Preformulation Studies of Nifedipine

ISSN 0975-2366 DOI:https://doi.org/10.31838/ijpr/2020.12.03.011 Research Article Preformulation Studies of Nifedipine J. SRINIVASARAO1, PROF. C. GOPINATH2 SM pharmaceuticals Sdn. Bhd.; Sungai petani; Malaysia JNTUA OTRI; Anantapur; Andhra Pradesh; India. Email: srinivasaraoj14@gmail.com Received: 24.01.20, Revised: 09.02.20, Accepted: 21.03.20 ABSTRACT The preformulation studies deals with physical; chemical properties of Nifedipine for the generation of bi-layer heterogeneous tablets formulation development studies. Solubility studies for saturated solutions were carried out over the pH range of 1.2-6.8. The solubility of Nifedipine in water at 37°C was 436.3mg/L. Further the solubility is not affected significantly (P < 0.05) in the buffer solutions in a pH range of 1.2 to 6.8. The PSD histograms represents the presence of Dv (50) 7.18µm; Dv (90) 19.3µm µm particles of Nifedipine. From the p-XRD studies; the diffraction line profiles are 2θ values for diffraction peaks at 11.7504°, 11.9178°, 12.9763° confirms to form D for Nifedipine. By the UV-Visible; FTIR; NMR and Mass spectroscopy studies Characterized to Nifedipine. The DSC thermo gram conforms to of Nifedipine at 172.77°C. The Assay content results of three APIs were within 99%101%. The Nifedipine contains nitro phenyl pyridine, nitroso phenyl pyridine analogues, and methyl 3-amino but-2enoate is <0.1% with >99.9% purity and all other unknown impurities were not detected in the drug substance. The compatibility of binary mixtures of drug and excipients, stored at 40°C ± 2°C/ 75% ± 5% RH for 1 month was assessed and the % results of Assay and impurities from initial period to after 30th day not have significant difference; P<0.01. The preformulation results conforms, the Excipients PVP K-30; Croscarmellose Sodium; HPMC K-15/E-5, Carbomer 974p, Ethyl cellulose, Xanthan gum, ZnO, and SiO2 were compatible with Nifedipine and selected for development of control release bilayer tablets. Key words: Preformulation studies; DSC studies; NMR Studies; FTIR Studies; p-XRD studies INTRODUCTION The oral route is the most preferred, accepted, desirable route for administrating therapeutically active agents for systemic effects over the different routes of administration are available for the delivery of drugs as it is a convenient, safe, good patient compliance and adaptable to accommodate more drugs. Even for sustained release formulations have been investigated for oral route of administration because of flexibility in designing dosage forms. The Nifedipine was selected taking into account of their physicochemical, and biopharmaceutical properties. all recent clinical guidelines still considered ACE inhibitors, Thiazides, Calcium channel antagonists and are as primary treatment for organization of hypertension [1]. The combination drug therapy at lower dose is recommended to permit medications of different mechanism of action, to complement each other and mutually effectively lower the blood pressure than the highest doses of the individual [2]. The primary intention of release drug delivery is to make sure safety and to improve efficacy of drugs in addition to patient compliance [3]. Nifedipine is dihydropyridine chemical class selective Calcium channel blocker, chemically defined as Dimethyl 2,6-dimethyl-4-(2-nitrophenyl)-1,4[4] dihydropyridine-3,5-dicarboxylate . Peak plasma concentration is achieved after 6 hours. Linear pharmacokinetic profile is over the dose of 30-180mg [5]. It is bio transformed in the liver by cytochrome P450 system. 