63 ISSN 0976-1047 2229-7499 International Journal of Biopharmaceutics Journal homepage: www.ijbonline.com IJB FORMULATION, CHARACTERIZATION AND BIOPHARMACEUTICAL EVALUATION OF ALLOPURINOL TABLETS Amal A. Ammar* 1, Ahmed M. Samy 2, Maha A. Marzouk 3, Maha k. Ahmed 4 1, 3 & 4 Department of Pharmaceutics, Faculty of Pharmacy (Girls), Al-Azhar University, Cairo, Egypt 2 Department of Pharmaceutics, Faculty of Pharmacy (Boys), Al-Azhar University, Cairo, Egypt ABSTRACT Allopurinol is a poorly water-soluble drug so the solubility is the main constraint for its oral bioavailability. Solid dispersions consisting of Allopurinol with different types of polymers were prepared by melting or solvent evaporation technique. Three formulations exhibited highest drug release after 45 min were incorporated into tablet matrix. Fourier transform Infrared spectroscopy (FTIR) and differential scanning calorimeter (DSC) were performed to identify any physicochemical interactions between Allopurinol and the used tablet excipients. Tablet formulations were also subjected to stability studies. FT1 and FT2 contained solid dispersions with urea and mannitol respectively showed the highest shelf stability t 90 (year) 2.29, 7.11 respectively. On the basis of the in-vitro release profile and stability data both FT1 and FT2 were subjected to bioavailability studies in human, and were compared with commercial tablets. Tablets containing solid dispersion showed higher AUC compared to the commercial tablets. These results suggest that the Allopurinol solid dispersion loaded tablets can be utilized to improve its bioavailability. Keywords: Allopurinol tablets; Bioavailability; Dissolution enhancement; Pharmacokinetics; Xanthine oxidase inhibitor. INTRODUCTION Allopurinol is an inhibitor of the enzyme commonly known as xanthine oxidase. Allopurinol is an analogue of hypoxanthine. It is effective for the treatment of both primary hyperuricemia of gout and secondary hyperuricemia related to hematological disorders or antineoplastic therapy (Clark s, 2004; Derek and Da-peng wang, 1999). It is a very weak acid with a dissociation constant (pka) of 9.4 and is therefore essentially unionized at all physiological ph values (Benzra and Bennett, 1978). Its lipid solubility is quite low as is indicated by its octanol: water partition coefficient of 0.28 (Day et al., 2007). Allopurinol is a polar compound with strong Corresponing Author Amal A. Ammar E-mail: amal_ammar@azhar.edu.eg intermolecular hydrogen bonding and limited solubility in both polar and non polar media (Samy et al., 2000; Ammar and El-Nahhas, 1995; Hamza and Kata, 1989 ). Oral bioavailability of a drug depends on its solubility and or dissolution rate, therefore efforts to increase dissolution of drugs with limited solubility is often needed. Solid dispersion techniques have been widely used to improve the dissolution properties and bioavailability of poorly water soluble drugs (Jagdale et al., 2010). Solid dispersion refers to a group of solid products consisting of at least two different components, generally a hydrophilic matrix and a hydrophobic drug. The matrix can be either crystalline or amorphous. The drug can be dispersed molecularly, in amorphous particles (clusters) or in crystalline particles. It is also defined as being a product formed by converting a fluid drug-carrier combination to the solid state (Aggarwal et al., 2010).
