Chromatography
Improved method by HPLC for Ibuprofen, Pseudoephedrine Hydrochloride and Chlorpheniramine maleate active ingredients and degradation products for the finished product in syrup dosage form
Mar 11 2021
Author: Yasemin Gökdeniz*, Gizem Alkan, Suna Beyoglu, Asiye Sezgin, Figen Onuk Gören, Pharmactive Pharmaceutical Company, Research and Development Center, Çerkezköy, Tekirdag, Turkey
*Corresponding Author on behalf
of Pharmactive Pharmaceutical Company
The objective of this paper is to develop a simple, precise, accurate, and reproducible improved reversed phase high performance liquid chromatographic method for the quantitative determination of Chlorpheniramine Maleate, Ibuprofen and Pseudoephedrine Hydrochloride possible degradation product in a combined pharmaceutical dosage form.
The objective of this paper is to develop a simple, precise, accurate, and reproducible improved reversed phase high performance liquid chromatographic method for the quantitative determination of Chlorpheniramine Maleate, Ibuprofen and Pseudoephedrine Hydrochloride possible degradation product in a combined pharmaceutical dosage form.
Introduction
3-(4-chlorophenyl)-N,N-dimethyl-3-pyridin-2-ylpropan-1-amine is the IUPAC name for Chlorpheniramine Maleate. The empirical formula for Chlorpheniramine Maleate is C20H23ClN2O4 (Figure 1. chemical structure of Chlorpheniramine Maleate salt). Chlorpheniramine Maleate is a H-1 receptor blocker and acts as an antihistamine used to relieve symptoms of allergy, hay fever, and the common cold. These symptoms include rash, watery eyes, itchy eyes/nose/throat/skin, cough, runny nose, and sneezing. 2-[4-(2-methyl-propyl)phenyl]propanoic acid is the IUPAC name of the Ibuprofen. The empirical formula for Ibuprofen is C13H18O2 (Figure 2. chemical structure of Ibuprofen). Ibuprofen is a nonselective inhibitor of COX-2, an enzyme involved in prostaglandin synthesis of the arachidonic acid pathway. Its pharmacological effects are believed to be due to inhibition of COX-2 which decrease the synthesis of prostaglandin involved in mediating inflammation pain, fever, and swelling [1-5]. (1S,2S)-2-(methylamino)-1-phenylpropan-1-ol hydrochloride is the IUPAC name of the Pseudoephedrine Hydrochloride. The empirical formula for Pseudoephedrine Hydrochloride is C10H16ClNO (Figure 3. chemical structure of Pseudoephedrine Hydrochloride) The most common use for Pseudoephedrine is as a decongestant, for conditions including nasal congestion, sinus congestion, and eustachian tube congestion, as it shrinks swollen nasal mucous membranes and reduces tissue hyperemia and edema [6] Other uses include vasomotor rhinitis, first-line treatment for priapism, and off-label use for hyperprolactinemia, while veterinarians often use pseudoephedrine off-label to treat incontinence in dogs and cats [6]. In conjunction with other medications, Pseudoephedrine is often used to treat allergic rhinitis, croup, sinusitis, otitis media, and tracheobronchitis [7].
Figure 1. Chemical structure of Chlorpheniramine Maleate.
Figure 2. Chemical structure of Ibuprofen.
Figure 3. Chemical structure of Pseudoephedrine Hydrochloride.
There is no related substances method reported for the simultaneous estimation of Chlorpheniramine Maleate (CPM), Ibuprofen (IBU), and Pseudoephedrine Hydrochloride (PSEH) in combined dosage form. The present study involved the development and validation of RP HPLC method for the estimation of CPM, IBU, and PSEH in combined pharmaceutical dosage form.