60 to 80% of absorbed drug is excreted in urine as highly water-soluble dormant metabolites and 5-15% is in biliary emission. The Elimination half-life is 7hours [6]. Figure 1: Typical Chemical structure of Nifedipine MATERIALS The active pharmaceutical ingredient Nifedipine BP procured from Unique Chemicals, Gujarat, 71 | International Journal of Pharmaceutical Research | Jul - Sep 2020 | Vol 12 | Issue 3 Preformulation Studies of Nifedipine India; The Excipients namely Polyvinylpyrrolidone K-30 BP; Croscarmellose Sodium BP; Hydroxy propyl methyl cellulose K-15 and E-5 BP, Carbomer 974P BP, Ethyl cellulose BP, Xanthan gum BP, Zinc oxide BP, and colloidal silicon di oxide were donated by SM Pharmaceuticals SDN BHD, Malaysia. compatibility studies were carried out. The excipients used were; Hydroxy propyl methyl cellulose K-15 and E-5, Carbomer 974p, Ethyl cellulose, Polyvinylpyrrolidone K-30; Croscarmellose Sodium; Xanthan gum, Zinc oxide, colloidal silicon di oxide Formic acid, Hydrochloric acid, Sodium Hydroxide, Potassium di-hydrogen phosphate, Di Potassium hydrogen phosphate, Sodium dihydrogen phosphate, Sodium Lauryl sulphate analytical grade chemicals and Methanol, Acetonitrile HPLC Grade solvents were purchased from Merck, Mumbai. Solubility studies A significant Physical-chemical property of a drug candidate is solubility, particularly aqueous solubility. A drug should have aqueous solubility for therapeutic effectiveness in the PH range of 1 to 8. The drug must be in solution form to enter into systemic circulation, to exert therapeutic effect. The solubility studies were conducted by using UV –Visible Spectrophotometer; UV1900; Shimadzu. The solubility of Nifedipine, Enalapril maleate, and Hydrochlorothiazide were determined in distilled water; 0.1N hydrochloric acid PH1.2; 0.1N hydrochloric acid PH1.2 with 1% w/v sodium lauryl sulfate; acetate buffer pH 3.0, 4.0, and phosphate buffers pH 5.0 and 6.8 phosphate buffer. The PH of all media were verified by using pH meter; 827 pH Lab; Metrohm. Precisely measured volume of each solvent was poured in screw-capped glass vials followed by addition of excess drug. The glass vials were sealed and positioned on a mechanical shaker at 37 °C for 24 hours. Thereafter aliquots were withdrawn, centrifuged, and filtered. The filtrates were diluted and absorbance was recorded using UV–Visible Spectrophotometer. The reference materials Nifedipine BPCRS # 2997 was procured from British Pharmacopoeia. The impurity standards of Nifedipine impurity A CRS # 1(nitro phenyl pyridine analogue), Nifedipine impurity B CRS # 2(nitroso phenyl pyridine analogue) for peak identification were procured from European Pharmacopoeia. EQUIPMENT / INSTRUMENTS The sophisticated instruments HPLC; Model: 2689; Make: Waters; UPLC; Model: H-Class; Make: Waters; UV –Visible Spectrophotometer; Model: UV 2600; Make: Shimadzu; Semi Micro Balance; Model: GR202; Make: AND Company Limited; Analytical Balance; Model: AL 204; Make: Mettler Toledo; Stability chamber; Model: TH 400 S/G; Make: Thermolab; Differential Scanning Calorimeter; Model: Q20; Make: TA Instruments; Particle size analyser; Model: Mastersizer 3000; Make: Malvern Panalytical Ltd; FTIR spectrometer; Model:Alpha; Make: Bruker; Nuclear Magnetic Resonance Spectrometer 500MHz; Model: ECZ400S/L1; Make: JEOL; Xray diffractometer (p-XRD); Model: Aeris; Make: Malvern Panalytical Ltd; pH meter; Model: 827 pH Lab; Make: Metrohm; Magnetic Stirrer; Model: MS7-H550-S; Make: DLab; LC-MS; Model: SQD2; Make: Waters; UPLC-MS/MS; Model: Xevo TQS; Make: Waters; Burrel Scientiphic wrist action laboratory shaker; Model: 75; Make: Fisher; Halogen Moisture Analyzer Model: HS153; Make: Mettler Toledo. METHODS The Physico-Chemical properties of drug substances have a great impact on the selection of controlled release formulation and manufacturing process. The physico-chemical characteristics of Nifedipine, and Drug and individual Excipient Particle size distribution Particle size considerably affects several quality parameters such as solubility, lack of grittiness, content uniformity, rate of dissolution, and ultimately bioavailability. Various methods to determine the particle size comprise cascade impaction, laser holography, microscopy, sieving, sedimentation rate, light energy diffraction. Particle size distribution studies of Nifedipine was carried out on a particle size analyser; Master sizer 3000; Malvern. Laser diffraction technique was employed to measure size distribution of powder particles in a dry dispersion apparatus. Based on distribution data, the particle size of powder was defined by various means like D (3, 2) D (4, 3), and the maximum particle size was measured by percentiles as Dv10, Dv50, Dv90. 