64 Several carrier systems have been used in the preparation of fast release solid dispersions. The technique provides a disposition of the drug on the surface of certain materials that can alter the dissolution properties of the drug. Once the solid dispersion is exposed to aqueous media and the carrier is dissolved, the drug is released as very fine colloidal particles (Zedong et al., 2008; Devi, 2003; Vippagunta et al., 2002). This results in a greatly enhanced surface area, thus prompting expectations of a high dissolution rate and level of bioavailability for poorly water-soluble drugs (Goldberg, 1966). The aim of the present study was to formulate tablets of Allopurinol solid dispersions in order to improve dissolution and aqueous solubility and to facilitate faster onset of action. In a previous study (Samy AM et al., 2010) several carriers were used in the preparation of solid dispersions, including urea, mannitol and PVP K30. In the current work we choose the three formulations which exhibited highest drug release after 45 min and incorporated them into compressed tablets. The prepared tablets were also evaluated for their uniformity of weight, thickness, hardness, friability, disintegration time and drug content uniformity. The shelf storage stability testing at room temperature for one year was carried out on the formulated Allopurinol tablets. Formulations showed improved dissolution and higher stability data were chosen for in-vivo absorption study in comparison with commercial product. MATERIALS & METHODS Materials Allopurinol (Allo) powder was kindly provided by Alexandria Company for pharmaceutical industries, (Alexandria, Egypt); Urea, Sodium chloride, Lactose, Anhydrous sodium acetate, salicylic acid, acetic acid, disodium hydrogen phosphate, potassium di-hydrogen ortho phosphate, ethyl alcohol (absolute) and Hydrochloric acid were supplied from El-Nasr Pharmaceutical chemicals Co., (Egypt); Mannitol, Magnesium stearate, Polyvinylpyrrolidone (PVP) K30 and Polyethylene glycol (PEG) 4000 were kindly provided by Amoun Company for pharmaceutical industries, (Cairo, Egypt); Avicel PH101, Fluka AG, CH 9470 Buchs., Mittler Teilchengrosse (Switzerland); Acetonitrile and methanol (HPLC grade), Scharlau chemie S.A., European Union. (Zyloric 100 mg tablet), Glaxo smithkline, Egypt. METHODS Study of Physicochemical interaction of Allopurinol with tablet excipients Differential scanning calorimetric (DSC) studies Possible interaction of Allopurinol with the tablet excipients was investigated using DSC. Approximately 5 mg of samples were weighed and hermetically sealed in the aluminium pans. Samples of drug alone, each excipient alone, physical mixtures of Allopurinol with the investigated excipients (1:1 W/W) were measured with Shimadzu, (model DSC-50, Japan) thermal analyzer. The DSC thermogram were obtained over a temperature range of 25 400 ºC with a thermal analyzer equipped with an advanced computer software program at a scanning rate of 10ºC / min and nitrogen gas purge of 40 ml/ min. The instrument was calibrated with pure indium as a reference. Fourier Transforms Infrared Spectroscopy (FTIR) Samples of 1-2 mg of drug alone, each excipient alone, physical mixtures of Allopurinol with the investigated excipients (1:1 W/W) prepared by simple and perfect mixing and solid dispersion (1:1 W/W) were mixed with KBr (IR grade) compressed into discs in the compression unit under vacuum and were scanned from 4000 400 cm -1 with an empty pellet holder as a reference. The spectrophotometer was Perkin-Elmer, FTS-1710, Beaconsfield (UK). Formulation of Allopurinol tablets. Allopurinol solid dispersions with different carriers like (urea, mannitol and PVP K30) in different ratios prepared by melting or solvent evaporation techniques were incorporated into tablet formulations. Tablet compression machine with flat-faced single punch, first medicine machinery, factory of Donghai branch, (China, Shanghai) was used for the manufacturing of the directly compressed Allopurinol tablets. Additives used in preparation of tablets (Avicel PH101, magnesium stearate and lactose) were incorporated by the ratio showed in table (1). Quality control study of the prepared tablets The prepared tablets from each formulation were subjected to the tablets quality control tests as drug content, weight uniformity, tablets thickness, disintegration time, hardness and friability. Drug release was assessed using a USP type II dissolution apparatus at 75 rpm in 900 ml 0.1N HCl maintained at 37ºC ± 0.5ºC (Abd-Elazeem, 2001). Sample of 5ml was withdrawn at regular intervals and replaced with the same volume of prewarmed (37ºC ± 0.5ºC) fresh dissolution medium. The samples withdrawn were filtered through Whatman filter paper (No. 1, Whatman, Maidstone, UK) and drug content in each sample was analyzed after suitable dilution, the amount of Allopurinol dissolved was determined spectrophotometrically at 250 nm. Plain Allopurinol tablets were used as a control. Shelf Stability study of Allopurinol tablets Stability studies were conducted on Allopurinol tablets containing its solid dispersions to assess their shelf stability with respect to their drug content, after storing them at room temperature for one year.