Experimental
Reagents and Materials
Analytically pure standards of CPM, IBU, and PSEH were obtained from Supriya Lifescience (Mumbai, India), Hubei (Hubei, China) and Malladi (Guindy, Chennai) respectively. Analytically pure impurity standards of Ephedrine, Ibuprofen Impurity E and Chlorpheniramine Maleate Impurity C were obtained from Ph.Eur (European Pharmacopoeia), USP (United States Pharmacopeia) and Pharmaffiliates(India) respectively. HPLC grade Methanol and Acetonitrile were obtained from JT Baker. HPLC grade Decan-1 Sulphonic Acid Sodium Salt, Triethylamine and Phosporic Acid 85% were obtained from Merck. HPLC grade Sodium Hydroxide were obtained from Sigma Aldrich. HPLC grade Potassium Dihydrogen Phosphate were obtained from Panreac. The water was distilled and deionised by using Millipore Milli Q Ultrapure system. Syrup formulation (Pharmactive Pharmaceutical Company, Turkey) containing labelled amount of 2 mg of Chlorpheniramine Maleate, 200 mg of Ibuprofen, and 30 mg of Pseudoephedrine Hydrochloride was used for the study.
Apparatus and Chromatographic Conditions
The liquid chromatographic system consists of Shimadzu LC20 series equipped with a PDA detector, quaternary pump, and automatic injector, 20 µL fixed loop. The analytes were monitored at 215 nm for Ibuprofen and Pseudoephedrine Hydrochloride and 225 nm for Chlorpheniramine Maleate. Chromatographic analysis was performed using a Direct In-Line Filter and 1 cm Guard Cartridge holders guard column and YMC Triart C18 column (5 µm × 250mm × 4.6 mm). All the drugs and chemicals were weighed on Mettler Toledo XP26, XP2U,XP205 electronic balance. Chromatographic separation was achieved using timed gradient mode (Table 1). The mobile phase consisted of A: pH 2.5 buffer (3.4 g KH2PO4, 5 g Decan-1-sulphonic acid sodium salt, and 5 mL of Triethylamine in 1000 mL deionised water adjusted to pH 2.5 with H3PO4) and B: Methanol (100%). The flow rate of the mobile phase was 0.7 mL/min. The column temperature was 45˚C and sample temperature was 15˚C. The injection volume was 20 μL. Diluent was pH:7.2 Buffer:Methanol (500:500 v/v).
Table 1. Gradient Mode.
Time (min.) Mobile Phase A (%) Mobile Phase B (%) FlowRate
0.01 48 52 0.7
27.00 48 52 0.7
27.01 35 65 0.7
37.00 35 65 0.7
37.01 35 65 0.6
55.00 35 65 0.6
55.01 35 65 0.7
60.00 35 65 0.7
60.01 48 52 0.7
75.00 48 52 0.7
Diluent Preparation
Preparation of pH 7.2 Buffer: 6.8 g of Potassium Dihydrogen Phosphate and 1.2 g of Sodium Hydroxide were dissolved in 1000 mL of distilled water. pH is adjusted to 7.2 with ortho-phosphoric acid.
Preparation of Diuent: 500 mL of pH 7.2 Buffer and 500 mL of Methanol was mixed, filtered through 0.45 μm membrane filter, and degassed.
Standard Solution Preparation
PSEH stock standard solution was prepared by transferring an accurately weighed 9 mg of Pseudoephedrine Hydrochloride working standard into a 20-mL volumetric flask. 10 mL of diluent was added followed by sonication for 5 min using an ultrasonic source. The solution was brought up to final volume with the diluent. CPM stock standard solution was prepared by transferring an accurately weighed 6 mg of Chlorpheniramine Maleate working standard into a 100-mL volumetric flask. 60 mL of diluent was added followed by sonication for 5 min using an ultrasonic source. The solution was brought up to final volume with the diluent. IBU stock standard solution was prepared by transferring an accurately weighed 12 mg of Ibuprofen working standard into a 20-mL volumetric flask. 10 mL of diluent was added followed by sonication for 5 min using an ultrasonic source. The solution was brought up to final volume with the diluent. For preparation of standard solution, 2 mL of PSEH stock standard solution, 0.5 mL of CPM stock standard solution, 2 mL of IBU stock standard solution transferred into a 100-mL volumetric flask and the solution was brought up to final volume with the diluent (CIbuprofen= 0.012 mg/mL, CPseudoephedrine HCl= 0.009 mg/mL, CChlorphenamine Maleate= 0.0003 mg/mL).
Sample Solution Preparation
Approximately 3 mL syrup (equivalent to 60 mg Ibuprofen, 9 mg Pseudoephedrine HCl, and 0.6 mg Chlorpheniramine Maleate) was weighed into a 10-mL volumetric flask, dissolved with 5 mL of diluent was added followed by sonication for 2 min using an ultrasonic source. The solution was brought up to final volume with the diluent and vortex for 1 min and filtered through 0.22 μm PVDF filter and injected into the HPLC system. (CIbuprofen= 6.0 mg/mL, CPseudoephedrine HCl= 0.9 mg/mL, CChlorpheniramine Maleate= 0.06 mg/mL).
Method Validation
Precision
Precision was determined as both repeatability and intermediate precision, in accordance with ICH recommendations. Standard solution prepared at 100% working concentration was injected six times to evaluate the system precision. Peak areas obtained from injections and Relative Standard Deviation (RSD%) value between them were calculated. The RSD % of not more than 5% was recommended. Repeatability (precision of the method) and intermediate precision were also evaluated. For repeatability, 6 syrup spiked samples, which were prepared by adding known impurities at the limit concentration and the analysis carried out. For intermediate precision, 6 syrup spiked samples were prepared and analysed on a different day using a different device and column by a different analyst. Repeatability and intermediate precision results, for IBU, PSEH and CPM impurities were calculated. The difference between result means of the two analysts were also calculated.
Linearity and range
To prove the linear response relation, peak areas of the standard solutions between 0.015% LOQ (Limit of Quantification) % and 1.2% of specification at 7 different concentrations (0.015%, 0.1%, 0.3%, 0.5%, 0.8%, 1.0%, 1.2% ) for PSEH, the standard solutions between 0.002 % LOQ (Limit of Quantification) % and 0.24% of specification at 7 different concentrations (0.002%, 0.05%, 0.06%, 0.1%,0.16%, 0.2%, 0.24%) for IBU, the standard solutions between 0.1% LOQ (Limit of Quantification) % and 0.6 % of specification at 6 different concentrations (0.1%, 0.15%, 0.25%, 0.4%, 0.5%, 0.6%) for CPM, the standard solutions between 0.015% LOQ (Limit of Quantification) % and 1.2% of specification at 7 different concentrations (0.015%, 0.1%, 0.3%, 0.5%, 0.8%, 1.0%, 1.2%) for Ephedrine, the standard solutions between 0.002% LOQ (Limit of Quantification) % and 0.24% of specification at 7 different concentrations (0.002%, 0.05%, 0.06%, 0.1%, 0.16%, 0.2%, 0.24% ) for Ibuprofen Impurity E, the standard solutions between 0.1% LOQ (Limit of Quantification) % and 0.6% of specification at 6 different concentrations (0.1%, 0.15%, 0.25%, 0.4%, 0.5%, 0.6%) for Chlorpheniramine Maleate Impurity C were measured and linearity curves were plotted. The prepared linearity solutions were analysed, y=ax+b threshold was found and regression analysis was performed (y: area, a: slope, b: intercept, x: concentration, mg/mL). Correlation coefficient between concentration and areas of more than 0.99 was sought. The response factor for Ephedrine, Ibuprofen Impurity E, Chlorfeniramine Maleate Impurity C impurities with respectively calculated according to PSEH, IBU, CPM. Then this response factor was used in denomination of calculation formula of % Impurity.
Detection (LOD) and Quantification (LOQ) Limits
LOD and LOQ were determined by the Signal/Noise (S/N) method. All of the Impurities and active substances were injected and obtained S/N were evaluated.
Specificity
Specificity test was performed to demonstrate the ability of the analytical method to measure only the intended substances in a given sample. Standard, unspiked sample, placebo, impurity standards, spiked sample, diluent were injected into the HPLC system and analysed. Chromatogram of the injected solutions were evaluated.