72 | International Journal of Pharmaceutical Research | Jul - Sep 2020 | Vol 12 | Issue 3 Preformulation Studies of Nifedipine Crystallinity and Polymorphism Crystallinity and polymorphism are one of the important features of preformulation studies. Crystals and polymorphs are characterized by microscopy, thermal analysis and X-ray diffraction techniques. Importance of identification of shape of crystals and its internal structure can affect the solubility and stability. Powder X-ray diffraction (pXRD) technique is a powerful nondestructive technique for recognition of polymorphic forms of sample and provides specific pattern for a given form. The pattern appeared as a fingerprint of phase and is represented as a plot of relative or absolute intensity verses the angular parameter 2θ. The P-XRD pattern is a record of diffracted intensity of a crystalline sample in one dimensional as a function of diffraction angle. The diffraction studies of Nifedipine, Enalapril maleate and Hydrochlorothiazide, each containing 500mg, were conducted on an X-ray diffractometer; Aeris; Malvern Panalytical.Goniometer was used to maintain the angle and rotate the sample p- XRD patterns were recorded using following scan parameters: A. Target material Cu B. Voltage 40 Kv C. Current 15 mA. D. Angular parameter 2θ 2º to 50º at a step size of 0.022º E. Length of 2θ 3.02 F. Scan time 36s G. Specimen length 10mm H. Temperature 25ºC I. CuKαα radiation 1.5418Å. J. K-β 1.392, K. K-α2/K-α1 0.5 CHARACTERIZATION UV-Visible Spectroscopy The UV spectra of APIs were scanned using a UV spectrophotometer; UV-2600; Shimadzu. Standard stock solution of Nifedipine was prepared individually by dissolving 30mg in 100 ml methanol. 30ppm standard solution was prepared by suitable dilution of stock solution with the same solvent and scanned in the range of 200-400nm to determine the wavelength of maximum absorbance. Fourier Transformed-Infrared (FT-IR) Spectroscopy Infrared spectra covering the region of 4000 to 650cm-1 were conducted for Nifedipine pure drug using FTIR spectrometer; Alpha; Bruker with ATR accessory. Obtained spectra were the average of 16 scans at a resolution of 4 cm−1 Nuclear Magnetic Resonance (NMR) Spectroscopy 1 H and 13C NMR spectra were run on NMR instrument 500MHz; ECZ400S/L1; JEOL equipped with 1H and 13C operating at 500 MHZ using tetra methyl silane (TMS) as an internal standard. About 10 mg and 30 mg of Nifedipine pure drug for H1 NMR and C13 NMR were taken individually into a clean NMR tube and diluted with the deuterated di methyl sulfoxide (DMSOd6) and the spectra were scanned by using below instrument parameters as given in the Table 1. Table 1: Nuclear Magnetic Resonance (NMR) spectroscopy Conditions H1- NMR instrument Parameters C13- NMR instrument Parameters X-Domain Proton X-Points 16384 Offset 7ppm X-Sweep 18ppm X-Pre scans 1 Solvent DMSO-D6 BF or S. Exp 0.30 Hz Relaxation delay 2 sec Pulse 45 Degree Scans 16 X-Domain Carbon-13 X-Points 32768 X-Offset 100 ppm X-Sweep 250 ppm X-Pre scans 4 Solvent DMSO-D6 BF or S. Exp 2.0 Hz Relaxation delay 2 sec Pulse 30 Degree Liquid chromatography–Mass Spectrometry (LC-MS) Mass analysis and detection for pure drug Nifedipine was carried out using LC-MS system; SQD-2; Waters equipped with electron spray ionization operated in combined mode. The mobile phase used was 5mM ammonium acetate in water and acetonitrile in a ratio of 20:80 in Isocratic method at a flow rate of 0.2ml/min. Run time was 3min. The optimized source dependent conditions were as follows. Desolvation gas flow- 1000 L/hr • • • • Cone gas flow - 25L/hr Capillary Voltages - 3.5 kV Cone Voltage- 50 V Desolvation temperature - 350°C 73 | International Journal of Pharmaceutical Research | Jul - Sep 2020 | Vol 12 | Issue 3 Preformulation Studies of Nifedipine Thermal Analysis by Differential Scanning Calorimetric Thermal properties of pure drugs were analyzed by using differential scanning calorimeter; TA Instruments; Q-20 V24.11 Build 124. 