65 Bioavailability study The selected tablet formulations with the highest dissolution profile (FT1, FT2) and commercial tablets (Zyloric 100mg tablets, Glaxo-Wellcome) were subjected to a single dose relative pharmacokinetic study. The study was performed following standard protocol in 6 healthy male volunteers, weighing 60 to 85 kg and of 22 to 30 years old in a cross-over design with two weeks wash out period in accordance with all applicable regulations. The study was reviewed and approved by the Ethical and Institutional Review Committee of the Pharmaceutical Analytical Unit Faculty of Pharmacy, (Boys), Al-Azhar University, Nasr City, Cairo, Egypt. Before initiating the study, informed consent was obtained from volunteers after the nature and possible consequences of the study were explained. All the subjects were in good health on the basis of their medical history and complete physical examination. The volunteers did not smoke and were not on any kind of medication before or during the experiment. Venous blood samples were collected pre-dose (0hours) and at0.5, 0.75, 1, 1.25, 1.5, 2, 3, 4 and 8 h post-dosing. The blood samples were centrifuged at 5,000 rpm for 10 min, and the plasma obtained were stored at -80 C until analysis. To compare the rate and extent of absorption of Allopurinol, the following pharmacokinetic variables were calculated for each volunteer using actual blood sampling times. The maximum plasma concentration (C max ) and the time required to reach this concentration (T max ) were read directly from the arithmetic plot of time vs. plasma concentration for Allopurinol. The overall elimination rate constant (ke) was calculated from the slope of the terminal elimination phase of a semilogarithmic plot of concentration vs. time after subjecting it to linear regression analysis. The elimination half life (t 1/2 ) was obtained by dividing 0.693 by ke. The absorption rate constant (ka) was calculated using the method of residuals (Gibaldi and Perrier, 1990). The area under the plasma Allopurinol concentration vs. time curve (AUC 0-8 ) was determined by means of the trapezoidal rule. The relative bioavailability of Allopurinol from matrix tablets in comparison to reference formulation (Zyloric 100mg tablets, Glaxo-Wellcome) commercial tablets was calculated by dividing its AUC 0-8 by that of the commercial tablet dosage form. Validation of the HPLC method Allopurinol was subjected to analytical validation in human plasma using an HPLC method according to USP guidelines, from which the recovery of the prepared tablets can be calculated. Preparation of standard solutions Stock solution of Allopurinol was prepared by dissolving 10 mg of Allopurinol in 100 ml methanol. This solution was used to prepare working standard solutions daily for different concentrations by dilution with methanol. The internal standard solution was prepared by dissolving 10 mg salicylic acid in 100 ml methanol. The working internal standard was prepared by taking 3 ml from this solution in 10 ml methanol (30µg/ml). Linearity The linearity of the method was evaluated using a calibration curve in the range of 0.5-8 µg/ml Allopurinol. Thirty μl injections were made in triplicate for each concentration and chromatographed on a C18 column using a freshly prepared mobile phase consisted of 2.72 gm of sodium acetate per liter distilled water adjusted to ph 4.5 with a mixture of acetic acid: acetonitrile (96:4). The mobile phase was degassed and filtered through a 0.45 µm filter (Millipore, Sainet-Quentin, Y-velines). The flow rate was 1 ml/minute. The detection wave length was 254 nm. The run time was 20 minutes. The calibration curve was obtained by plotting the peak area as a function of drug concentration and the regression parameters were determined. Intra-day and inter-day reproducibility of Allopurinol The intra-day and inter-day reproducibility were determined by replicate analysis of three sets of samples spiked with different concentrations of Allopurinol (0.5, 1, 2, 4, 6, and 8 μg/ml) within one day or on three consecutive