Pharmaceutical nanoparticle technologies: An approach to improve drug solubility and dissolution rate of Piroxicam

 

Tuti Sri Suhesti^1,2*, Achmad Fudholi², Ronny Martien² , Sudibyo Martono²

¹Department of Pharmacy, Faculty of Health Sciences, Jenderal Soedirman University, Jl. Dr. Soeparno, Karangwangkal, Purwokerto 53122 Indonesia

²Faculty of Pharmacy Gadjah Mada University, Sekip Utara Yogyakarta, 55281 Indonesia

 

ABSTRACT:

Pharmaceutical nanotechnology is employed to improve poor aqueous solubility of drug compounds which have  limited in vivo bioavailability because of their low dissolution rate in the gastrointestinal fluids. Nanoparticle technology reduced particle size of piroxicam, proved to be effective in improving the oral bioavailability as a result of enhanced solubility and dissolution rate. To achieve this objective, the formulations were prepared by evaporative antisolvent precipitation methode. The first-line drug is dissolved in an organic solvent, and then quickly mixed with an aqueous solution of stabilizer media. The physical characteristics of nanoparticles, as shape, particle size, solubility and disolution rate of nanopiroxicam were evaluated. FTIR (Infrared Fourier Transform Spectroscopy),  XRD (X-ray diffraction) and differential scanning calorimetr (DSC) of the microcrystals were studied. The results showed that the nanoprecipitation method was used to prepare biodegradable drugs of reproducible sizes of piroxicam in the range (300-400 nm) with spherical shape by addressing the effects of processing parameters. Nanopiroxicam successfully improved the solubility and in vitro dissolution is expected due to of amorphous from revealed by X-ray and Differential Scanning Calorimetry (DSC) studies. The dissolution rate of the nanoparticles was markedly enhanced by reducing the particle size and a subsequently increased surface area. Nanopiroxicam formulations improved solubility and dissolution rate of piroxicam. Dissolution test showed that the C₅ and C₆₀ values of nanopiroxicams were greater than that of the commercial drugs and that of untreated Piroxicam, respectively.

 

 

INTRODUCTION:

Poor solubility is one of the major challenges in drug development today ⁽¹⁾. An estimated 40% of drugs fall under BCS Class 2 (low solubility and high permeability) or Class 4 (low solubility and low permeability)^(2,3,4). These drugs show limited bioavailability because of their low solubility ^(5,6,7). Piroxicam (Prx) is a non steroidal anti inflammatory drug, used as analgesic and therapy in acute or long-term treatment of arthritis. Prx has the characteristics of high lipophilicity, very poorly soluble in water (23 mg/mL), high molecular weight, included in the class of drugs (BCS II) with a critical point on the issue of drug dissolution^(8,9,10).  Prx is slightly soluble and consequently it critically affects its analgesic effect onset. Low solubility is a major problem in the development of drug formulation. In many cases, low solubility will result in low bioavailability ^(11,12).

 

Several approaches to overcome these problems. Different techniques have been studied to enhance the dissolution rate of Prx. Application formation of complex molecule using beta-cyclodextrin ⁽¹³⁾ has result advantages such as better powder flow and compressibility and improved Prx bioavailability. Other methods such as liquid–solid compacts ⁽⁵⁾, wet granulation using betalactose and PVP ^(14,15), and solid dispersions ^(16,17) have been proposed to enhance the dissolution rate of Prx. Another approach that began many developed to improve the dissolution rate of poor aqueous solubility of a drug entity can be addressed with various pharmaceutical particle technologies in nanotechnology ^(18,19). Nanosizing with mechanical micronization technique was utilized by Tanuwijaya et.al.,⁽²⁰⁾ to incerase the solubility of Prx. It is an energy intensive process which may bring about pharmaceuticallly dentrimental changes in crystalline form of  Prx ^(21,22). That are limited for some drugs due to their low efficiency, sometimes leading to thermal and chemical degradation of drugs, and resulting in non-uniform sized particles ^(23,24).