2.08mg of the sample was weighed and placed on the thematically sealed pan; an empty pan was used for blank. Now keep both the pans on respective slots. Then samples were heated by maintaining a temperature range of 20 to 350°C at a ramped temperature of 10 °C / min, using nitrogen at a flow rate of 50 mL/min for maintaining the inert atmosphere. Quantitative Assay Content The content of Nifedipine was determined by UPLC H-Class with PDA detector; Make: Waters using Acquity UPLC BEH C18; 1.7µm; 2.1mm I.D x 50mm Length (Make: Waters) previously stabilized at 40ᴼC with 0.5mL per minute flow rate of 0.1%v/v formic acid in water as buffer; a mixture of Methanol and acetonitrile in a ratio of 10:90 v/v from the gradient table 2. The standard and sample solution was prepared with a concentration of 30µg/mL of Nifedipine in methanol followed by make up the dilutions with aqueous buffer containing 0.1% v/v formic acid. The chromatography is carried with 1.0µL injection volume; detection of analytes was measured at 254nm. Table 2: Typical liquid chromatography gradient program Time Line A Line B (minutes) (%buffer v/v) (%Solventv/v) 0.0 95 5 0.5 95 5 1.2 70 30 2.0 50 50 3.0 95 5 4.0 95 5 Quantitative Estimation of Related Substance The impurities related to Nifedipine were quantified by using UPLC H-Class equipped with Tandem mass spectrometer detector (MS/MS); Waters using Acquity UPLC BEH C18; 1.7µm; 2.1mm I.D x 50mm Length; Waters previously stabilized at 40ᴼC with 0.4mL per minute flow rate of 0.1%v/v formic acid in water as buffer; Methanol and acetonitrile in a ratio of 10:90 v/v from the gradient table 4 with mass spectrometric conditions as given in the table 3. Table 3: Typical MS conditions for related substance quantification Parameter Setting Mode ESI + Capillary voltage 2.5 kV Cone voltage 20 V Collision Energy Ramp (MSMS experiment) 15 – 45 V Source temperature 120 C Desolvation Temperature 350 C Desolvation Gas Flow 650 L hr o o Table 4: Typical gradient elution program for related substance quantification Time (min) Flow (mL/min) %A %B Curve 0 0.4 98 2 - 0.5 0.4 98 2 6 7 0.4 60 40 6 8 0.4 98 2 11 10 0.4 98 2 11 Drug- Excipients Compatibility Studies This is an important feature of pre formulation studies used to evaluate the impact of excipients on physical and chemical stability, dissolution and ultimately bioavailability of the formulation. FT-IR Spectroscopy, DSC studies for the physical mixture, and Accelerated Stability studies for binary mixture of pure drug and individual excipients were performed to assess the compatibility of excipients with Nifedipine. The blend was mixed and filled in Type I glass vials (open vials & vials sealed with rubber stopper) and were stored in Stability chamber; Model: TH 400 S/G (Make: thermo lab) at 40ºC + 2 ºC / 75% R.H for 1 month. The stored compatibility mixtures were tested individually by UPLC, HPLC analysis to assess the drug substance purity and impurity profile. RESULTS AND DISCUSSION Solubility studies Solubility studies for saturated solutions were carried out over the pH range of 1.2-6.8. The 74 | International Journal of Pharmaceutical Research | Jul - Sep 2020 | Vol 12 | Issue 3 Preformulation Studies of Nifedipine data of the solubility studies are shown in table 5. The solubility of Nifedipine in water at 37°C is 436.3mg/L. Further the solubility is not affected significantly (P > 0.05) in the buffer solutions in a pH range of 1.2 to 6.8. From the table 5, it is seen that Nifedipine is having limited solubility < 450mg/Lower the entire pH range studied. According to the Biopharmaceutical Classification System (BCS), Nifedipine is a Class II compound with low solubility and high permeability. The