days. Recovery Absolute recovery of Allopurinol was determined in triplicate, using blank human plasma samples spiked with Allopurinol; the mean peak area was compared to that obtained from the standard drug with the same concentration. RESULTS AND DISCUSSION Study of physicochemical interaction of Allopurinol with tablet excipients DSC studies DSC thermogram of Allopurinol and the tablet excipients presents in figure (1) in which Allopurinol is characterized by a sharp endothermic peak at 386 corresponding to its melting point. Magnesium stearate and anhydrous lactose had sharp peaks at 117.08 and 245.11 C, respectively while Avicel PH101 had a broad peak at 134.1 C. On the other hand, thermogram for all physical mixtures indicate that there was no appreciable shift in the melting peak of Allopurinol with all excipients. This indicates the absence of possible interactions between the components. FTIR spectroscopy Figure (2) shows FTIR spectra for Allopurinol alone and each tablet excipient alone and Allopurinol with tablet excipients in physical mixtures. FTIR spectrum of pure Allopurinol characterized by the absorption bands
66 at 3167 cm-1 at high frequency, most probably attributed to N-H stretching band of secondary amine group, at 3034 cm-1 denoting C-H stretching vibration of pyrimidine ring. At low frequencies the band at 1693 cm-1 indicating C=O stretching vibration of the keno form of 4-hydroxy tautomer. Also the bands at (1581-1469.96) cm-1 are attributed predominantly to C-N stretching and C-C ring stretching respectively. Bands at 1234.55-698.29 cm -1 denote CH in plane deformation. An IR spectrum of Magnesium stearate alone exhibited a major bands at 3541.62 cm- 1 for O-H stretching bands of COOH acid group, also bands at 2856.83, 2640.79 and 2328.29cm-1 were for C-H aliphatic bands. While at 1541 cm -1 for C=O of carboxylic acid group. The spectra of Allopurinol and Magnesium stearate physical mixture show the same bands of both Allopurinol and magnesium stearate at the same position. Anhydrous lactose spectrum is dominated firstly by the strong primary alcohol (OH) stretching vibration showing peaks at 3466.39, 3364.16, 3300.50 and 3248.42 cm -1. These (OH) groups in lactose are hydrogen bonded to each other; free hydroxyl groups would manifest themselves at higher frequency at 3852.19, 3765.39 and 3630.36 cm -1. At low frequencies, C-O stretching present in primary and secondary alcohols respectively, R-CH2- OH and R-CH-OH-R dominates and shows strong absorption band at 1431.31 and 1089.88 respectively. The spectra of Allopurinol and anhydrous lactose physical mixture show the absorption bands of both Allopurinol and lactose at the same position. An IR spectra of Avicel PH101 which characterized by a peak at 3460.61, 3408.52, 3234.92 and 3140.40 cm -1 corresponding to the strong primary alcohol (OH) stretching vibration. These (OH) groups in Avicel PH101 are hydrogen bonded to each other; free hydroxyl groups would manifest themselves at higher frequency at 3805.9, 3759.6 and 3674.73 cm -1. Bands at 1375.37 and 1039.73 cm -1 corresponding to that of C-O stretching present in primary and secondary alcohols respectively. While bands at 2846.83, 2717.95, 2526.98, 2332.15 and 2264.63cm -1 were corresponding to C-H group. The IR spectra of the physical mixture of Allopurinol and Avicel PH101 seemed to be only the summation of drug and Avicel PH101 spectra. This result suggested that there was no interaction between drug and Avicel PH101 in the physical mixture. It was clear that all characteristic bands of Allopurinol and tablet excipients (magnesium stearate, anhydrous lactose and Avicel PH101 were appeared in the same regions and at the same ranges and there was no new bands appeared, although the shape of the functional group regions in the spectrum of the drug and the excipients used was not identical with that of pure drug alone. This might be indicative of absence of interaction between Allopurinol and excipients. Quality control of the prepared tablets Physical Properties The results of the uniformity of