CHARACTERIZATION AND VALIDATION OF IMPURITIES RELATED TO PHARMACEUTICAL BULK DRUG (API) BY USING SOME ANALYTICAL TECHNIQUES

CHARACTERIZATION AND VALIDATION OF IMPURITIES RELATED TO PHARMACEUTICAL BULK DRUG (API) BY USING SOME ANALYTICAL TECHNIQUES

Champa P. Maurya ¹ and Manohar V. Lokhande ^*2

Department of Chemistry ¹, Jagdish Prasad Jhabarmal Tibrewala University, Jhunjhunu - 333001, Rajasthan, India.

Department of Chemistry ², Sathaye College, Vile Parle (E), Mumbai - 400057, Maharashtra, India.

ABSTRACT: Active Pharmaceutical Ingredient (API) of pharmaceutical bulk drug, four impurities were identified and were detected by a newly developed Reverse phase high performance liquid chromatographic (HPLC) method. The R.S.D.’s for SM-I, SM-II, SM- III and SM were found to be 1.48%, 1.71%, 1.67%, 6.37% respectively. These values are within the acceptance criteria of 10%.The limit of quantification values for SM-I, SM-II and SM-III were found to be 0.006 %, 0.006 %, and 0.006 % w.r.t. analyte concentration (500 μg /cm³), respectively. For determining method accuracy, known as unclean LOQ, the specified limit of 80%, 100% and 120% of the SM bulk sample (test preparation) was pointed. For confirming method precision six different test preparations of samples of SM were analyzed. Identified impurities were characterized by LC/MS/MS method. Identified impurities were unknown. Structural determination of such impurities was carried out by LC/MS/MS using electro spray ionization source and an ion trap mass analyzer. Structural identification by using nuclear magnetic resonance (NMR) and infrared (IR) spectroscopy.  The method was validated according to ICH guidelines with respect to Specificity, Precision, Accuracy, Linearity and Robustness.

Impurities, HPLC, LCMS/MS/MS, NMR and Validation

INTRODUCTION: Sulfa methoxypyrazine (SM) such as N1 - (3-Methoxypyrazin – 2 - yl) Sulphanilamide is a long-acting sulfonamide which has been used orally for the treatment of respiratory and urinary tract infections ¹. It gives the combination with pyrimethamine ^{2, 3} in the treatment of malaria. It has also been given in the ratio 4 parts of Sulfa methoxypyrazine to 5 parts of trimethoprim as a combination with uses similar to those of co-trimoxazole ⁴.

A few bio-analytical techniques were reported in the literature for the quantitative determination of Sulfa methoxypyrazine (SM) concentration in biological fluids using liquid chromatography and mass spectroscopic method ^{5, 6, 7}. However, so far there is no published report, describing the complete characterization of related impurities in Sulfa methoxypyrazine as active pharmaceutical ingredient (API). Some part of the paper is reported by using LC/MS/MS and isolation/synthesis of related substances in Sulfa methoxypyrazine Active Pharmaceutical Ingredient ⁸.

Impurity profile of a drug substance is critical for its security valuation and manufacturing process. It is required to identify and characterize the impurities in the pharmaceutical bulk drugs, if impurities are present above then the acceptable limits of 0.1 % ⁹. The present study deals with the identification and structural explanation of the process related impurities, which were found in the pharmaceutical bulk drugs Sulfa methoxypyrazine. Though, different methods of synthesis of Sulfa methoxypyrazine are reported, the selected route was safe, feasible & economical ¹⁰. However, the literature survey does not give any details regarding these impurities. Impurity profiling of drugs is the most important issue in the modern pharmaceutical analysis ^{11, 12} for developing process technology to manufacture high purity drug substance. During process development studies, four impurities were detected in both crude and pure samples of SM using a newly developed gradient reversed phase HPLC method. This paper also deals with the analytical method validation of a new HPLC method for quantitative determination of these impurities.

