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British Journal of Pharmaceutical Research, ISSN: 2231-2919,Vol.: 14, Issue.: 4


Development and Validation of Bio Analytical Method for Estimation of Bortezomib in k3 EDTA Human Plasma Using HPLC-ESI-MS/MS and Its Application to a Bioequivalence & CME Studies


Ravi Pratap Pulla1*, K. Vanitha Prakash1, U. B. Abhini1, B. Naga Maheswari1, V. Divya1, M. V. Shushmitha1 and V. Ravali1

1SSJ College of Pharmacy, V. N. Pally, Gandipet, Hyderabad-75, Telangana State, India.

Article Information
(1) Rahul S. Khupse, Pharmaceutical Sciences, University of Findlay, USA.
(1) Marina Quartu, University of Cagliari, Italy.
(2) Tommasina Guglielmelli, San Luigi Hospital, Italy.
Complete Peer review History: http://www.sciencedomain.org/review-history/17605


Purpose: To develop a highly selective, reproducible & precise rugged bio analytical method for estimation of Bortezomib (BTZ), “A Protease Inhibitor” in human plasma by validating the developed method in accordance to US-FDA guidelines.

Methodology Envisaged: BTZ D3 was used as an internal standard (ISTD) for the determination of BTZ in human plasma using a rapid & specific liquid chromatographic – Electron Spray Ionization –Mass spectrometric method. The analytical method was moduled with liquid-liquid phase extraction by using annular centrifugal contactor & the samples were analyzed by HPLC, on a column - ACE 5CN (150 x 4.6 mm 5 µm), using mobile phase consisting of ammonium formate buffer: ACN (25:75 v/v), delivered at 1.0 ml/min & 90% flow spitting. Applied Bio system MDS Sciex API 3000 Triple Quadruple MS equipped with Turbo Ion Spray (TIS) as LC/MS interface was used in for MS detection. TIS with multiple reaction monitoring (MRM) were acquired by ESI mass spectra, using the transitions m/z 362.95→310.21 & m/z 172.64→146.06 to quantify BTZ & BTZ D3 respectively.

Results: % variability was ≤ 5.52 & ≤ 6.15 [that was ≤ 15], indicating the specificity of the method, showing no matrix interferences across the elution system. Acceptance is ranging between -8.30 to 2.83 & -4.32 to 1.00% (< 5% CV) & accuracy in the range of 92.73 – 102.20 (< 10% difference) was observed over a linear range of 2.00 – 1000 ng/mL. The mean (n=3) correlation coefficient was 0.9991 & overall mean recovery was 85.62%. Retention time for drug & ISTD is found out to be 0.08 & 0.07; % CV of area ratio is 1.91% & area ratio ