|Year : 2012 | Volume
| Issue : 4 | Page : 286-291
Development and validation of stability indicating UPLC assay method for ziprasidone active pharma ingredient
Sonam Mittal1, Abhishek Gupta2, Balasubramanian Narasimhan3, Kona S Srinivas2, Ravi Shankar Gupta2, Vinod Prasad Semwal2
1 Department of Pharmaceutical Chemistry, Vaish Institute of Pharmaceutical Education and Research (VIPER), Rohtak, Haryana, India
2 Department of Analytical Chemistry, Daiichi Sankyo India Pharma Pvt. Ltd., Gurgaon, Haryana, India
3 Department of Pharmaceutical Sciences, Maharshi Dayanand University, Rohtak, Haryana, India
|Date of Web Publication||1-Nov-2012|
Department of Analytical Chemistry, Daiichi Sankyo Research Centre in India, Plot-20, Sector-18, Gurgaon, Haryana
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Background: Ziprasidone, a novel antipsychotic, exhibits a potent highly selective antagonistic activity on D2 and 5HT2A receptors. Literature survey for ziprasidone revealed several analytical methods based on different techniques but no UPLC method has been reported so far. Aim: Aim of this research paper is to present a simple and rapid stability indicating isocratic, ultra performance liquid chromatographic (UPLC) method which was developed and validated for the determination of ziprasidone active pharmaceutical ingredient. Forced degradation studies of ziprasidone were studied under acid, base, oxidative hydrolysis, thermal stress and photo stress conditions. Materials and Methods: The quantitative determination of ziprasidone drug was performed on a Supelco analytical column (100×2.1 mm i.d., 2.7 ΅m) with 10 mM ammonium acetate buffer (pH: 6.7) and acetonitrile (ACN) as mobile phase with the ratio (55:45-Buffer:ACN) at a flow rate of 0.35 ml/ min. For UPLC method, UV detection was made at 318 nm and the run time was 3 min. Developed UPLC method was validated as per ICH guidelines. Results and Conclusion: Mild degradation of the drug substance was observed during oxidative hydrolysis and considerable degradation observed during basic hydrolysis. During method validation, parameters such as precision, linearity, ruggedness, stability, robustness, and specificity were evaluated, which remained within acceptable limits. Developed UPLC method was successfully applied for evaluating assay of Ziprasidone active Pharma ingredient.
Keywords: Antipsychotic, stability indicating assay, ultra performance liquid chromatographic, validation, ziprasidone
|How to cite this article:|
Mittal S, Gupta A, Narasimhan B, Srinivas KS, Gupta RS, Semwal VP. Development and validation of stability indicating UPLC assay method for ziprasidone active pharma ingredient. Chron Young Sci 2012;3:286-91
|How to cite this URL:|
Mittal S, Gupta A, Narasimhan B, Srinivas KS, Gupta RS, Semwal VP. Development and validation of stability indicating UPLC assay method for ziprasidone active pharma ingredient. Chron Young Sci [serial online] 2012 [cited 2019 Mar 24];3:286-91. Available from: http://www.cysonline.org/text.asp?2012/3/4/286/103097
| Introduction|| |
(5-[2-[4-(1, 2-benzisothiazol-3-yl)-1-piperazinyl] ethyl]-6-chloro-1, 3-dihydro-2 H-indol-2-one) [Figure 1], a novel antipsychotic, exhibits a potent highly selective antagonistic activity on D 2 and 5HT 2A receptors. It has a high affinity for 5HT 1a , 5HT 1d , 5HT 2c receptor subtypes that could contribute to its overall therapeutic effect. , Ziprasidone is most effective in schizophrenia, a chronic illness that requires lifelong treatment spread over approximately 1% of the world's population. ,
Literature survey for ziprasidone revealed several analytical methods based on different techniques, viz, LC-MS ,, assay for their quantification in plasma and brain, high performance liquid chromatography (HPLC) ,, method for simultaneous determination of ziprasidone in capsule formulation, and HPLC-UV  methods for determination ziprasidone in human plasma and urine, LC  with fluorescence for determination of plasma ziprasidone, and capillary zone electrophoresis  for determination of ziprasidone in pharmaceutical formulations.
