25 C 290 nm 25 C 290 nm40C 290 nm 40 C 290 nm 40 C 290 nm Observations and

25 c 290 nm 25 c 290 nm40c 290 nm 40 c 290 nm 40 c

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25 °C & 290 nm 25 °C & 290 nm 40°C & 290 nm 40 °C & 290 nm 40 °C & 290 nm Observations and corrections to next trial Trial No. 1 Base line is good but the retention of the sample in the column was too much, fronting, no resolution between the peaks, buffer should be used within 10mM to 50mM with slightly basic pH within the range of Pka value to elute the sample early, stable bond column is needed. Trial No. 2 Column was changed to stable bonded C8, reduction in fronting, base line is good, no peak resolution, buffer used to stabilize the pH, peak elution is too early. Trial No. 3 Base line is good, resolution between the sulfone and PNT peak is more than 2 but no peak resolution to others. Trial No. 4 Baseline is good, column was changed to stable bonded C18, gradient flow is used to increase the resolution between the peaks but still no resolution between DBT and Sulfone. Trial No. 5 Base line is good, peak resolution between DBT and Sulfone & PNT and sulfide are not good, gradient should be changed. Trial No. 6 No interference of any peak with the main peak and to each other, resolution between the sulfone and Pantoprazole peak is more than 2. MP- Percentage mobile phase, NHP- Disodium hydrogen phosphate, KHP- Disodium hydrogen phosphate, OPA - Orthophosphoric acid, ACN- HPLC grade acetonitrile, TEA- triethylamine. Our preliminary trials using different columns (Phenomenex Luna C8, Zorbax SB C8), different mobile phases consisting of water-acetonitrile and sodium phos- phate buffer- acetonitrile using isocratic elution did not give good peak shape and resolution. For the impurities and PNT, retention times were found near dead point. Therefore, in such case gradient elution is preffered over isocratic. To obtain efficient gradient elution pattern, small linear gradient was employed for development. Several, permutations and combination of ACN-buffer and buffer- ACN with varying P H have been scanned to finalize the components of mobile phase. It has been observed that ACN (mobile phase-A) and phosphate buffer adjusted to pH 7.0 with orthophosphoric acid (mobile phase-B) with gradient elution provide best peak shape and resolution. The diluent preffered is 0.1M sodium hydroxide: ACN (50:50). Hypersil ODS a C18 column is preferred over C8 column because of its hydrophobicity so that to rec- tify retention problems. Table I includes all the trials for optimization of method. Forced degradation studies In forced degradation method pantoprazole sodium was found to degrade substantially under acidic, oxidative conditions while stable under alkaline, thermal condition and under photolytic conditions was miniscule (Table II). The major impurity under acidic and oxidative condition was Sulfide (RRT 1.63) and Sulfone (RRT 0.80) respec - tively (Figure 2). Method validation PNT and its related impurities were well resolved with no interference from the blank and mobile phase. For all compounds the purity angle was found to be less than
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S. Pandey, P. Pandey, D. Mishra, U. K. Singh 180 purity threshold value. The value of relative retention time and purity data is listed in table II. The described method
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