 

An appropriate method can be selected by considering the properties of drug to be formulated and the properties of desired dosage form. Nanoparticle formulation for Piroxicam delivery system used bottom up technique was selected for current study ⁽²⁵⁾. Evaporative methods of anti-solvent precipitation is one of the bottom up techniques used to prepare the nanoparticles Prx⁽²⁶⁾. The method was simple and convenient to reduce the drug particle size and increase the surface area and thus enhance the solubility and dissolution of poorly soluble drugs. The benefits of which this method is cost-effective, quick to perform, reproducible and is suitable for scaling up⁽²⁷⁾. Several important factors contribute to the effectiveness of this method in preparing particles with acceptable size range, shape and the percentage of  the drug load, namely the amount of   polymer, percentage of surfactant and volume of organic and aqueous phases⁽¹³⁾.

 

This research formulated nanopiroxicam particle used Polivinyl pirollidonpolymer as carriers and Chitosan as a co-stabilizer to increase stability. Evaluation of the characteristics of nanoparticles that are formed are analyzed physico-chemical and analysis to compare solubility and dissolution rate of the nanoparticles with commercial Prx and Prx not formulated.

 

MATERIALS AND METHODS:

Materials

Piroxicam was provided by Nantong Jinghua  Pharmaceutical Co Ltd, Jiangsu China, Chitosan was a Biotech Surindo, Polyvinylpyrrolidone (PVP) K30 and Sodium tripoly phosphat (STPP) were procured locally. Acetate acid, dichloromethane, Aceton, methanol and ethyl acetate were from Merck and Led for analytical grade.

 

Methods

A.    Preparation of Nanosuspensions

PRX nanosuspensions were prepared on the optimum formula by evaporative antisolvent precipitation technique⁽²⁾. The solvent in which Prx that showed highest solubility was used for preparing the nanosuspensions. Required amount of Prx was dissolved in sufficient volume of selected solvent. The polymers/surfactants each were dissolved in water (100 mL) separately and the resulting mixture was stirred at 4,000 rpm by using a mechanical stirrer. After formation of a homogenous solution, drug solution was added all at once with continuous stirring. After complete addition of the drug solution, stirring was continued for 30 minutes. The final nanosuspension Prx is dryed using spray-dryer methode.

 

B.    Particle Size Measurement

The nanosuspensions were subjected to particle size analysis based on dynamic light scattering. The measurements were made in average particle size, polydispersity index, and zeta potential. Measurements of particle size were determined by laser diffraction using Horiba (SZ-100, Z type Instruments), which is also known as low angle laser light scattering ⁽²⁾. Nanosuspensions were  suspended in bidistilled water (Merck) for each nanosuspension sample.

 

C.     Surface Morphology

Morphology of the spraydrying nanoparticle that was investigated by a scanning electron microscope (SEM). The drug particles were sputter coated with gold before observation. Particle size and morphological microstructure of nanosuspension Prx were also visualized utilizing transmission electron mycroscopy (TEM) methode.

 

D.    Determination of solubility studies

The solubility of piroxicam in water was determined by taking excess  quantity of Prx in 50 ml water. The suspensions were stirred at 22°C, filtered, and the drug content was determined by Spectrofotometer. Each sample was analyzed in triplicate. The concentration of Prx was analyzed at ƛ_max 352 nm.

 

 

E.    Dissolution studies of microparticles

The dissolution of piroxicam pure sample, nanoparticle formulations and commercial Prx  were  determined  by  using  USP  dissolution  apparatus XXIV‐Type II.  Dissolution  medium  was  900 ml aquabidest. Samples were taken at minute 0, 5, 10, 15, 30, 45, 60, and 90 as much as 5,0 mL. Sampel yang diambil diganti dengan medium disolusi baru dalam jumlah yang sama sehingga volume medium disolusi tetap. The sample taken was replaced with a new dissolution medium in the same amount. The amount of dissolved drug was determined using UV spectrophotometric method  (UV  1601  A Shimadzu, Japan) at 358 nm. The readings were taken in triplicate.

 

RESULT AND DISCUSSION:

Among the solvents, dichloromethane (DCM/€ =8.93) was selected as the solvent for PRX due to its higher solubilization potential for Prx and low boiling point. These are two important parameters as rapid evaporation of the solvent that is essential for higher supersaturation and rapid nucleation, all prerequisites for ultrafine crystal size ⁽²⁾.In this study, nanosuspension Prx were prepared using polymers with PVP K30, chitosan and STPP in ratio of 1:1:1 in combination were chosen as optimum formula.

 

Particle size analysis

Morphology and microstructure of the nanosuspension Prx were examined using TEM (Fig. 1). The nanoparticles appear to suspend as individual particles with a spherical shape. TEM images show the particle surface morphology to be smooth and non porous. The average particle size (Zav) of the plain drug suspension was found to be 200-300 nm.

