Quantitative Application of NMR In Ropivacaine ...

1 International Journal of ChemTech Research CODEN( USA): IJCRGG ISSN : 0974-4290. , , pp 312-321, Jan-Mar 2013. Quantitative Application of NMR In Ropivacaine Hydrochloride And Its Related Impurity-A With Correlation By Alternate Techniques Kokila Parmar*, Anilkumar Mahato, Rinku Patel and Sarju Prajapati Department of Chemistry, Hem. North Gujarat University, Patan- 384265, Gujaarat, India. * : Abstract: The present work is Application of Quantitative NMR is increasingly used as a practical tool with respect to elucidation of structure and Quantitative analysis. In the present study, we show the ability of 1H NMR.

2 To determine the content of drug and its related impurity-A in Ropivacaine hydrochloride. Rapid, specific and accurate proton NMR method developed for the drug Ropivacaine hydrochloride using Internal standard. For the drug quantification, proton signal at and for Tetrachloronitrobenzene (TCNB) were used. Assay method for proton NMR were validated for specificity, precision and intermediate precision, linearity, solution stability and Robustness. Impurity profiling of impurity-A (2,6 dimethyl aniline) were also developed and validated for specificity and precision. LOD and LOQ were also established.

3 Keywords: NMR, Ropivacaine Hydrochloride,Related Impurity Impurity-A of Ropivacaine hydrochloride, Quantitative Application . Introduction analytic. The determination is based on the ratio of the integration of a specific signal of the analyte and Ropivacaine hydrochloride is (2S)-N-(2,6- the internal reference standard. dimethyl-phenyl)-1-propyl-2-piperinecarb oxamide is The aim of the present study is to determine an anesthetic drug. The drug act by blocking the the assay of Ropivacaine by Quantitative NMR. conduction of impulses in the target nerve structures, spectroscopy using internal standard method and primarily located within the subarachinoid space [1- related impurity 2,6-dimethylaniline , a genotoxic 3].

4 The drug reported in the Literature survey reveals impurity by area integral method [10-13]. that the reported methods were HPLC determination in USP2. Several methods using GC [4], HPLC [5, Experimental 6] and HPLC-MS [7] or amperometric detection [8]. and capillary electrophoresis [9] have been High purity analytical grade substances were developed to analyse the drug and their impurities in used. Authentic sample of Ropivacaine was obtained pharmaceutical formulation or biological fluids. The from Local pharmaceutical company and used as present study uses the proton Nuclear Magnetic such.

5 Internal standard used was resonance Spectroscopy (NMR) to quantify Tetrachloronitrobenzene ( , Methanol-d4. Ropivacaine and its related impurity 2, 6- ( ) were purchased from Merck. dimethylaniline NMR is a Quantitative spectroscopy tool as the intensity of resonance line is proportional Instrumentation to the number of resonant nuclei. This principle is NMR instrument used was Bruker Make employed for the Quantitative determination of Avance-400 operating at ( ) for compounds. The most advantage of Q-NMR is that proton equipped with a 5mm multinuclear broad the determination does not require standard of the band observe (BBO) probehead.)

6 Kokila Parmar et al ,5(1) 313. Structure interpretation of drug and Impurity by Where, NMR, IR spectroscopy and Mass spectrometry It Integration of the test sample NMR Parameters Is Integration of the standard sample NMR analysis was performed on Ws Weight of the standard sample Ropivacaine sample measured at 400 MHz Wt Weight of the test sample Spectrometer. Typically 16 scans were collected for Mt Molecular weight of the Test sample assay and 64 scans for related impurity into 32768 Ms Molecular weight of the Standard sample data points using 90 pulse length, spectral width of Ns Number of Standard sample protons , digital resolution of Nt Number of test sample protons , dead time of 6 s and acquisition P Strength / assay of the standard sample time of s.

7 A delay time of 20s was used for Ropivacaine determination and impurity Specificity of NMR Method : determination which was sufficient to relax the The available related compound or impurity protons. The FID were apodised with 1Hz has to be sufficiently well separated from those of exponential line broadening function before Fourier the main drug and other impurities to allow Transformation. Automatic phase correction and Quantitative NMR. Sometimes when the impurities baseline correction were employed. merge with spinning side bands it becomes important to run the sample using different spinning Procedure for determination (assay) of drug : rates or without spinning.

8 Following table gives the Weigh and transfer accurately about 10mg chemical shift values of impurity A and the main to 15mg each of of Ropivacaine hydrochloride compound. sample and TCNB into a 10ml glass vial, to it is First the identification were done for the added of MeOD and sonicated to dissolve. Ropivacaine and impurity A by proton NMR, The solvent is chosen during method development Carbon 13 NMR, FTIR and Mass spectrometry. such that it does not exhibit resonance peaks that Chemical name overlap those of the sample being analyzed. The 1-propylpiperidine-2-carboxylic acid (2,6-dimethyl solvent should have good solubility properties and phenyl)amide hydrochloride monohydrate does not exhibit resonance peaks which overlap Chemical Formula C17H27 ClN2O.

9 Those of the specimen being analyzed. Normally, Molecular weight g/mole TMS or similar is incorporated in the solvent as a chemical shift reference. 1. H NMR analysis 1. H NMR spectral analysis of impurity-A, Formula of assay / content is was performed on NMR spectrometer of Bruker 400. % assay , w/w UltraShield, model-Avance-II, in Methanol-d6, = It x Ns x Mt x Ws x P. using Tetramethylsilane as an internal standard. Is x Nt x Ms x Wt Table 1 Observation Impurity-A of Ropivacaine hydrochloride by 1H NMR spectroscopy Chemical shift, , ppm Multiplicity, proton number Assignments singlet, 6H Methyl protons at C-2, C-6.

10 , Doublet, 2H Aromatic protons at C-3, C-5. Triplet, 1H Aromatic protons at C-4. 3 2. 1. 4 NH2. 5 6. Kokila Parmar et al ,5(1) 314. ppm Fig. 1 Identification Impurity-A of Ropivacaine hydrochloride by 1H NMR Spectroscopy Remark protons at C-1, C-2 and C-3 appear as multiple at From the Table and Figure 1 observation Methine protons C-7 appear at and discussion, it is concluded that the chemical , while methylene protons at C-12, C-11, shift in the 1H NMR spectrum of Impurity-A are in C-13, C-10, C-8, C-9 appear at , accordance with the position and the proton number , and respectively.