Accuracy and recovery
Accuracy was determined by the standard addition method. Accuracy of method was perform at LOQ%, release limit level and 120% of the shelf life limit of the working concentration by adding Ephedrine, Ibuprofen Impurity E and Chlorpheniramine Maleate Impurity C to the sample solution. In totally, 9 samples were prepared (3 for each level) and injected triplicate. Recovery (%) and RSD (%) were calculated for each concentration.
Robustness
The robustness of the developed method was measured to evaluate the influence of deliberate variation in the chromatographic conditions. The robustness of the method was evaluated by changing the flow rate (0.6 and 0.8 mL/min), column temperature (40˚C and 50˚C) column brand (Inertsil ODS-4 column 250 x 4.6 mm 5 µm), and mobile phase pH change (pH:2.4-pH:2.6). Robustness testing was performed in order to obtain information about those critical parameters affecting the response (peak area, retention time).
Results and discussion
Precision
System precision was determined by performing injection repeatability standard solution and the obtained RSD values for PSEH, IBU and CPM were found 0.11%, 0.06% and 3.4% respectively. The low RSD values indicate that the system is precise. Also in comparison of repeatability and intermediate precision results of differences between means of two analysts sample results for the precision.
Table 2. Repeatability data of validated method (%).
Injection No Ephedrine (%) Ibuprofen Chlorpheniramine Impurity E (%) Maleate Impurity C (%)
1 1.03 0.21 0.48
2 1.04 0.20 0.50
3 1.03 0.20 0.50
4 1.03 0.20 0.50
5 1.03 0.20 0.50
6 0.98 0.20 0.46
Average 1.02 0.20 0.49
SD 0.02 0.00 0.02
RSD (%) 2.1 2.0 3.4
Table 3. Intermediate precision data of validated method (%).
Injection No Ephedrine (%) Ibuprofen Chlorpheniramine Impurity E (%) Maleate Impurity C (%)
1 1.05 0.20 0.50
2 1.05 0.20 0.51
3 1.04 0.20 0.48
4 1.04 0.20 0.49
5 1.03 0.20 0.48
6 1.02 0.20 0.51
Average 1.04 0.20 0.50
SD 0.01 0.00 0.01
RSD (%) 1.1 0.0 2.8
Table 4. Repeatability and Intermediate precision data mean and differences results between two analyst (%)
Repeatability (%) Intermediate Differences (%)
Precision(%)
Ephedrine 1.02 1.04 0.02
Ibuprofen Impurity E 0.20 0.20 0.00
Chlorpheniramine
Maleate Impurity C 0.49 0.50 0.01
The method is found to be precise with respect to the criteria of the system precision, intermediate precision and repeatability.
Linearity and range
The calibration curve for PSEH was found to be linear in the range of 0.000135–0.010801 mg/mL with a correlation coefficient of 1.00. The calibration curve for Ephedrine was found to be linear in the range of 0.000135–0.010782 mg/mL with a correlation coefficient of 1.00. The calibration curve for IBU was found to be linear in the range of 0.000121–0.014537 mg/mL with a correlation coefficient of 1.00.
The calibration curve for Ibuprofen Impurity E was found to be linear in the range of 0.000120–0.014371 mg/mL with a correlation coefficient of 1.00. The calibration curve for CPM was found to be linear in the range of 0.0000610–0.000366 mg/mL with a correlation coefficient of 1.00. The calibration curve for Chlorpheniramine Maleate Impurity C was found to be linear in the range of 0.0000447–0.000268 mg/mL with a correlation coefficient of 0.99.
Table 5. Linearity Data for PSEH of validated method.
Concentration Level in (%) Theoretical Concentration of Response (Area)
PSEH (mg/mL)(x-value) (y value)
0.015% 0.000135 5422.2
0.1% 0.000900 37886.2
0.3% 0.002700 108824.5
0.5% 0.004500 182038.8
0.8% 0.007201 289913.0
1.0% 0.009001 361572.8
1.2% 0.010801 434768.3
Table 6. Linearity Data for Ephedrine of validated method.