MATERIALS AND METHODS:

Materials: Samples of API were obtained from pharmaceutical laboratories, Chemical Research Division, Mumbai, India. HPLC grade CH₃CN and perchloric acid (70%) were purchased from Merck India Limited. Chloroform(d) and DMS (d) (for NMR) were purchased from Aldrich Company.

Methods: High Performance Liquid Chromatography: The Samples were analyzed on Alliance 2690 HPLC (Waters, Milford, MA, USA) system equipped with 2487 UVdetector. A Unisphere C18 column (150 mm x 4.6 mm i.d. 5 μm) was used for chromatographic separation. The mobile phase consisting of A: 1 cm³ perchloric acid (70%) in 1000 cm³ of water and B: acetonitrile, with timed gradient programmer Tmin /A: B: T0/85:15; T10/85:15; T30/50:50; T40/85:15; T45/85:15 with flow rate of 0.8 ml per minute were used. The column oven temperature was maintained at 30 °C. The injection volume was 20μL and the detector wavelength was fixed at 270 nm.

Liquid Chromatography-Tandem Mass Spectrometry (LC/MS/MS): The MS and MS/MS studies were performed on LCQ Advantage (Thermo Electron, San Jose, CA) ion trap mass spectrometer. The source voltage was maintained at 3.0 kV and capillary temperature at 250 °C. Nitrogen was used as both sheath and auxiliary gas. The mass to charge ratio was scanned across the various range. MS/MS studies were carried out by keeping normalized collision energy at 25-30% and an isolation width of 6 amu. The HPLC consisted of an Agilent-1100 series quaternary gradient pump with a degasser, an auto sampler and column oven. A C18 column (ProntoSIL Kromabond column 150 mm x 4.6 mm i.d. 5 μm) was used for separation. The mobile phase consisting of A: 1 cm³ Trifluoracetic acid in 1000 cm³ water and B: acetonitrile, with timed gradient programme Tmin/A: B: T0/85:15; T10/85:15; T30/50:50; T40/85:15; T45/85:15 with flow rate of 0.8 ml per minute were used.

NMR Spectroscopy: ¹H and ¹³C NMR spectra of the synthesized/isolated impurities were recorded on Bruker 400MHz instrument. The ¹H and ¹³C chemical shift values were reported on the δ scale (ppm) relative to CDCl₃ (7.26 ppm).

Preparative Liquid Chromatography: Impurities were isolated from the bulk sample using Waters Auto purification system consisting of 2525 binary gradient pump, a 2487UV detector and 2767 sample manager (Waters, Milford MA, USA). A Peerless Basic C18 column (150mm×21.2mm i.d., particle size 5μm) was used for the separation. The mobile phase was consisted of a mixture of water and acetonitrile in the ratio of 85:15 and was pumped at flow rate 25 cm³ /min. The detection was monitored at 270 nm.

Preparation of Solutions for Validation of HPLC Method: The test preparation solution of 500 μg/cm³ of SP bulk drug sample were prepared by using the diluents (mixture of 0.1% perchloric acid (75%) in water and acetonitrile, ratio is 85:15. A stock solution of mixture of impurities were prepared by dissolving 0.5mg/cm³ each of SM-I, SM-II, SM-III and SM. From this stock solution, a standard solution containing 0.5μg/cm³ each of SM-I, SM-II, SM-III and SM were prepared. This standard solution was also used for checking system suitability parameters.

RESULTS AND DISCUSSIONS:

Detection of Impurities by HPLC: Using HPLC analysis method are described and having the presence of four impurities at RRTs 0.20, 0.25, 0.54 and 1.34 with respect to principle peak. The target impurities under study are marked as Sulphanilamide used as a starting material, SM-I, SM-II, and SM-III, respectively. The typical chromatogram of crude SM sample highlighting the retention time of impurities.

