectively. Plotted data were fitted with linear regression to obtain regression equation and coefficient (r2) 31, 32.

Eq. (3)
Eq. (4)
Eq. (5)
Eq. (6)
Eq. (7)
Eq. (8)

Where, F denotes fraction of drug released up to time t; K0, Kf, KH, p, KP, K1/3, td and β are parameters of models.

Comparison of in-vitro drug release profile by model-independent methods:
Model independent methods i.e. ratio test and pair-wise procedure were used for comparison of in-vitro drug release profiles of NFH and NFH-NS. For ratio test, comparison was made on basis of mean dissolution time (MDT) calculated using following equation:

Eq. (9)

Where, j is sample number; n is number of dissolution sample times; t̂j and ∆Mj are time at midpoint and additional amount of drug dissolved between tj and tj–1, respectively.

For pair-wise comparison, difference factor (f1) which measured percent error between in-vitro drug release curves of NFH and NFH-NS over all time intervals was calculated using Eq. (10). Dissolution efficiency (% DE) used to characterize drug release profile was calculated applying Eq. (11) and (12) 31, 33, 34.

Eq. (10)
Eq. (11)
Eq. (12)
Where, n is sampling number; R and T is percent dissolved of NFH and NFH-NS at each time point j; SA = Shaded area under in-vitro drug release curve up to time t; R = Rectangle area (y100 × t) described by 100% dissolution up to time t and y is drug percent dissolved at time t.

Stability study:
Following ICH guidelines Q1A (R2), NFH-NS were kept in sealed glass vials at elevated temperatures and relative humidity in stability analysis test chamber (CHM 10S, REMI, India) at 25 ± 2°C/60 ± 5% RH and 40 ± 2°C/75 ± 5 % RH over a period of 12 months for conducting long term and accelerated stability testing, respectively 35, 36. Samples were withdrawn at preset time intervals of 3, 6 and 12 months and characterized for change in % EE and % DL (as per aforementioned protocol).

In-vivo evaluation:
Drug administration:
NFH and NFH-NS were dissolved in phosphate-buffer solution. Isofluorane was used for anesthesia in incisional pain model. Animals were randomly divided into several experimental groups: 1 - phosphate-buffer solution (5 ml kg–1) treated control group, 2 - NFH (10 mg kg–1) treated reference group, 3 - NFH-NS (10 mg kg–1) treated test group-I, 4 - NFH-NS (20 mg kg–1) treated test group-II, and 5 - NFH-NS (30 mg kg–1) treated test group-III. Sham group was included in incisional pain model. All drug preparations were administered by peroral (p.o) route using oral gavage.

Post-incisional pain study by incisional pain model:
Surgery: Surgery of rats was conducted after anaesthetization with 2% isofluorane. Using sterile technique, 1 cm longitudinal incision was made with 10 # scalpel, through skin and fascia of plantar aspect of left hind paw, starting 0.5 cm from proximal edge of heel and extending toward toes. Skin was opposed with two single interrupted sutures using 5-0 nylon with gentle pressure. Povidone-iodine antibiotic powder (Cipladine, Cipla Ltd. Mumbai, India) was sprinkled over wound site and animals were allowed to recover in their home cages 10, 12, 37. Sham-operated rats received anesthesia, but not an incision.

Treatment of injury:
Von Frey paw withdrawl thresold (VFWT) was determined prior to surgery (baseline) on day 0 and 24 h post-ligation (pre-dose) on day 1, for each rat of different groups. Drug preparations were administered immediately after the pre-dose test and continued daily to day 14 post-ligation. Behavioral test (mechanical allodynia) was recorded at different time intervals (0.5, 1, 2, 4, 6, 8, 10 h, 3, 5, 7 and 14 day) post-dose treatment.

Behavioral test (mechanical allodynia):
Automated electronic von Frey nylon monofilaments (Stoelting, USA) were used to determine VFWT to non-noxious mechanical stimulus. Each rat was positioned on wire mesh cage with an uplifted metal screen surface. Electronic von Frey employed a single nonflexible filament to which experimenter implemented an increasing force. Stimulus was applied progressively to plantar surface of left hind paw of individual animal. End point was contemplated as nocifensive paw withdrawal 38. Percent reversal of allodynia was determined using following equation:

Eq. (14)

Statistical Analysis:
All the results were represented as mean value ± standard deviation. Two-way ANOVA followed by bonferroni post-test for comparison and significance study was performed using GraphPad Prism version 5.01 for windows (GraphPad Software, San Diego California, USA). Statistical difference (p < 0.05) was considered significant. Dose that produced 50% of the maximum percent reversal (ED50) was calculated using the curve-fitting functions in GraphPad Prism 5.01.