weight, hardness, drug content, thickness, and friability of the tablets are given in Table 2. All the samples of the test product complied with the official requirements of uniformity of weight. The drug content ranged from 95.4 to 105.4% of the label claim for Allopurinol in all formulations. The low friability values (0.023%±0.0015 to 0.33%±0.004) indicate that the matrix tablets are compact and hard. In-vitro dissolution of Allopurinol from tablets The in-vitro release of Allopurinol from tablets prepared as a solid dispersion compared with control tablets in 0.1 N HCl at 37 C ± 0.5 C are represented in Figure (3). It was found that the release of Allopurinol according to their percent mean released at 45 minutes were 102±0.23 % for FT1; 103.7±0.5% for FT2 and 15±0.7% for FT3. One way analysis of variance (ANOVA) of Allopurinol tablets with respect to their % released at 45 minute followed by Tukey- Kramer multiple comparisons test, showed significant difference at p < 0.05. Kinetic treatment for the in-vitro release of Allopurinol tablets It was found that the in-vitro release of Allopurinol followed different kinetic orders and no definite kinetic order could express the drug release from different tablet formulations (table 3). Stability study Allopurinol tablets stability study according to the calculated t 90 after one year shelf storage was as follows: FT2 Allopurinol tablets containing mannitol (drug:carrier) 1:1> FT1 Allopurinol tablets containing urea (drug:carrier) 1:1> FT3 Allopurinol tablets containing PVP K30 (drug:carrier) 1:1 with t 90 7.11, 2.29 and 1.61 year respectively. So, FT1 and FT2 were selected for the invivo study compared with commercial tablet (zyloric 100 mg). Validation of the HPLC method Optimization of chromatographic conditions Different chromatographic conditions affecting the separation process were studied and optimized. Different compositions of the mobile phase, flow rates, and wavelengths were tried. Allopurinol's peak was resolved by using a reversed-phase Nucleosil C18 column (particle size: 5 μm, 250 mm 4.6 mm), a mobile phase consisted of 2.72 gm of sodium acetate per liter distilled water adjusted to ph 4.5 with a mixture of acetic acid: acetonitrile (96:4). The mobile phase was degassed and filtered through a 0.45 µm filter (Millipore, Sainet- Quentin, Y-velines). The flow rate was 1 ml/minute. The detection wave length was 254nm. Allopurinol and salicylic acid were resolved and the retention times were 6.2 and 17.5 minutes, respectively. No
67 interfering peaks were observed in the chromatogram of the blank human plasma. Salicylic acid is a good choice as internal standard due to its similar spectral properties to Allopurinol. Linearity According to peak area-response at 254 nm, Beer's law was obeyed over the range of 0.5-8 μg/ml of Allopurinol, with a high correlation coefficient (0.9997). The equation of linear regression was Y = 0.182X 0.023. Intra-day and inter-day reproducibility of Allopurinol assay Intra- and iner-day of Allopurinol assay were calculated. The average correlation coefficient was 0.996 and 0.996, respectively as shown in table 4. These results confirmed excellent linearity of the calibration lines and high reproducibility of the assay. Absolute recovery Using the proposed HPLC method, absolute recovery of the drug was 94.5359. Bioavailability studies The mean Allopurinol plasma concentration vs. time profiles is shown in Figure 4. The mean pharmacokinetic parameters calculated from individual plasma Allopurinol concentrations vs. time profiles are summarized in Table 5. Based on statistical data, pharmacokinetic parameters of the two preparations indicated bioequivalence. The relative bioavailability of Allopurinol tablet containing urea in (drug: carrier ratio 1:1) and Allopurinol tablets containing mannitol in (drug : carrier ratio 1:1) was found to be 118 and 54 % respectively. Figure 1. DSC thermograms of allopurinol and various tablet excipients Allo, allopurinol; PM, physical mixture; Mag. St., Magnesium stearate and Anhy. Lact. Anhydrous lactose.
68 Figure 2. FTIR spectrum of Allo, various tablet excipients and the physical mixtures Allo, Allopurinol; PM, physical mixture; Mag.St., Magnesium stearate and Anhy.Lact., Anhydrous lactose Figure 3. Percent drug released from tablets formulation compared to plain drug (c) and commercial tablet (Zyloric 100) Figure 4. Allopurinol mean plasma concentration-time curve after oral administration
69 Table 1. Composition of Allopurinol tablets Components (mg) Tablet code SD tecnique Allo. Urea Mannitol PVP Avicel Anhydrous Magnesium K30 PH101 lactose stearate FT1 melting 100 100 ------ --------- 30 67 3 FT2 melting 100 ----- 100 -------- 30 67 3 FT3 Solvent 100 ------ ---------- 100 evaporation 30 67 3 Control (C) 100 ------ --------- ----------- 15 39 1.5 Table 2. Quality control data of Allopurinol tablets Formula Quality control tests FT1 FT2 FT3 Weight variation (mg) ± S.D. 297.80 ±1.06 299.13 ±1.29 299.03 ± 2.28 Thickness (mm) ± S.D. 3.6811±0.01 3.528 ± 0.02 3.73 ±0.01 Hardness (Kg) ± S.D. 5.2 ± 0.141 4.793 ± 0.147 4.99 ± 0.43 Friability (% Loss) ± S.D. 0.023 ± 0.015 0.333 ± 0.004 0.07 ± 0.0141 Disintegration time (minutes) ± S.D. 9.66 ± 0.288 2.5 ± 0.502 50.00 ± 1.414 Drug content (%) ± S.D. 96.224 ± 0.89 95.63 ± 0.945 102.42 ± 1.55 Table 3. Kinetic treatments for the in-vitro release of Allopurinol tablets Correlation coefficient (r) Formula A Higuchi-diffusion Hixson-Crowel Baker-lonsdale Zero-order First-order Second-order model cube root law equation FT1 0.947-0.950 0.9176 0.979 0.996 0.994 FT2 0.853-0.986 0.917 0.9103 0.982 0.981 FT3 0.966-0.696 0.624 0.923 0.817 0.743 Z1 0.837-0.997 0.919 0.896 0.963 0.919 Z1(Zyloric 100mg Galaxo-Wellcome) Table 4. Intra-day and inter-day reproducibility of Allopurinol in human plasma by high-performance liquid chromatography Peak area ratio Spiked Conc. (μg/ml) Intra-day Inter-day Mean ± S.D Mean ± S.D 0.00 0.00 ± 0.00 0.00 ± 0.00 0.5 0.077 ± 0.07 0.077 ± 0.005 1 0.150 ± 0.15 0.153 ± 0.003 2 0.338 ± 0.33 0.348 ± 0.019 4 0.740 ± 0.74 0.732 ± 0.017 6 1.0008 ± 0.04 1.043 ± 0.04 8 1.459 ± 0.10 1.513 ± 0.02 Slope 0.177 ± 0.01 0.188 ± 0.003 Correlation coefficient(r) 0.9964 ± 0.004 0.996 ± 0.001 S.D. = Standard deviation
70 Table 5. Pharmacokinetic parameters of different Allopurinol treatments administered orally to human volunteers Pharmacokinetic parameters Volunteers orally administered TF1 TF2 Commercial tablets C max (μg/ml) 1.883 1.752 1.467 t max (hr) 1 1.208 1 t ½ ab (hr) -1.558-0.979-2.121 t ½ el (hr) 1.096 0.584 1.474 K ab (hr -1 ) -0.445-0.708-0.327 K el (hr -1 ) 0.666 1.247 0.475 AUC 0-8 (μg.hr/ml) 3.976 1.874 1 AUC 0- (μg.hr/ml) 4.463 2.040 1.467 RB % 118 54 ------- CONCLUSION Allopurinol tablet (FT1) which contains urea in (drug : carrier ratio 1:1) at dose of 100 mg has a best relative bioavailability, highest Cmax and AUC 0- and reasonable k ab and k el. ACKNOWLEDGMENTS