Concentration Level in (%) Theoretical Concentration of Response (Area)
Ephedrine (mg/mL) (x-value) (y value)
0.015% 0.000135 5493.8
0.1% 0.000898 37196.8
0.3% 0.002696 106510.0
0.5% 0.004493 179369.3
0.8% 0.007188 284115.0
1.0% 0.008985 358511.0
1.2% 0.010782 428597.8
Table 7. Linearity Data for Ibuprofen of validated method.
Concentration Level in (%) Theoretical Concentration of Response (Area)
Ibuprofen (mg/mL) (x-value) (y value)
0.002% 0.000121 12918.8
0.05% 0.003028 254683.8
0.06% 0.003634 296903.5
0.1% 0.006057 480526.3
0.16% 0.009691 782628.5
0.2% 0.012114 984041.7
0.24% 0.014537 1183409.2
Table 8. Linearity Data for Ibuprofen Impurity E of validated method.
Concentration Level in (%) Theoretical Concentration of Ibuprofen Response Impurity E (mg/mL) (x-value) (Area) (y value)
0.002% 0.000120 19923.0
0.05% 0.002994 252915.2
0.06% 0.003593 297940.5
0.1% 0.005988 486689.5
0.16% 0.009580 796728.5
0.2% 0.011976 1003007.7
0.24% 0.014371 1176968.8
Table 9. Linearity Data for Chlorpheniramine Maleate of validated method.
Concentration Level in (%) Theoretical Concentration Response (Area)
of CHL (mg/mL) (x-value) (y value)
0.1% 0.000061 3566.0
0.15% 0.000091 5309.0
0.25% 0.000152 8992.7
0.4% 0.000244 14439.0
0.5% 0.000305 17768.8
0.6% 0.000366 22039.5
Table 10. Linearity Data for Chlorpheniramine Maleate Impurity C of validated method.
Concentration Level in (%) Theoretical Concentration of Response CPM Impurity C (mg/mL) (x-value) (Area) (y value)
0.1% 0.000045 2732.0
0.15% 0.000067 2912.5
0.25% 0.000112 5461.2
0.4% 0.000179 8322.5
0.5% 0.000223 11825.3
0.6% 0.000268 13031.3
Figure 4. Linearity Plot of Pseudoephedrine HCl: A graph of Concentration (x value) against area response obtained (y- value) is plotted.
Figure 5. Linearity Plot of Ephedrine: A graph of Concentration (x value) against area response obtained (y- value) is plotted.
Figure 6: Linearity Plot of Ibuprofen: A graph of Concentration (x value) against area response obtained (y- value) is plotted.
Figure 7: Linearity Plot of Ibuprofen Impurity E: A graph of Concentration (x value) against area response obtained (y- value) is plotted.
Figure 8: Linearity Plot of Chlorpheniramine Maleate: A graph of Concentration (x value) against area response obtained (y- value) is plotted.
Figure 9: Linearity Plot of Chlorpheniramine Maleate Impurity C: A graph of Concentration (x value) against area response obtained (y- value) is plotted.
Table 11. Validation parameters for CPM, IBU, and PSEH.
Parameter Pseudoephedrine Ibuprofen Chlorpheniramine
Maleate
% Y-axis 802.27 1836.1 159.8
Intercept
Slope 4x107 8x107 6x107
Response Factor
for Division 1.00* 1.00* 1.00*
*It will be used for quantification of unknown impurities
Table 12. Response Factor of Impurities.
Parameter Ephedrine Ibuprofen Impurity E Chlorpheniramine
Maleate Impurity C % Y-axis 407.61 5933.8 29.17
Intercept
Slope 4x107 8x107 5x107
Response Factor
for Division 1.01** 0.99*** 1.21****
** Calculated based on Pseudoephedrine HCl
*** Calculated based on Ibuprofen
****Calculated based on Chlorpheniramine Maleate
Specificity
The applied method demonstrated excellent specificity. There are no interfering peaks in the chromatogram originating from the diluent, impurities and placebo. The spectrum of IBU, PSEH and CPM peaks did not interfere with the other peaks in Standard and Sample solution chromatograms. The selectivity of the method was proved. Spiked sample data is shown in Figure 10.