Identification of Impurities by LC/MS/MS: The previous work of characterization is ideological to produce the mass data for the parent drug molecule so that, it may be easily compared and achieve, the process related impurities may be formed during the synthetic reaction. The spectra of SM exhibits a protonated molecular ion peak [M+H]⁺ 281 (Fig. 1) (molecular mass of SM is 280) in electro spray ionization in positive mode, the most plausible position of protonation was at NH₂ and NH. The MS/MS spectrum taken for the protonated SM molecule showed prominent peak at 156 (Fig. 2) which is due to  NH-SO₂ bond giving rise to C₆H₆NO₂S⁺ probable fragmentation are shown in (Fig. 3).

FIG. 1:  PLAUSIBLE SCHEME FOR FRAGMENTATIONS OF SM

SM-I showed a protonated molecular ion peak [M+H]⁺ 267 having molecular mass of 266, which under goes fragmentation to form C₆H₆NO₂S⁺ for 156 by loss of C₄H₅N₃O ion.

FIG. 2: PLAUSIBLE SCHEME FOR FRAGMENTATIONS OF SM-I

SM-II similarly showed [M+H]⁺ of 251 for molecular mass of 250 and a loss of C₄H₅N₃^•+ giving daughter ion of mass 156.

FIG. 3: PLAUSIBLE SCHEME FOR FRAGMENTATIONS OF SM-II

SM-III, which is an isomer, showed similar fragmentation that of SM. From all the mass fragmentation as discussed above, showed similar daughter ions of 156 for C₆H₆NO₂S⁺ which revealed that these impurities are structurally similar.

SM-I: showed a protonated molecular ion peak [M+H]⁺ 267 having molecular mass of 266, which under goes fragmentation to form C₆H₆NO₂S⁺ for 156 by loss of C₄H₅N₃O ion.

SM-II: similarly showed [M+H]+ of 251 for molecular mass of 250 and a loss of C₄H₅N₃ •⁺ giving daughter ion of mass 156.

SM-III: which is an isomer, showed similar fragmentation that of SM (Fig. 4). Since all the mass fragmentation as discussed above, showed similar daughter ions of 156 for C₆H₆NO₂S⁺ indicated that such impurities are structurally similar. SM-III and SM were having same molecular mass and may be Regio isomer of each other; later it was fixed to confirm the structure by NMR. Hence NMR of all the impurities and the product was carried out for comparison and further confirmation of structure. Since SM-III and SM were having same molecular mass and may be diasteriomer, hence it was mandatory to confirm the structure by NMR. Hence NMR of all the impurities and the product was carried out for comparison and further confirmation of structure.

FIG. 4: PLAUSIBLE SCHEME FOR FRAGMENTATIONS OF SM-III

Analytical Method Validation by HPLC: The validation study allowed the assessment of the process for its appropriateness for routine analysis. The new advanced system for SM and its related impurities was validated according to ICH guidelines ⁹. The validation study was accepted for the analysis of SM-I, SM-II and SM-III. The system appropriateness parameters obtained for related substance process are given (Fig. 5). Forced degradation studies were also performed (Acid, Base) for SM bulk drug sample to demonstrate the stability indicating power of the newly developed HPLC method.

FIG. 5: CHROMATOGRAM OF SYSTEM SUITABILITY SOLUTION

Specificity: Specificity is the capacity of analytical process to found the amount of analyte response in the existence of its potential impurities and degradants.  To specificity the HPLC technique for the determination of injecting individual impurity samples, wherein no interference was observed for any other components. The chromatograms were checked for the presence of any extra peak. Peak purity of these samples under stressed conditions was verified using a PDA detector. The purity of the principle and other chromatographic peaks was found to be acceptable. This study confirmed the stability indicating power of the HPLC method.