Ultra performance liquid chromatography (UPLC) is a recent technique in liquid chromatography, which enables significant reduction in separation time and solvent consumption. Literature reports reveals that UPLC system allows about ninefold decrease in analysis time when compared with conventional HPLC system using 5 μm particle size analytical columns and about threefold decrease in analysis time in comparison to 3 μm particle size analytical column without compromise on overall separation. ,
Stability testing forms an important part of the process of drug product development. The purpose of stability testing is to provide evidence on how the quality of drug substance or drug product varies with time under the influence of variety of environmental factors such as temperature, humidity, and light and enables recommendation of storage conditions, retest periods, and shelf life to be established. The present investigation was undertaken to establish the stability indicating UPLC assay method for the estimation of ziprasidone as recommended by the International Conference on Harmonization (ICH) guidelines  and USP. 
| Materials and Methods|| |
Chemicals and reagents
Reference standard of ziprasidone hydrochloride (Purity 99.5% Expiry June 12) was gifted by Ranbaxy Research Laboratories, Gurgaon. Acetonitrice of HPLC grade was taken form Spectrochem Labs, Mumbai, India, and ammonium acetate and hydrogen peroxide of analytical reagent grade were obtained from Qualigens, Mumbai, India. Potassium dihydrogen phosphate, triethylamine, and sodium hydroxide of laboratory reagent grade were obtained from SD fine Chemicals Ltd., Mumbai, India.
Solution of ammonium acetate (0.01 M) was prepared by dissolving about 0.77 g of ammonium acetate in 1 l of water for HPLC. The pH of this solution was adjusted to 6.7 with acetic acid. The buffer preparation was found stable with respect to pH and visual clarity for 48 h.
Analyses were performed on Acquity UPLC TM system (Waters, Milford, MA, USA), consisting of binary solvent manager, sample manager, and PDA detector. The detector was set at sampling rate of 20 points/s and filter time constant of 0.2 s. System control, data collection, and data processing were accomplished using Waters Empower TM chromatography data software. The analytical column was 100 × 2.1 mm UPLC Supelco, 2.7 μm (Merck, USA). The separation of ziprasidone was achieved by isocratic elution using acetonitrile (ACN) and acetate buffer (pH 6.7; 0.01 M). The optimized conditions were as follows: isocratic elution (55:45-Buffer:ACN) with an injection volume of 5 μl and flow rate of 0.35 ml/min at 40° C and detection wavelength of 318 nm (absorbance maxima/lambda max being 318) with back pressure of 6500 psi. The run time finalized was of 3 min.
Preparation of standard solution
Standard solution was prepared by dissolving 80 mg of standard substance in water: ACN (60: 40 v/v) mixture to obtain solution containing 80 μg/ml of ziprasidone.
Sample solution (80 μg/ml) was prepared by weighing 80.0 mg of drug into 100 ml volumetric flask. Drug was dissolved into small volume of diluent, water: ACN (60: 40 v/v) and sonicated for 1 min. Then volume was made up to 100 ml with diluent. From the each stock solution, 10 ml solution was transferred to 100 ml volumetric flask and volume was made up to 100 ml to get the final sample concentration of 80 μg/ml.
System suitability parameters were measured so as to verify the system performance. System precision was determined on six replicate injections of standard preparations.
Forced degradation studies were performed to demonstrate selectivity and stability indicating capability of the proposed method. The samples of ziprasidone were exposed to acidic, alkaline, thermal oxidative, and photolytic degradation conditions. All the exposed standards and samples were than analyzed by proposed method.
Linearity was demonstrated from 70% to 130% of standard concentration using minimum six calibration level (70%, 80%, 90%, 100%, 120%, and 130%) for the compound. The method of linear regression was used for data evaluation. Peak areas of sample compound were plotted against respective concentrations.
Precision was investigated using sample preparation procedure for six real samples and analyzed by proposed method. Intermediate precision was studied using different column, performing analysis on different day and also by different analyst.
The robustness is a measure of method capacity to remain unaffected by small but deliberate changes in chromatographic conditions such as change in pH of buffer (±0.2 units), column temperature (±5°C), flow rate (±10%) as well as ratio of mobile phase (±2 units).
Stability of sample preparation
Stability of sample solution was established by storage of sample solution at ambient temperature for 24 h followed by its assay, which was then compared against fresh sample.