RESULT AND DISCUSSION:
Characterization of NFH-NS:
Entrapment efficiency and drug loading of NFH-NS was 84.972 ± 1.23% and 21.41 ± 2.02%, respectively. Entrapment and loading capacity of fabricated nanospheres was found satisfactorily high. Z-average and polydispersity index of nanospheres was found 648 ± 4.8 nm and 0.53, respectively (Fig. 2). Data illustrated homogeneity with comparatively narrow particle size distribution in nanometric range. Zeta potential (ζ) value of NFH-NS was determined + 4.48 mV (Fig. 3). Positive value of zeta potential may be attributed to cationic poly (meth) acrylates with quaternary ammonium group. Scanning electron micrographs asserted that nanospheres were substantially spherical in shape with smooth morphology (Fig.4).

Statistical Analysis:
All the results were represented as mean value ± standard deviation. Two-way ANOVA followed by bonferroni post-test for comparison and significance study was performed using GraphPad Prism version 5.01 for windows (GraphPad Software, San Diego California, USA). Statistical difference (p < 0.05) was considered significant. Dose that produced 50% of the maximum percent reversal (ED50) was calculated using the curve-fitting functions in GraphPad Prism 5.01.

RESULT AND DISCUSSION:
Characterization of NFH-NS:
Entrapment efficiency and drug loading of NFH-NS was 84.972 ± 1.23% and 21.41 ± 2.02%, respectively. Entrapment and loading capacity of fabricated nanospheres was found satisfactorily high. Z-average and polydispersity index of nanospheres was found 648 ± 4.8 nm and 0.53, respectively (Fig. 2). Data illustrated homogeneity with comparatively narrow particle size distribution in nanometric range. Zeta potential (ζ) value of NFH-NS was determined + 4.48 mV (Fig. 3). Positive value of zeta potential may be attributed to cationic poly (meth) acrylates with quaternary ammonium group. Scanning electron micrographs asserted that nanospheres were substantially spherical in shape with smooth morphology (Fig.4).

In-vitro dissolution study:
NFH was released from NFH-NS following biphasic pattern with an initial ‘burst release’ of loosely bound NFH on or near the surface of particles during first 4 h and then sustained release of remaining drug from core due to diffusion from polymer matrix over 24 h. However, NFH rendered a rapid release of drug within 6 h (Fig. 5). Mechanism of drug release from NFH-NS was resolved by finding R2 value for various mathematical kinetic models viz. zero-order kinetics, first-order kinetics, Hixon-crowell, higuchi, korsmeyer-peppas and weibull model (Fig.6). In-vitro drug release of NFH-NS was best explained by korsmeyer-peppas equation with regression coefficient (r2), 0.988 (Fig. 6e). This indicted that NFH-NS followed fickian diffusion drug transport mechanism 31.

MDT for pure NFH and NFH-NS was found to be 2.12 and 5.68 h, respectively, (Table 1) which indicated that in-vitro drug release from nanospheres was considerably slower than from pure drug. Reason behind this might be entrapment of drug in poly (meth) acrylates polymers which gets swell in presence of water and digestive fluids and exhibit slow release by diffusion. % DE for NFH and NFH-NS was found 18.95 and 68.9 %, respectively (Table 1). Higher dissolution efficiency of NFH-NS could be due to increased surface area to volume ratio. In order to consider similar dissolution profiles between reference and test formulation, f1 values should be lower than 15 (0-15). Difference factor (f1) between NFH-NS and NFH was 31.83 which indicated dissimilarity between dissolution profiles as the value was > 15.

TABLE 1: DISSOLUTION EFFICIENCY (% DE) AND MEAN DISSOLUTION TIME (MDT) OF NFH AND NFH-NS
Formulation % DE MDT
NFH 18.95 2.12
NFH-NS 68.90 5.68

Stability study:
Long term and accelerated stability testing of NFH-NS was conducted as per ICH guidelines Q1A (R2) and ICH Q1. NFH-NS were characterized for change in % EE and DL at predetermined time intervals of 3, 6 and 12 months. It was observed that there was no significant change (p > 0.05) in % EE and DL of samples kept at 40 ± 2°C/75 ± 5 % RH as compared to those placed at 25 ± 2°C/60 ± 5% RH (Fig. 7) which revealed stability of formulation on storage at elevated temperature and humidity conditions.