Accuracy and Recovery
Accuracy was determined by the standard addition method. Accuracy of the method was performed at LOQ%, release limit level and 120% of the shelf life limit of the working concentration by adding Ephedrine, Ibuprofen Impurity E and Chlorpheniramine Maleate Impurity C to the sample solution. In totally, 9 samples were prepared (3 for each level) and injected triplicate. Recovery (%) and RSD (%) were calculated for each concentration. Results are shown in Table 13-15. All results meet the acceptance criteria.
Table 13. Recovery data of validated method (%).
Solution Name Recovery of Ephedrine (%)
0.015 % 91.39
90.09
100.62
1.00% 99.99
101.07
100.96
1.20% 96.04
98.18
96.53
Average 97.21
SD 3.3
RSD % 3.4
Table 14. Recovery data of validated method (%).
Solution Name Recovery of Ibuprofen
Impurity E (%)
0.002% 101.87
98.68
90.66
0.2% 101.81
97.56
95.57
0.24% 96.03
96.30
97.92
Average 97.38
SD 3.4
RSD % 3.5
Table 15. Recovery data of validated method (%).
Solution Name Recovery of Chlorpheniramine
Maleate Impurity C (%)
0.1% 118.56
106.95
106.03
0.5% 103.69
108.06
109.68
0.6% 108.07
107.32
103.10
Average 107.94
SD 2.3
RSD % 2.1
Detection (LOD) and Quantification (LOQ) Limits
The LOD and LOQ of the method were show the Table 16 and Table 17. The method can be used for detection and quantification of trace amounts of Ibuprofen, Pseudoephedrine HCl, Chlorpheniramine Maleate impurities over a wide range of concentrations.
Table 16. LOD data of validated method (%).
Substances LOD Concentration S/N (Signal/Noise)
(mg/mL)
Ephedrine 0.00000405 3.4
Ibuprofen Impurity E 0.000036 13.3
Chlorpheniramine Maleate
Impurity C 0.000018 3.9
Pseudoephedrine HCl 0.00000405 3.4
Ibuprofen 0.000036 10.6
Chlorpheniramine Maleate 0.000018 4.5
Table 17. LOQ data of validated method (%).
Substances LOQ Concentration S/N (Signal/Noise)
(mg/mL)
Ephedrine 0.0000135 12.5
Ibuprofen Impurity E 0.00012 46.0
Chlorpheniramine Maleate
Impurity C 0.00006 10.6
Pseudoephedrine HCl 0.0000135 11.6
Ibuprofen 0.00012 26.9
Chlorpheniramine Maleate 0.00006 15.0
Robustness
With the Robustness parameter, the effect of changes in the method parameters and solution stability were examined. For this aim parameters which listed below were performed;
- Flow rate: 0.6 mL/min and 0.8 mL/min
- Column Temperature: 40°C and 50°C
- Mobile Phase pH Change: pH:2.3 and pH:2.5
- Column Brand Change: Inertsil ODS 4 250 mm x 4.6 mm x 5.0 µm
Table 18. Retention time of the Robustness of the method by small changes in chromatographic conditions.
Parameter Ephedrine Ibuprofen Chlorpheniramine
(min) Impurity E (min) Maleate Impurity C (min)
Normal 24.27 50.69 42.59
Flow Rate-0.6 mL/min 28.95 55.70 45.54
Flow Rate-0.8 mL/min 21.52 46.97 40.55
Column Temperature: 40°C 26.35 51.83 43.52
Column Temperature: 50°C 23.12 49.47 42.17
Mobile Phase pH Change: pH:2.3 24.00 50.44 42.75
Mobile Phase pH Change: pH:2.5 - - -
Different Column 20.65 47.19 40.82
Table 19. Retention time of the Robustness of the method by small changes in chromatographic conditions.