 

Figure 1. TEM nanosuspension piroxicam

 

The SEM micrographs of the plain drug of Prx (untreated Prx) are highly crystalline, angular and cubic or rod shape crystals with rounded (blunt) edges, whereas micrographs of nanoPrx revealed sperical shapes, amorphus forms and are obsviously smaller crystal (Fig. 2).

 

PRX

 

Nano PRX

Figure 2. Scanning electron micrograph of Piroxicam samples

 

Untreated Prx powder had particle size with a mean diameter of 13-15 µm. The size distributions of the nanoparticles of the optimized formulations had a mean diameter of 316-319 nm.

 

Figure 3. Particle size distribution of nanoPRX

 

As shown in Figure 3, the size distribution of nanoPrx were narrow and uniform, with polidispersity index (PI) of 0,265. The narrow and uniform size distribution of the particles is one of the important advantages of the nanocrystallization process, in contrast with other particle size reduction methods as milling.⁽²³⁾

 

Particle reduction technology made an increase in particle surface area and increase in Gibbs free energy thus making thermo dynamically unstable. Chitosan as stabilizers is used in an attempt to reduce agglomeration of nanoparticles. Zeta potensial of the nanoparticles of the optimized formulations had a mean value of 18,1 mV (Fig. 4)

 

Figure 4. Zeta Potensial of Nanopiroxicam sample

 

Solubility Analysis Dissolution studies

The solubility analysis results that pure piroxicam was capable to produce solubility of 26,77 µg/mL in aquadest (37ºC), Whereas nanoparticles Prx produce solubility of 147,101 µg/mL. Figure 5 represents a comparison of the dissolution profiles of pure (untreated) Prx, the commercial and the nanoparticles of Prx. Tests carried out using a dissolution medium in water. The experiment revealed higher dissolution rate for the nano formulation compared to the untreated and the commercial Prx. The nano Prx particles dissolved up to 40% within 5 minutes. In contrast, only 9,47% of the untreated Prx and 15,305% of the commercial Prx dissolved, respectively, during the same period. These results demonstrate that the rate and extent of drug dissolution were markedly enhanced by the nanoparticles. The increased dissolution rate of Prx could be atributted to the pronounced reduction in particle size, the coresponding increased surface area, the enhanced solubility, and the amorphous nature of the drug in preparation.

Figure 5. Drug dissolution profiles of NanoPrx compared with Prx branded and untreated Prx (in water)

Table 1. Piroxicam release (%)

Sr. No.	Formulation	Piroxicam release (%)
		C5 ± SD	C60 ± SD
1	Untreated Prx	9,47 ± 1,394	23,67 ± 1,054
2	Commercial Prx	15,31 ± 1,579	37,45 ± 1,534
3	Nano Prx	40,48 ± 2,617	89,95 ± 1,643

 

Dissolution rates of nanoPrx were faster than those of the untreated Prx (Table 1). Particles size and, crystalline form are considered the main determinant factors for dissolution rate. In crystals of nanopiroxicam, higher dissolution rates appeared to result from reduced particle size and a subsequently increased surface area⁽⁹⁾. Mixing the drug with polymers resulted in increased wettability and dissolution surface area and reduced interfacial tension between the dissolution medium and the hydrophobic drug surface.⁽¹⁴⁾

 

FTIR spectroscopy

FTIR spectroscopy was used characterize possible changes in the chemical structure and drug–additive interactions in the untreated Prx and nanoPrx.

 

FTIR spectra of Piroxicam reported characteristic peaks of aromatic C-H stretching, aromatic CH bending, C-S stretching, C=O stretching, C=C, C=N ring  stretching, asymmetric S(=O)2 stretching, symmetric S(=O)2 stretching, secondary amine N-H stretching and OH stretching at 3022, 877, 690, 1741, 1527, 1348, 1149, 3338 and 3645 respectively. NanoPRX reported values of characteristic peaks were obverted with insignificant changes (fig. 6 and table 2).

 

The principle absorption bands of piroxicam appear in the regions of 3749, 1627, and 1350 cm^-1, and these bands are related to the functional groups of OH, C=O, and S=O, respectively showed  in table 2. These bands were slightly shifted to the regions of 3749, 1651, and 1327 cm^-1 in the FTIR spectra of nanoPrx. As there is no significant difference in the location of the bands, the internal structures were not different ⁽²⁸⁾, and there was no incompatibility between the drug and excipients during the procedure ⁽³⁰⁾.