FIG. 7: % EE AND % DL OF NFH-NS AGAINST STORAGE TIME AT 25 ± 2 oC/60 ± 5% RH AND 40 ± 2 oC/75 ± 5 % RH 25oC

In-vivo evaluation:
Postoperative pain study was performed by incisional pain model to compare NFH (10mg kg-1) and NFH-NS (10-30 mg kg–1) employing mechanical allodynia for behavioral testing of rats. VFWT determined by electronic von Frey assay prior to incision was 75.1 ± 2.4 (baseline). All rats underwent VFWT testing before treatment (pre-dose) followed by post-dose testing. Control group exhibited strong allodynia during experimental period as compared to NFH-NS treated groups (p < 0.001). On the contrary, phosphate-buffer treated sham group did not display mechanical allodynia during experimental period. NFH-NS 10-30mg kg–1 treated group produced significant difference in VFWT till 14 day of drug administration as compared to NFH 10 mg kg–1 treated reference group (p < 0.05, p < 0.01, p < 0.001) (Fig. 8). Data illustrated sustained release of drug from NFH-NS. Furthermore, it was investigated that NFH-NS 20-30 mg kg–1 treated animals exhibited significantly higher VFWT as compared to NFH-NS 10 mg kg–1 treated animals (p < 0.001, at all time points) which manifested that NFH-NS produced dose-dependent attenuation of pain.

Minimum effective dose (MED), onset and duration of action for NFH-NS was retrieved from the curve obtained in incisional pain model (Fig.8). MED, minimum dose that elicited statistically significant response as compared to vehicle-treated controls was 10 mg kg–1. Onset of action, earliest detected statistically significant response with MED was found 0.5 h. NFH-NS exhibited long duration of action as indicated by statistically significant VFWT at various time points post-dose at MED (10 mg kg–1) (Fig. 8, Table 2).

FIG. 8: VFWT FROM VARYING DOSES OF NFH-NS ADMINISTERED VIA P.O. ROUTE (N = 5). ASTERISK INDICATES SIGNIFICANT DIFFERENCE BETWEEN NFH-NS AND NFH (*P < 0.05, ɸP < 0.01, #P < 0.001). BL INDICATES BASELINE

Dose response curve was constructed between dose (NFH-NS 10-30 mg kg–1) and maximum % reversal of allodynia at corresponding dose. Maximum percent reversal was reported for highest studied dose 30 mg kg–1 (Fig. 9). Dose that produced 50 % of maximum percent reversal (ED50) as calculated by curve-fitting functions in GraphPad Prism 5.01 was found 16.4 mg kg–1 (Table 2).

FIG. 9: MAXIMUM % REVERSAL OF ALLODYNIA PRODUCED BY NFH-NS (10–30 MG KG–1, P.O.) (N = 5)

TABLE 2: REPORTS OF NFH-NS AGAINST TACTILE ALLODYNIA IN RAT MODEL OF INCISIONAL PAIN
Compound NFH-NS
Doses (mg kg–1) 10, 20, 30
Route p.o
MED (mg kg–1) 10
Onset of action (h) by 0.5
Duration of action long
Maximum percent reversal 72, 84, 92 %
ED50 (mg kg–1) 16.4

CONCLUSION: Our findings clearly demonstrated that poly (meth) acrylates nanospheres of nefopam hydrochloride synthesized by quasi solvent diffusion technique followed fickian diffusion for drug transport with an initial ‘burst release’ succeeded by sustained release over 24 h. Long term and accelerated stability testing revealed stability of NFH-NS on storage at 25 ± 2°C/60 ± 5% RH and 40 ± 2°C/75 ± 5 % RH. It was investigated that maximum reversal of mechanical allodynia was achieved after p.o administration of NFH-NS (30 mg kg–1) in a rat model of post-incisional pain study. We have researched the potency and efficacy of NFH-NS in a rat model of post-incisional pain. MED and ED50 of NFH-NS was found 10mg kg–1 and 16.4mg kg–1, respectively. Current investigation conclusively manifested sustained as well as dose dependent effect of NFH-NS. These findings should prove useful in the study of post-incisional pain and assessment of novel treatments.

ACKNOWLEDGEMENTS: The authors wish to thank Chitkara University for providing platform to conduct this research work. The support for providing access to Science Direct and anti-plagiarism software from department of Research, Innovation and Consultancy, IKG Punjab Technical University, Jalandhar, is greatly acknowledged. The authors are grateful to Evonik Industries AG, Mumbai, India for providing gift sample of eudragit RL 100 and RS 100.