The authors would like to thank Alexandria Company for pharmaceutical industries, (Alexandria, Egypt) for their donation of Allopurinol and Amoun Company for pharmaceutical industries, (Cairo, Egypt) for providing the other used polymers. REFERENCES Abd-Elazeem M, Khattab I, samy E, Tous S, Joachim J and Reynier J. Improvement availability of Allopurinol from pharmaceutical dosage forms: II- capsules and tablets. Az. J. Pharm. Sci. 2001; 27: 387-399. Aggarwal S, Gupta G D and Chaudhary S. Solid dispersion as an eminent strategic approach in solubility enhancement of poorly soluble drugs, I.J.P.S.R 2010, 1(8). Ammar HO and El-Nahhas SA. Improvement of some pharmaceutical properties of drugs by cyclodextrin complexation. Pharmazie. 1995; 50: 49-50. Benzra SA and Bennett TR. (1978) in Analytical profiles of drug substances and excipients, Academic press, Inc, 7, pp.1-18. Clark s analysis of drugs and poisons in pharmaceutical body fluids and postmartum materials (2004), 3 rd Ed., Pharmaceutical press, London, pp. 601-603. Day RO, Grham GG, Hicks M, Mclchlan AJ, Stocker SL and Williams KM. Clinical pharmacokinetics and pharmacodynamics of Allopurinol. Clin. Pharmacokinet. 2007; 46: 623-644. Derek K and Da-peng wang TL. Formulation development of Allopurinol suppositories and injectables. Drug Dev Ind Pharm. 1999; 25: 1205-1208. Devi VK, Vijayalakhmi P and Avinash M. Preformulation studies on celecoxib with a view to improve bioavailability. Indian J Pharm Sci. 2003; 65:542-545. Gibaldi M, Perrier D. Pharmacokinetics. New York, NY: Marcel Dekker; 1990. Goldberg AH, Galbaldi M and Kanig KL. Increasing dissolution rates and gastrointestinal absorption of drugs via solid solutions and eutectic mixtures III. Experimental evaluation of griseofulvin-succinic acid solution. J Pharm Sci. 1966; 55:487-492. Hamza YE and Kata M. Influence of certain non-ionic surfactants on solubiliziton and in-vitro availability of Allopurinol. Pharm Ind 1989; 51: 1441-1444. Jagdale SC, Kuchekar BS, Chabukswar AR, Musale VP and Jadhao MA. Preparation and in vitro evaluation of Allopurinol- Gelucire 50/13 solid dispersions. Int J. Advances in Pharm. Sci. 2010; 1: 60-67. Kramer, W.G. and Feldman, S. High performance liquid chromatographic assay for allopurinol and oxipurinol in human plasma, J. Chromatogr. 1979; 162, 94-97 Reinders, M.K.; Nijdam, L.C.; VanRoon, E.N.; Movig, K.L.L.; Jansen, T.L.Th.A.; Van de Laar, M. A.F.J. and Brouwers, J.R.B.J. A simple method for quantification of Alloprinol and oxipurinol in human serum by high performance liquid chromatograghy with UV-detection, J. Pharm. Biomed. Anal. 2007; 45, 312-317. Samy AM, Marzouk MA, Ammar AA, Ahmed MK. Enhancement of the dissolution profile of allopurinol by a solid dispersion technique. Drug Discov Ther. 2010; 4(2):77-84.
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