FIG. 6: A TYPICAL CHROMATOGRAM OF SM SAMPLE

FIG. 7(a): CHROMATOGRAM OF SM-I IN IDENTIFICATION

FIG. 7 (b): PDA PEAK PURITY SPECTRUM OF SM-I

FIG. 8 (a): CHROMATOGRAM OF SM-II IN IDENTIFICATION

FIG. 8 (b): PEAK PURITY PDA SPECTRUM OF SM-II

FIG. 9 (a): CHROMATOGRAM OF SM-III IN IDENTIFICATION

FIG. 9 (b): PEAK PURITY PDA SPECTRUM OF SM-III

Precision:  The ability of the method to precisely quantify impurities was determined by calculating the relative standard deviation (RSD) for response (peak area) of each impurity in the standard solution (a mixture of impurities) a copy of the injections. The R.S.D.’s for SM-I, SM-II, SM and SM-III were found to be 1.48%, 1.71%, 6.37% and 1.67%, respectively. These values are within the acceptance criteria of 10%. For confirming method precision six different test preparations of samples of SM were analyzed. The determined R.S.D. of these results was found to be under acceptable limit.

FIG. 10(a):  CHROMATOGRAM OF MIX STANDARD SOLUTION (INJECTION-01) IN PRECISION

FIG. 10(b): CHROMATOGRAM OF MIX STANDARD SOLUTION (INJECTION-02) IN PRECISION

FIG. 10 (C): CHROMATOGRAM OF MIX STANDARD SOLUTION (INJECTION-03) IN PRECISION

TABLE 1: EVALUATION DATA OF PRECISION STUDY

TABLE 2:  SYSTEM SUITABILITY REPORT IN PRECISION

Accuracy: For determining method accuracy, known as unclean LOQ, the specified limit of 80%, 100% and 120% of the SM bulk sample (test preparation) was spiked into. The unclean recovery was calculated individually.

FIG. 11:  CHROMATOGRAM OF MIX STANDARD SOLUTION OF ACCURACY

FIG. 12: CHROMATOGRAM OF PARENT SAMPLE IN ACCURACY

TABLE 3: ACCURACY OF IMPURITIES

Limit of Detection (DL) and Limit of Quantification (QL): Detection limit and quantitation limit for all impurities was estimated by signal to noise (S/N) method. The limit of detection values for SM-I, SM -II and SM -III were 0.002 %, 0.002 % and 0.002 % w.r.t. analyte concentration (500 μg /cm³) respectively. The limit of quantification values for SM-I, SM-II and SM-III were found to be 0.006 %, 0.006 %, and 0.006 % w.r.t. analyte concentration (500 μg /cm³), respectively.

FIG. 13: CHROMATOGRAM OF LOQ STANDARD SOLUTION

FIG. 14: CHROMATOGRAM OF LOD STANDARD SOLUTION

Linearity:  On drawing a plot at six concentration levels in triplicate, covering a range of LOQ to 150%, linear calibration plots were achieved. For SM-I, the corresponding regression equation was y = 28876x+348.97, with correlation coefficient (r) is 0.9996. For SM-II, corresponding regression equation was y = 52747x−0.2397, with correlation coefficient (r) is 0.9990. For SM-III, corresponding regression equation was y = 63972x+434.54, with the correlation coefficient (r) is 0.9983. The results showed a good correlation between the peak area and concentration of impurities.

TABLE 4: LINEARITY TABLE FOR SM-I IMPURITY

Correlation coefficient =0.9996

Slope =28876.00

Intercept =348.97

FIG. 15: LINEARITY CURVE FOR SM-I IMPURITY

TABLE 5: LINEARITY TABLE FOR SM-II IMPURITY

Correlation coefficient =0.9990

Slope =52746.63

Intercept =-0.24

FIG. 16: LINEARITY CURVE FOR SM-II IMPURITY

TABLE 6: LINEARITY TABLE FOR SM-III IMPURITY

Correlation coefficient = 0.9983

Slope =63972.03

Intercept =434.54

FIG. 17: LINEARITY CURVE FOR SM-III IMPURITY

Robustness: In all the deliberately varied chromatographic conditions (column temperature, flow rate and column make), the chromatogram for system suitability solution for related substance showed no significant change in system suitability parameters.

TABLE 8: CHANGE IN COLUMN TEMPERATURE [± 3 °C ]

* Data taken from Precision study

FIG. 18: CHROMATOGRAM OF SYSTEM SUITABILITY AT COLUMN TEMPERATURE 27 °C

FIG. 19: CHROMATOGRAM OF SYSTEM SUITABILITY AT COLUMN TEMPERATURE 33 °C

TABLE 9: CHANGE IN FLOW RATE  [± 0.2 cm³/min ]

* Data taken from Precision study

FIG. 20: CHROMATOGRAM OF SYSTEM SUITABILITY FLOW RATE 0.6 cm³ /min

FIG. 21: CHROMATOGRAM OF SYSTEM SUITABILITY FLOW RATE 1.0 cm³/min

TABLE 10: CHANGE IN COLUMN MAKES

* Data taken from Precision study

FIG. 23:  CHROMATOGRAM OF SYSTEM SUITABILITY USING PRONTO SIL KROMABOND COLUMN (150*4.6*5um)

FIG. 24: CHROMATOGRAM OF SYSTEM SUITABILITY USING PRONTO SIL KROMAPLUS COLUMN (150*4.6*5um)

FIG. 25: CHROMATOGRAM OF SYSTEM SUITABILITY USING KROMASIL C18 COLUMN (150*4.6*5um)

Solution Stability: The solution stability of SM sample and its related impurities was carried out by leaving both solutions in tightly capped HPLC vials at 25 °C for 16 hours in an auto sampler. No significant changes were observed in the area of impurities in standard solution after 16 hours.

FIG. 26: CHROMATOGRAM OF MIX STANDARD FRESHLY PREPARED IN SOLUTION STABILITY [0 HRS]

FIG. 27: CHROMATOGRAM OF TEST SOLUTION IN SOLUTION STABILITY [AFTER 16 HRS]

Isolation and structural elucidation of SM-I :  During the synthesis of SM i.e. from SCP to SM due to the basics condition of the reaction mass there is hydrolysis of methoxy group taking place which give rise to SM-I impurity which is then isolated by preparative HPLC (described in Section 2.3.6). 95% of chromatographic purity found.¹H and ¹³C NMR spectral data (refer Table 11) the proposed structure was confirmed. The MS / MS spectrum of the isolated Faux direct infusion mode using a combination of MS / MS spectrum was the same as the match SM-I (Refer Fig. 28 and 29).

Synthesis and structural elucidation of SM-II: Since SM-II cannot be isolated from the reaction mixture SM synthesis, it was synthesized independently. Due to the presence of 2-chloropyrazine as an impurity in 2,3-dichloropyrazine used as raw material in synthetic route of SM there is formation of SM-II which remains unreacted and get carry forward to SM final. This impurity was prepared synthetically by using the same synthetic route as that of SM but instead of 2,3-dichloropyrazine the starting material used was 2-chloropyrazine (Fig. 30). The chromatographic purity was found to be 97%. ¹H and ¹³C NMR spectral data (refer to Table 11) confirmed the proposed structure. Direct infusion of the compound synthesized from unclean condition using MS / MS spectrum was a match to the MS/MS spectrum of SM-II.

Synthesis and structural elucidation of SM-III: 2,6-dichloropyrazine which is isomer presence in 2,3-dichloropyrazine as an impurity under goes similar reaction of SM formed SM-III. SM-III is synthesized by using 2,6-dichloropyrazine instead of 2,3-dichloropyrazine in synthetic process of SM (Fig. 31). The chromatographic purity was found to be 96%. ¹H and ¹³C NMR spectral data (refer to Table 11) confirmed the proposed structure. Direct infusion of the compound synthesized from unclean condition using MS / MS spectrum was a match to the MS/MS spectrum of SM-III.