REFERENCES:
1. Suphiya P, Misra R, Sahoo SK: Nanoparticles: a boon to drug delivery, therapeutics, diagnostics and imaging. Nanomedicine: Nanotechnology, Biology, and Medicine 2012; 8:147-166.
2. Gautier JC, Grangier JL, Barbier A, Dupont P, Dussossoy D, Pastor G, Couvreur P: Biodegradable nanoparticles for subcutaneous administration of growth hormone releasing factor (hGRF). Journal of Controlled Release 1992; 20:67-78.
3. Gulyaev AE, Gelperina SE, Skidan IN, Antropov AS, Kivman GY, Kreuter J: Significant transport of doxorubicin into the brain with polysorbate 80-coated nanoparticles. Pharmaceutical Research 1999; 16:1564-1569.
4. Jiao Y, Ubrich N, Marchand-Arvier M, Vigneron C, Hoffman M: In-vitro and in-vivo evaluation of oral heparin-loaded polymeric nanoparticles in rabbits. Circulation 2002; 105:230-235.
5. Couvreur P: Nanoparticles in drug delivery: Past, present and future. Advanced Drug Delivery Reviews 2013; 65:21-23.
6. Joshi G, Kumar A, Sawant K: Enhanced bioavailability and intestinal uptake of Gemcitabine HCl loaded PLGA nanoparticles after oral delivery. European Journal of Pharmaceutical Sciences 2014; 60:80-89.
7. Li X, Xu Y, Chen G, Wei P, Ping Q: PLGA nanoparticles for the oral delivery of 5-Fluorouracil using high pressure homogenization-emulsification as the preparation method and in-vitro/in-vivo studies. Drug Development Industrial Pharmacy 2008; 34:107-115.
8. Sarmento B, Ribeiro A, Veiga F, Ferreira D, Neufeld R: Oral bioavailability of insulin contained in polysaccharide nanoparticles. Biomacromolecules 2007; 8:3054-3060.
9. Kapfer B, Alfonsi P, Guignard B, Sessler DI, Chauvin M: Nefopam and ketamine comparably enhance postoperative analgesia. Anesthesia & Analgesia 2005; 100:169-174.
10. Suto T, Obata H, Tobe M, Oku H, Yokoo H, Nakazato Y, Saito S: Long-term effect of epidural injection with sustained-release lidocaine particles in a rat model of postoperative pain. British Journal of Anaesthesia 2012; 109:957-967.
11. Evans MS, Lysakowski C, Trame`r MR: Nefopam for the prevention of postoperative pain: quantitative systematic review. British Journal of Anaesthesia 2008; 101:610-617.
12. Whiteside GT, Harrison J, Boulet J, Mark L, Pearson M, Gottshall S, Walker K: Pharmacological characterization of a rat model of incisional pain. British Journal of Pharmacology 2004; 141: 85-91.
13. Piercey MF, Schroeder LA: Spinal and supraspinal sites for morphine and nefopam analgesia in the mouse. European Journal of Pharmacology 1981; 74:135-140.
14. Kyung HK, Salahadin A: Rediscovery of nefopam for the treatment of neuropathic pain. Korean Journal of Pain 2014; 27:103-111.
15. Löscher W, Schmidt D: New Horizons in the development of antiepileptic drugs: Innovative strategies. Epilepsy Research 2006; 69:183-272.
16. Torres GE, Gainetdinov RR, Caron MG: Plasma membrane monoamine transporters: structure, regulation and function. Nature Reviews Neuroscience 2003; 4:13-25.
17. Aymard G, Warot D, Demolis P, Giudicelli JF, Lechat P, Le Guern ME, Alquier C, Diquet B: Comparative pharmacokinetics and pharmacodynamics of intravenous and oral nefopam in healthy volunteers. Pharmacology Toxicology 2003; 92:279-286.
18. Heel RC, Brogden RN, Pakes GE, Speight TM, Avery GS: Nefopam: a review of its pharmacological properties and therapeutic efficacity. Drug 1980; 19:249-267.
19. Tigerstedt I, Sipponen J, Tammisto T, Turunen M: Comparison of nefopam and pethidine in postoperative pain. British Journal of Anaesthesia 1977; 49:1133-1138.
20. Du MB, Aubrun F, Langlois M, Le Guern ME, Alquier C, Chauvin M, Fletcher D: Randomized prospective study of the analgesic effect of nefopam after orthopaedic surgery. British Journal of Anaesthesia 2003; 91:836-841.