Parameter Pseudoephedrine Ibuprofen Chlorpheniramine
HCl (min) (min) Maleate (min)
Normal 21.64 52.09 41.64
Flow Rate-0.6 mL/min 25.72 57.12 44.81
Flow Rate-0.8 mL/min 19.13 48.26 39.66
Column Temperature: 40°C 23.20 53.37 42.70
Column Temperature: 50°C 20.73 50.82 41.14
Mobile Phase pH Change: pH:2.3 21.41 51.88 41.80
Mobile Phase pH Change: pH:2.5 - - -
Different Column 18.81 48.26 40.20
Table 20. Relative Standard Deviation of the standard solution by small changes in chromatographic conditions.
Parameter Pseudoephedrine Ibuprofen Chlorpheniramine
HCl (%) (%) Maleate (%)
Normal 0.27 0.21 1.52
Flow Rate-0.6 mL/min 0.12 0.16 0.80
Flow Rate-0.8 mL/min 0.15 0.08 0.76
Column Temperature: 40°C 0.12 0.16 3.71
Column Temperature: 50°C 0.09 0.13 0.76
Mobile Phase pH Change: pH:2.3 0.27 0.21 0.69
Mobile Phase pH Change: pH:2.5 - - -
Different Column 0.32 4.45 4.09
Obtained results showed that there was no significant change in system suitability parameters for any components except pH change. The method is robust in respect of changes to flow rate, column temperature, different manufacturer column. The method is sensitive to pH change
Stability
The stability of the drug in solution during analysis was determined by repeated analysis of the standard and spiked sample solutions at autosampler temperature (15°C). Peak areas were recorded and similarity % was calculated. It was proved that standard solution is stable at 15°C for 117 hours and sample solution is stable at 15°C for 49 hours.
Table 21. Stability of the Standard Solution for Pseudoephedrine HCl.
Time (hours) Standard Solution Change %
(Pseudoephedrine) (Area)
0 300558 -
15 301195 0.2
24 301889 0.4
49 300232 0.1
83 303030 0.8
91 305897 1.8
117 300282 0.1
Table 22. Stability of the Standard Solution for Ibuprofen.
Time (hours) Standard Solution (Ibuprofen) Change %
(Area)
0 933057 -
15 927982 0.5
24 928964 0.4
49 929594 0.4
83 930633 0.3
91 931667 0.1
117 937448 0.5
Table 23. Stability of the Standard Solution for Chlorpheniramine Maleate.
Time (hours) Standard Solution Change %
(Chlorpheniramine Maleate) (Area)
0 16134 -
15 16000 0.8
24 16507 2.3
49 15953 1.1
83 16258 0.8
91 16469 2.1
117 16649 3.2
Table 24. Stability of the Spiked Sample Solution: Ephedrine.
Time (hours) Ephedrine Change %
(Area)
0 309912 -
15 310737 0.3
24 310114 0.1
37 310348 0.1
49 311437 0.5
83 314325 1.4
Table 25. Stability of the Spiked Sample Solution: Ibuprofen Impurity E
Time (hours) Ibuprofen Impurity E (Area) Change %
0 1051222 -
15 1043505 0.7
24 1044317 0.7
37 1039344 1.1
49 914473 1.1
83 913500 1.2
Table 26. Stability of the Spiked Sample Solution: Chlorpheniramine Maleate Impurity C.
Time (hours) Chlorpheniramine Maleate Change %
Impurity C (Area)
0 13080 -
15 13151 0.5
24 13036 0.3
37 12124 7.3
49 13124 0.3
83 11031 15.7
Conclusion
This HPLC method is sensitive, accurate, precise, reproducible, specific and stability-indicating. This method was found to be accurate and precise as indicated by the recovery studies and relative standard deviation. The Relative Response factor for impurities determined from validation can be used for quantification of impurities in Pseudoephedrine, Ibuprofen, Chlorpheniramine Maleate in routine testing, by which there is no need to use impurities’ standard. Furthermore, the HPLC method could also be suggested for the routine analysis of Ibuprofen, Pseudoephedrine Hydrochloride, and Chlorpheniramine Maleate and their related substances in pharmaceutical dosage formulations.
References
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