absorbance of Nifedipine in 0.1N Hydrochloric acid and 1% Sodium lauryl sulfate media is in between 0.4212 to 0.9664 abs from Figure 2 solubility graphs and these absorbance values are significantly greater than from all other media. The representing solubility graphs are shown in Figure 2. Table 5: Solubility data of Nifedipine Sr. No 1 2 3 3 4 5 6 7 Name of Solubility Media Water 0.1N HCl, pH 1.2 0.1NHCl, pH 1.2 + 1% SLS Acetate buffer, pH 3.0 Acetate buffer, pH 4.0 Phosphate buffer, pH 5.0 Phosphate buffer, pH 6.8 0.1N Sodium hydroxide 1.2000 Nifedipine Solubility (mg/L) 436.3 462.2 471.4 456.3 453.1 451.6 447.3 434.5 Water 1.0000 0.1N HCl; pH 1.2 0.8000 0.6000 0.4000 0.2000 0.0000 0.00 100.00 200.00 300.00 400.00 500.00 0.1N HCl, pH 1.2 + 1%SLS Acetate buffer pH 3.0 Acetate buffer pH 4.0 Concentration (mg/L) Figure 2: Typical Solubility graph for Nifedipine Particle size distribution study The study was carried out by using laser diffraction method on Master sizer 3000 particle size analyzer. The Nifedipine PSD histogram represented in the Figure 3 conforms the presence of Dv (10) 1.67µm; Dv (50) 7.18µm; Dv (90) 19.3µm and Dv (95) 23.9 µm particles in crystalline power by dry dispersion technique. 75 | International Journal of Pharmaceutical Research | Jul - Sep 2020 | Vol 12 | Issue 3 Preformulation Studies of Nifedipine Figure 3: Typical PSD Histogram for Nifedipine X-Ray diffraction studies The main characteristics of diffraction line profiles are 2θ position, peak height, peak area, and shape (characterized by, e.g., peak width, or asymmetry, analytical function, and empirical representation). A crystalline structure of Nifedipine implies that the structural units (i.e. the unit cells) are repeated in a long-range order. The atoms and/or molecules do not possess a distinguishable crystal lattice. From the diffract graph represented in Figure 4, the diffraction line profiles are 2θ values for diffraction peaks are at 8.0627°, 10.4088°, 11.7504°, 11.9178°, 12.9763°, 13.3525°, 14.6803°,16.1915° confirms to form D solid phase. Figure 4: Pattern of X- ray Diffraction of Nifedipine 76 | International Journal of Pharmaceutical Research | Jul - Sep 2020 | Vol 12 | Issue 3 Preformulation Studies of Nifedipine CHARACTERIZATION UV-Visible Spectroscopy The UV spectra of APIs were scanned by using the Shimadzu UV-2600 instrument in a range of 200nm to 400nm. The 30ppm of Nifedipine standard solution is prepared in methanol. The ƛmax values observed at 235 nm and at 329nm from the figure 5. Figure 5: Typical UV Spectrum for Nifedipine Fourier Transformed Infrared Spectroscopy The FT-IR spectrum of Nifedipine pure drug from the Figure 6 shows characteristic peaks which consist of a broad peak at 3332 cm-1 indicating the >N-H stretching ( pyridine moiety), the absorption peaks at 1529cm-1 is due to (N-O) asymmetric stretching, and at 1349 cm-1 is due to (N-O) symmetric stretching ( Aryl- nitro), at 1680 cm-1 is due to >C=O stretch (ester group), at 1227 cm-1 is due to C-O stretch, at 2953cm-1, 2996 cm-1 are due to C–H stretching and absorption at 1623 cm-1 representing ring breathing band (pyridine). Figure 6: Typical FTIR Spectrum for Nifedipine Nuclear Magnetic Resonance Spectroscopy Chemical shift values are reported on the δ scale in ppm relative to TMS (δ 0.00) as internal standard. The 1H NMR spectrum of compound from figure: 7 shows chemical shift values in the region of 7.68 to 7.24ppm corresponding to protons at C-3, C-5, C-4, and C-6 of phenyl group. The Prominent signals at 5.66ppm ascribed to -CH proton and at 5.72ppm to -NH proton of the dihydropyridine ring. The singlet observed at 2.34ppm represents methyl group at C-2, and C-6 on the dihydropyridine ring. The spectrum also displays a singlet at 3.59ppm attributable to methoxy group at C-3, and C-5 of the dihydropyridine ring. Chemical shifts values assigned to various protons are listed in Table: 7. 