21. Mimoz O, Incagnoli P, Josse C, Gillon MC, Kuhlman L, Mirand A, Soilleux H, Fletcher D: Analgesic efficacy and safety of nefopam vs. propacetamol following hepatic resection. Anaesthesia 2001; 56:520-525.
22. Brun H, Paul M, Razzouq N, Binhas M, Gibaud S, Astier A: Cyclodextrin inclusion complexes of the central analgesic drug nefopam. Drug Development Industrial Pharmacy 2006; 32:1123-1134.
23. Dordoni PL, Della VM, Stefanelli A, Iannace E, Paparella P, Rocca B, Accorra F: Effect of ketorolac, ketoprofen and nefopam on platelet function. Anaesthesia 1994; 49:1046-1049.
24. Gasser JC, Bellville JW: Respiratory effects of nefopam. Clinical Pharmacology and Therapeutics 1975; 18:175-179.
25. Ranjan AP, Mukerjee A, Helson L, Vishwanatha JK: Scale up, optimization and stability analysis of Curcumin C3 complex-loaded nanoparticles for cancer therapy. Journal of Nanobiotechnolgy 2012; 10:1-18.
26. Tang J, Xu N, Ji H, Liu H, Wang Z, Wu L: Eudragit nanoparticles containing genistein: formulation, development, and bioavailability assessment. International Journal of Nanomedicine 2011; 6:2429-2435.
27. Leroueil-Le VM, Fluckiger L, Kim YI, Hoffman M, Maincent P: Preparation and characterization of nanoparticles containing an antihypertensive agent.
European Journal of Pharmaceutics and Biopharmaceutics 1998; 46:137-143.
28. Holm R, Müllertz A, Christensen E, Høy CE, Kristensen HG: Comparison of total oral bioavailability and the lymphatic transport of halofantrine from three different unsaturated triglycerides in lymph-cannulated conscious rats. European Journal of Pharmaceutical Sciences 2001; 14:331-337.
29. Kathleen D, Jo V, Guy VM, Annick L: Evaluation of ciprofloxacin-loaded Eudragit RS100 or RL100/PLGA nanoparticles. International Journal of Pharmaceutics 2006; 314:72-82.
30. Sahana B, Santra K, Basu S, Mukherjee B: Development of biodegradable polymer based tamoxifen citrate loaded nanoparticles and effect of some manufacturing process parameters on them: a physicochemical and in-vitro evaluation. International Journal of Nanomedicine 2010; 5:621-630.
31. Costa P, Sousa Lobo JM: Modeling and comparison of dissolution profiles. European Journal of Pharmaceutical Sciences 2001; 13:123-133.
32. Karasulu E, Yesim KH, Ertan G, Kirilmaz L, Guneri T: Extended release lipophilic indomethacin microspheres: formulation factors and mathematical equations fitted drug release rates. European Journal of Pharmaceutical Sciences 2003; 19:99-104.
33. Khan KA: The concept of dissolution efficiency. Journal of Pharmacy and Pharmacology 1975; 27:48-49.
34. Khan KA, Rhodes CT: Effect of compaction pressure on the dissolution efficiency of some direct compression systems. Pharmaceutica acta Helvetiae 1972; 47:594-607.
35. Madaswamy SM, Si-Shen F: Pharmaceutical stability aspects of nanomedicines. Nanomedicine 2009; 4:857-860.
36. Mulik R, Mahadik K, Paradkar A: Development of curcuminoids loaded poly (butyl) cyanoacrylates nanoparticles: physicochemical characterization and stability study. European Journal of Pharmaceutics and Biopharmaceutics 2009; 37:395-404.
37. Brennan TJ, Vandermeulen EP, Gebhart GF: Characterization of a rat model of incisional pain. Pain 1996; 64:493-501.
38. Stuesse SL, Crisp T, McBurney DL, Schechter JB, Lovell JA, Cruce WL: Neuropathic pain in aged rats: behavioral responses and astrocytic activation. Experimental Brain Research 2001; 137:219-227.

How to cite this article:

Singh S, Sharma N, Singla YP Arora S and Madan J: Poly (Meth) Acrylates Nanospheres of Centrally Acting Analgesic Drug for Postoperative Pain: In-Vitro and In-Vivo Characterization Study. Int J Pharm Sci Res 2016; 7(5): 1967-77.doi: 10.13040/IJPSR.0975-8232.7(5).1967-77.

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