FIG. 28: MASS SPECTRUM OF SM P,   (b) ms/ms SPECTRUM OF SM

FIG. 29: (a) MASS SPECTRUM OF SM – I, (b) ms/ms SPECTRUM OF SM -I

FIG. 30: (a) MASS SPECTRUM OF SM-II, (b) ms/ms SPECTRUM OF SM-II

FIG. 31: (a) MASS SPECTRUM OF SM-III, (b) ms/ms SPECTRUM OF SM-III 

TABLE 11: NMR DATA OF SM-I, II, III AND SM

SM					SM-I				SM-II				SM-III
Posi tion	Integr ation	δ ppm	Multipli city J(H)z	13C &ppm	Integ ration	δ ppm	Multipl icity J(H)z	13C& ppm	Integra tion	δ  ppm	Multip licity J(H)z	13C& PPM	Inter griti on	δ ppm	Multip licity J(H)z	13C & ppm
1	2Ha	6.01	Brs	-	2Ha	4.39	Brs		2Ha	6.08	Brs		2Ha	6.08	Brs
2	-	-	-	153.3	-	-		151.6				153.8				151.6
3	2Ha	6.57	d(8.8)	112.6	2Ha	6.61	d(8.8)	116.6	2Ha	6.58	d(8.8)	153.9	2Ha	6.56	d(8.8)	116.6
4	2H	7.67	d(8.8)	130.2	2Ha	7.81	d(8.8)	128.1	2Ha	7.58	d(8.8)	129.8	2Ha	7.55	d(8.8)	128.1
5	-	-	-	125.6	-	-		129.1				124.5				129.7
6	-	-	-	-	-	-
7	1Hb	10.34	Brs	-	1Hb	11.7	Brs		1Hb	11.0	Brs		1Hb	11.11	Brs
8	-	-	-	149.9	-	-		152.7				7.8				154
9	1Ha	-	d(1.2)	138.8	1Ha	-		145.7	1Ha	7.8		7.8	1Ha	8.32	d(1.2)	123
10	-	-	-	-	-	-				7.8
11	1Ha	7.71	S	133.9	1Ha	6.05	d(4.3)	125.5	1Ha				1Ha	8.18	s	132.4
12	1Ha	7.71	S	133.6	1Ha	6.9	d(4.3)	125.5	1Ha				1Ha	8.18	s	161.1
13	-	--	-	--	-	-
14	-	-	-	-	1Hb	8.44	Brs
15	2Hb	3.89	S	54.1	-	-			3H	3.73	S	3.73				55.9

s-singlet, d-doublet, brs-broad singlet. ^a Refer the structural formula in Figure.^b1H-¹H coupling constants

CONCLUSION: The present study details the identification and determination of structure of four process related impurities are found in the product (SM). Reverse phase HPLC using a gradient of a newly developed method, the impurities were detected in samples of crude and refined SM during process development studies. An exhaustive study was carried out LC / MS / MS for the identification of impurities using. The spectroscopy using different technologies and their synthesis was followed by characterization. For a quantitative estimate of the contaminants in the paper elaborates on a new HPLC method validation. The specificity of the HPLC method was confirmed by injecting samples of individual impurity and it was observed that there was no interference for any of the components.

The ability of the method to precisely quantify impurities was determined by calculating the relative standard deviation (RSD) for response (peak area) of each impurity in the standard solution (a mixture of impurities) a copy of the injections. For determining method accuracy, known as unclean LOQ, the specified limit of 80%, 100% and 120% of the SM bulk sample (test preparation) was spiked into. Signal to noise (S/N)

method was used for Detection limit (DL) and quantitation limit (QL) for all impurities. On drawing a plot at six concentration levels in triplicate, covering a range of LOQ to 150 %, linear calibration plots were achieved. The stability of solution of SM sample and its related impurities was determined by placing both solutions in tightly capped HPLC vials at 25 °C for 16 hrs in an auto sampler. There are no significant changes were noted in the area of impurities in standard solution after 16 hours. But in sample solution area of SM-I have been increased and one unknown impurity at RRT 0.28 is generated after 16 hours. From stability data generated it is concluded that standard solution is stable for 16 hours and sample solution should be prepared freshly.