 

Figure 6. FTIR spectra of (A). STPP, (B).Chitosan, (C). PVP, (D).  Untreated Piroxicam and (E). NanoPrx

 

 

Table 2. Functional groups and corresponding IR peaks of Prx

Group	Reported Values	Observed values
		PRX	NanoPRX
Aromatic C-H stretching	3100-3000	2931,80	2931,80
Aromatic C-H bending	900-675	833,25	840,96
C-S, stretching	700-600	624,94	570,93
C=O stretching	1870-1540	1627,92	1651,07
C=C, C=N ring stretching	1600-1430	1527,62	1566,20
Asymmetric S(=O)2 stretching	1350-1430	1350,17	1327,03
Symmetric S(=O)2 stretching	1160-1120	1126,43	1080,14
Secondary amine N-H stretching	3350-3310	3340,71	3433,29
OH stretching	3650-3584	3749,62	3749,62

 

X-ray Diffraction Studies

 

Figure 7. X-ray diffraction patterns of (A) Untreatted Prx and (B). NanoPrx

The XRD patterns of the nanoPrx are different to that of the Prx untreated crystalline powder. The major peaks are present in the diffractograms of the nano formulations but have less intensity than those for the untreated crystalline drug. Figure 7 shows that the XRD patterns of the nanoPrx formulations and the untreated drug powder are different. X-ray diffraction of piroxicam displayed four majorpeaks marked as 1, 2, 3 and 4 at 2θ: 8.27 (1276), 14.13 (1187), 17.32 (1736), 27.04 (1392).  These  four  major  peaks  of nanoPRx were reduced to  89,81%, 87,11%, 80,24% and 91,16% intensity respectively (Table 3). Because peak height is influenced by crystal size and crystallinity, the reduction of the height of the peaks indicates reduction of the particle size and formation of the microcrystalline form of the drug ⁽²⁷⁾. NanoPrx crystals also showed such a phenomenon in their XRD patterns ⁽²⁸⁾.

 

Table 3. XRD results and % reduction of intensity of peaks for piroxicam

No.	Major Peaks Piroxicam	Major Peaks NanoPrx	% reduction of intensity of peaks
1. 2. 3. 4.	8.27 (1276) 14.13 (1187)   17.32 (1736) 27.04 (1392)	8.28 (130) 13.76 (153) 17.36 (343) 27.56 (123)	89,81 87,11 80,24 91,16

 

Differential Scanning Calorimetry

Figure 8 shows the DSC  thermogram untreated Prx and nanoPrx, the final results for melting point and entalphy are also listed in table 4. As apparent from the figure 8, piroxicam has a sharp melting endotherm at 205.59⁰C. Cross PVP has melting at 96.96 ⁰C. However, drug melting endotherm for nano piroxicam was observed but it was slightly shifted to 117,69⁰C and an additional endotherm was also observed at 75,03⁰C  and at 198,78⁰C indicating that Prx was changed to different form.                

 

Figure 8. DSC thermograms of Untreated Prx(A) and NanoPrx (B)

 

Tabel 4. DSC results of Untreated Prx and NanoPrx

 

 

 

The main peak of piroxicam were omitted, this could mean that the entire product was changed to the amorphous form ⁽²⁹⁾. Present results in association  with  prominent  lessening  in  peaks  in  X-ray  diffraction  suggest  formation  of amorphous forms or change of different crystal lattice.

 

 

CONCLUSION:

Nanosizing is a classical approach to enhance solubility and dissolution velocity of poorly water-soluble drugs. Evaporative antisolvent precipitation technology is a simple and cost effective approach for producing nanoparticles. The drawback of solvent residue can be tackled by the use of highly volatile solvents in limited quantities. The nanoprecipitation method was used to prepare biodegradable nanosuspensions of reproducible sizes in the range (300-400 nm) with spherical shape by addressing the effects of processing parameters. Nanopiroxicam succesfully improved the solubility and in vitro dissolution is expected due to of amorphous from revealed by X- ray and Differential Scanning Calorimetry (DSC) studies. Moreover, nanoparticle technology is widely used in pharmaceutical industry and is also reported to enhance solubility.

 

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Accepted on 28.06.2016           © RJPT All right reserved