Results and Discussion
For analysis of ziprasidone, different chromatographic conditions were tried on HPLC and UPLC and results obtained were compared. Isocratic elution is simple and requires only one pump and flat baseline separation for easy and reproducible results. The results from HPLC [Figure 2]b involve few main problems: longer run time, lack of good peak shape, and low response, while with UPLC all these problems are solved and thus isocratic run using UPLC provide proper peak with a good baseline in 3 min time [Figure 2]a [Table 1], a-c was selected for the analysis of ziprasidone.
|Figure 2a: UPLC chromatogram of ziprasidone|
Figure 2b: HPLC chromatogram of ziprasidone
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|Table 1: Different chromatographic conditions applied and results obtained|
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Among various columns available for UPLC analysis, Supelco C18, 2.7 μm, (100 × 2.1) mm i.d. column was preferred, because it provides appreciable peak shape and resolution and absorbance were good. Among different mobile phase used, the mobile phase consisted of ACN and 10 mM ammonium acetate with an apparent pH adjusted to 6.7±0.1 with ammonia and acetic acid was found to be suitable for analysis of ziprasidone. Further, a flow rate of 0.35 ml/min, an injection volume of 5 μl, and UV detection at 318 nm for drug were found to be suitable for analysis. [Figure 3] indicates the peak obtained for the sample by the selected method.
Analytical parameters and validation
After satisfactory development of method, it was subjected to method validation as per ICH guidelines. The method was validated to demonstrate that it is suitable for its intended purpose by the standard procedure to evaluate adequate validation characteristics (precision, linearity, robustness, stability indicating capability).
Results of other system suitability parameters such as theoretical plates, purity angle, and purity threshold are presented in [Table 2]. The data presented in [Table 2] indicated the acceptable system suitability parameters, as the % RSD is not more than 2%. Tailing factor was not more than 2 and theoretical plates are more than 1000 and purity angle was less than purity threshold.
The percentage (%) RSD of area count of six replicate injections was below 2 which indicated the system precision.
The results of forced degradation studies are given in [Table 3]. Ziprasidone was found sensitive to basic hydrolysis. The assay value was decreased to 73.52% and degradation peaks were observed in the chromatogram. Ziprasidone was found stable to acid hydrolysis yielding assay value of 101.80%. Chromatograms of base and acid degraded samples for ziprasidone are presented in [Figure 4] and [Figure 5]. Ziprasidone was sensitive to oxidative conditions and the assay value decreased to 75.66%. Chromatogram of oxidation degraded ziprasidone is shown in [Figure 6]. Ziprasidone was found to be stable under thermal and photo stress degradation studies. Peaks due to ziprasidone were investigated for spectral purity in the chromatogram of all exposed samples and standards and found to be spectrally pure.
The response was found linear from 70% to 130% standard concentration. The correlation coefficient (R 2 ) was greater than 0.0.99 [Table 4] and [Figure 7].
The intra-assay precision of ziprasidone was 81.67% with a % RSD of 1.49%. Inter-assay precision for ziprasidone was 80.65% with % RSD of 0.79% [Table 4]. Low values of RSD indicated that the method is precise.
No significant effect was observed on system suitability parameters such as theoretical plates, purity angle, and purity threshold, when small but deliberate changes were made to chromatographic conditions such as change in flow rate (±10%), temperature (±5 units), pH (±0.2 units), and organic content (±2%). The results are presented in [Table 2], along with system suitability parameters of normal methodology. Thus, the method was found to be robust with respect to variability in above condition
Stability in sample solution
Sample solution did not show any appreciable change in assay value when stored at ambient temperature up to 24 h. Assay results are presented in [Table 4].
| Conclusion|| |
A novel UPLC method was successfully developed and validated for determination of ziprasidone. The total run time was 3 min, within which drug got eluted. Method validation results have proved the method to be selective, precise, accurate, robust and stability indicating. Sample solution stability was established for determination of assay and impurities. This method can be successfully applied for the routine analysis and stability study. Also, it can be used for determination of content uniformity and dissolution profiling of this product, where sample load is higher and high throughput is essential for faster delivery of results. Overall, the method provides high-throughput solution for determination of ziprasidone with excellent selectivity, precision, and accuracy.
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[Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7]
[Table 1], [Table 2], [Table 3], [Table 4]