77 | International Journal of Pharmaceutical Research | Jul - Sep 2020 | Vol 12 | Issue 3 Preformulation Studies of Nifedipine The 13CNMR spectrum from figure 8, reveals chemical shift values in between 123.8ppm to 132.7ppm assigned to carbons of phenyl group. The signal at 34.5ppm attributed to C-4 carbon. The signal at 51.0ppm was assigned to two methoxy group. The chemical shift value at 19.5ppm ascribed to two methyl group. The signal observed at 103.7ppm was assigned to quaternary carbons of C-3 andC-5. The chemical shift values at 142.0 and144.7 ppm was assigned to quaternary carbons of C-2 and C-6. The signal at 167.5ppm assigned to carbon atoms of two keto group. Chemical shifts values assigned to various carbons are listed in Table: 8. Finally, H1/C13–NMR spectra confirms to Nifedipine Chemical Structure. Table 7: 1HNMR chemical shift values for Nifedipine Position 2 and 6 3 and 5 1 4 3’ of phenyl 5 ‘of phenyl 4 ‘of phenyl 6’ of phenyl Group CH3 OCH3 NH CH CH CH CH CH δ H(ppm) 2.34 (s, 6H) 3.59(s, 6H) 5.72(s, 1H) 5.66(s, 1H) 7.68 (dd, 1H, J=8.3 Hz, 1.4Hz) 7.51 (dd, 1H, J=8.3 Hz, 1.4Hz) 7.45 (td, 1H, J=7.6 Hz, 1.4Hz) 7.24(m,1H) Table 8: 13CNMR chemical shift values for Nifedipine Position C- 2 and C-6 C- 4 C-3 and C-5 C-3 and C-5 C-3 ‘ C-4 ‘ C-5 ‘ C-1 ‘ C- 2 C- 6 C-3 and C-5 Type of Carbon CH3 CH OCH3 qC CH CH CH C qC qC C=O δ C (ppm) 19.5 34.5 51.0 103.7 123.8 127.0 131.0 132.7 142.0 144.7 167.5 78 | International Journal of Pharmaceutical Research | Jul - Sep 2020 | Vol 12 | Issue 3 Preformulation Studies of Nifedipine Figure 7: 1HNMR spectrum for Nifedipine Figure 8: 13CNMR spectrum for Nifedipine Mass Spectrometry The molecular ion provides the molecular mass of the analyte. The molecular ion masses for Nifedipine was predicted in ES combined mode. The molecular ion (M-1) -detected at m/z 345.2 in negative ionization mode, based on this, conformed that the molecular weight of Nifedipine is 346.3 as in the mass spectrum shown in Figure 9. 79 | International Journal of Pharmaceutical Research | Jul - Sep 2020 | Vol 12 | Issue 3 Preformulation Studies of Nifedipine Figure 9: Typical Mass spectrum for Nifedipine Thermal Analysis by Differential Scanning Calorimetric The differential scanning calorimeter (DSC) is a fundamental tool in thermal analysis. Melting is an endothermic process, requiring the absorption of heat. The temperature remains constant during melting despite continued heating from 20 to 350°C with a rate of 10°C/min. The DSC thermo gram of Nifedipine from the Figure 10, the peak was observed at 172.77°C and the onset value is 171.27°C represents the characteristic endothermic peak of Nifedipine and energy required for melting is 111.2J/g. Figure 10: Typical DSC graph for Nifedipine 80 | International Journal of Pharmaceutical Research | Jul - Sep 2020 | Vol 12 | Issue 3 Preformulation Studies of Nifedipine Assay Content and Related Substances The purity of Nifedipine by UPLC is >99.8%. The % of impurities A and B and other impurities by UPLC-MS/MS are <0.1% and <0.01% respectively and results are shown in Table 9. Drug-Excipients Compatibility studies To meet the target product profile, tablet excipients with appropriate functionality were assessed. Based on scientific and prior knowledge, the chosen excipients had been used successfully for a roller-compacted formulation. The FT-IR spectra of the Nifedipine and excipients were overlaid to identify probable chemical interaction between them. The FT- IR spectra of HPMC E5 and HPMC E15 show stretching vibrations of the hydroxyl group at 3445cm-1 and 3455cm-1. Both the polymers show bending vibrations of hydroxyl group at same wavelength that is 1644 cm-1, as well as aliphatic stretching of C-H appear at 2905cm-1. Characteristic absorption bands of C-O in C-O-C group of glucose molecules of HPMC E5 and E15 observed at 1059cm-1 and 1060cm-1 and both polymers show at C–H bending at 1337 cm–1. The spectrum of ethyl cellulose shows characteristic absorption bands for C-O stretching vibration in the C-O-C linkage of glycoside bonds is at 1052 cm–1 and C–H stretching