ACKNOWLEDGEMENT: The authors are thankful to Mukesh Kumar Gupta, Nitin Rathod from IPCA Laboratories and M.R. Chaudhary for their help and co-operations.

CONFLICTS OF INTEREST: Nil

REFERENCES:

1. Schwenger EM, Tejani AM and Loewen PS: Probiotics for preventing urinary tract infections in adults and children. Cochrane Database System Reviews2015; 23 (12):1-18
2. Penali L and Jansen FH: Single-day, three-dose treatment with fixed dose combination articulate /sulfa methoxypyrazine/pyrimethamine to cure Plasmodium falciparum malaria. International Journal Infectious Diseases2008; 12(4): 430-437.
3. Duke O, Yusuf A and Claire N: Derivatized Electrodes in the Electro analysis of Pyrimethamine/2-Sulfanilamido-3-Methoxypyrazine (Metakelfin) and Lumefantrine/ Artemeter (Coartem). Journal of chemical and chemical engineering 2014; 8: 215-225
4. Sweet man SC: The Complete Drug Reference. Pharmaceutical Press, London, 36^th edition 2009.
5. Storme ML, Jansen FH, Goeteyn W and Bocxlaer JVF: Simultaneous quantitative analysis of the antimalarials pyrimethamine and sulfa methoxypyrazine in plasma samples using liquid chromatography/tandem mass spectrometry., Rapid Communication of Mass Spectrometry 2006; 20: 2947-54.
6. Dua VK, Sarin R, Gupta NC and Sharma VP: Sulfalene concentrations in plasma and blood cells of Plasmodium falciparum malaria cases after treatment with metakelfin using high-performance liquid chromatography. Journal of Chromatography-B and Biomedical Sciences Applications 1998; 714: 390-98.
7. Narasimha R, Devanna KN and Suresh Reddy KVN: Method Development and Validation Study for Quantitative Determination of 3-Ethynylaniline Content in Erlotinib by Liquid Chromatography–Tandem Mass Spectrometry. Indian Journal of Advances in Chemical Science 2016; 4(2): 208-213.
8. Lokhande MV, Gupta MK and Rathod NG: Characterization and Quantitative Determination of Process Related Impurities in Sulfa methoxypyrazine By Some Analytical Techniques. International Journal of Medicine and Pharmaceutical Sciences, 2013; 3(4): 37-48.
9. ICH Q3D Guideline, Impurities in New Drug Substances, Step- 5 version 2014.
10. Turin VG, Oranto G: Buracchi, United States Patent 4299962.1981.
11. Thummala V, Raghava R,  Noru AK,Seshadri RK, Annarapu MR,   and  Ivaturi MR: Development and Validation of a Stability-Indicating RP-HPLC Method for Simultaneous Estimation of Dextromethorphan Impurities in Pharmaceutical Formulations. Chromatography Research International 2013; 2013:1-12.
12. Primer: Pharmaceutical impurities analysis solution, Agilent technology 2015.
13. Blessy M, Patel RD, Prajapati PN and Agarwal YK: Development of forced degradation and stability indicating studies of drugs—A review.Journal of Pharmaceutical Analysis 2014; 4(3): 159–165.
    

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    How to cite this article:

    Maurya CP and Lokhande MV: Characterization and validation of impurities related to pharmaceutical bulk drug (API) by using some analytical techniques. Int J Pharm Sci Res 2017; 8(8): 3325-40.doi: 10.13040/IJPSR.0975-8232.8(8).3325-40.

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