bands at 2890 cm–1 and 2980 cm–1. The absorption at 1369 cm–1 corresponds to C–H bending. The FT IR spectrum of Xanthan gum exhibits a prominent absorption band at 3400 cm-1 is due to OH groups interacting with a water molecule, the absorption peak at 1692cm-1 is ascribed to C=O stretching of esters, and at 1615cm-1 is for COOstretching vibration. The absorption band at 1052cm-1 corresponds to the C-O stretching of CO-C glycoside linkage bonds. Compared to the absorption bands from each component, it was observed that the FT-IR spectrum of Nifedipine showed a superposition of peaks from the excipients, indicated that there were no intermolecular interactions between them from Figure 11. The thermal profile of Xanthan gum discloses a melting point in between 90-120ºC, the endothermic peak observed for Xanthan gum at 100.2ºC. While the drug melting peak is quite visible in the physical mixture of Nifedipine and Xanthan gum (1:1) thermogram at 173.5ºC, Therefore the thermograms of the physical mixture showed that no interaction between Nifedipine and Xanthan gum. The thermograms of pure Nifedipine and Xanthan gum /Nifedipine mixture are shown in the Figure 12 In the physical mixture (1:1) thermogram, the drug melting peak still appears at 173.5ºC, indicating that there was no physiochemical interaction between Nifedipine and HPMC. The thermograms of pure Nifedipine and Nifedipine /HPMC mixture are shown in the Figure 13. The DSC thermogram of pure Nifedipine shows a sharp endothermic peak at 172.7ºC and a similar endothermic peak was observed at 173.5ºC for the controlled release granules of Nifedipine indicated that all selected excipients were compatible with Nifedipine. The Assay result from Table 9 for Nifedipine is greater than 99% and the relative standard deviation is <1.0% from the initial results to 30thday results. The calculated confidence interval at 95% level from descriptive statistics is 0.21, represents no significant change in the assay results from the initial. The related impurities Nifedipine nitrophenyl pyridine analogue (impurity A), Nifedipine Nitroso phenyl pyridine analogue (impurity B) were <0.1% from entire study intervals. 81 | International Journal of Pharmaceutical Research | Jul - Sep 2020 | Vol 12 | Issue 3 Preformulation Studies of Nifedipine Figure 11: Overlay FT-IR spectra for Nifedipine and Excipient Figure 12: DSC thermogram for Nifedipine and Nifedipine and Xanthan gum mixture 82 | International Journal of Pharmaceutical Research | Jul - Sep 2020 | Vol 12 | Issue 3 Preformulation Studies of Nifedipine Figure 13: DSC thermogram for Nifedipine and Nifedipine and HPMC mixture Table 6: Compatibility results of Nifedipine API and Nifedipine with Excipients Mixture Mixture Name Nifedipine API Nifedipine + HPMC K-15 Nifedipine + Carbomer 934P Nifedipine + HPMC E-5 Nifedipine + Colloidal silicon dioxide Nifedipine + Ethyl cellulose Nifedipine + PVP K-30 Nifedipine + Condition at 40°C/75%RH Time period Initial (0 Hour) 7th Day 15th Day 30th Day Initial (0 Hour) 7th Day 15th Day 30th Day Initial (0 Hour) 7th Day 15th Day 30th Day Initial (0 Hour) 7th Day 15th Day 30th Day Initial (0 Hour) 7th Day 15th Day 30th Day Initial (0 Hour) 7th Day 15th Day 30th Day Initial (0 Hour) 7th Day 15th Day 30th Day Initial (0 Hour) 7th Day Assay (%) 99.8 99.7 99.9 99.6 99.7 99.8 99.6 99.8 99.6 99.5 99.8 99.7 99.4 99.6 99.7 99.5 99.6 99.8 99.4 99.6 99.7 99.8 99.4 99.5 99.6 99.5 99.8 99.7 99.7 99.8 A (%) 0.03 0.03 0.04 0.06 0.04 0.03 0.05 0.04 0.06 0.05 0.05 0.06 0.04 0.03 0.06 0.05 0.06 0.04 0.05 0.06 0.06 0.05 0.07 0.07 0.04 0.06 0.06 0.07 0.06 0.05 % Impurities B Any other (%) impurity (%) 0.04 BDL (<0.01%) 0.05 BDL (<0.01%) 0.06 BDL (<0.01%) 0.07 BDL (<0.01%) 0.05 BDL (<0.01%) 0.04 BDL (<0.01%) 0.07 BDL (<0.01%) 0.06 BDL (<0.01%) 0.06 BDL (<0.01%) 0.07 BDL (<0.01%) 0.07 BDL (<0.01%) 0.05 BDL (<0.01%) 0.05 BDL (<0.01%) 0.06 BDL (<0.01%) 0.04 BDL (<0.01%) 0.07 BDL (<0.01%) 0.06 BDL (<0.01%) 0.05 BDL (<0.01%) 0.06 BDL (<0.01%) 0.07 BDL (<0.01%) 0.08 BDL (<0.01%) 0.04 BDL (<0.01%) 0.04 BDL (<0.01%) 0.05 BDL (<0.01%) 0.03 BDL (<0.01%) 0.07 BDL (<0.01%) 0.05 BDL (<0.01%) 0.04 BDL (<0.01%) 0.07 BDL (<0.01%) 0.04 BDL (<0.01%) 83 | International Journal of Pharmaceutical Research | Jul - Sep 2020 | Vol 12 | Issue 3 Preformulation Studies of Nifedipine Crosscormellose sodium Nifedipine + Xantham Gum 15th Day 30th Day Initial (0 Hour) 7th Day 15th Day 30th Day Nifedipine + Initial (0 Hour) red ferric oxide 7th Day 15th Day 30th Day Nifedipine + Initial (0 Hour) Zinc oxide 7th Day 15th Day 30th Day *BDL: Below detection limit CONCLUSION Pre formulation play a significant part in the selection of drug candidate, formulation components, and drug product manufacturing process. Pre formulation studies give directions for design of formulation in choice of drug form, excipients and composition. Physical structure helps in adjustment of biopharmaceutical and pharmacokinetic properties. Pre formulation studies on Nifedipine have been performed. The water solubility of nifedipine is 436.3mg/L. According to the Biopharmaceutical Classification System (BCS), Nifedipine is a Class II compound with low solubility and high permeability. The PSD of Nifedipine indicated that it is a crystalline powder. X-ray diffraction pattern of Nifedipine diffraction peaks confirms the D form solid state. The chemical structure of nifedipine was confirmed by UV-Visible Spectroscopy, FT-IR Spectroscopy, NMR Spectroscopy, Mass Spectrometry and melting point was determined by thermal analysis by DSC. The solid-state stability by FT-IR, DSC, accelerated stability studies in presence of tablet excipients has been conducted. The FT-IR overlay spectral studies indicated that there were no intermolecular interactions between Nifedipine and selected excipients. In the DSC thermograms of physical mixture, and DSC graph of Nifedipine controlled release granules, the drug melting peak was quite visible confirms that no physiochemical interaction between Nifedipine and excipients. Accelerated stability studies of blend at 40ºC + 2 ºC / 75% R.H for 1month, the % purity of Nifedipine is > 99% and % of Nifedipine impurities A and B with in prescribed limits. Therefore, the selected 99.6 99.5 99.6 99.7 99.8 99.6 99.5 99.6 99.4 99.3 99.4 99.3 99.6 99.7 0.03 0.04 0.04 0.03 0.05 0.06 0.06 0.08 0.06 0.04 0.06 0.07 0.06 0.04 0.06 0.06 0.05 0.03 0.04 0.07 0.07 0.05 0.06 0.06 0.07 0.08 0.05 0.06 BDL (<0.01%) BDL (<0.01%) BDL (<0.01%) BDL (<0.01%) BDL (<0.01%) BDL (<0.01%) BDL (<0.01%) BDL (<0.01%) BDL (<0.01%) BDL (<0.01%) BDL (<0.01%) BDL (<0.01%) BDL (<0.01%) BDL (<0.01%) excipients were compatible with Nifedipine and employed to assist in the preparation of controlled release granules of this therapeutic agent and further development of anti-hypertensive bilayer tablets. Acknowledgements This research was sponsored and supported by M/s SM Pharmaceuticals Sdn. Bhd, Malaysia for procuring Materials, Chemicals, Reference Materials, and for their valid support for testing of samples of the research study. Ethical Clearance This part of research work has not carried on any animal or Human, and this work was carried on chemical and instrumental analysis. Source of Funding This research was in part of my Ph. D research work and the funding was sourced from SM Pharmaceuticals Sdn Bhd, Malaysia. Conflict of Interest The SM Pharmaceuticals Sdn Bhd, Research and Development Centre had no association in the writing of the manuscript as well as in the decision to submit the article for publication. The authors have indicated that they have no competing interest regarding the content of this article REFERENCES 1. Alan J. Zillich, Jay Garg, Sanjib Basu, George L. Bakris, & Barry L. Carter. (2006). Thiazide Diuretics, Potassium, and the Development of Diabetes. Hypertension, 48, 219–224. 2. Marvin Moser and Henry, R. Black (1998). The role of combination therapy in the treatment of hypertension. 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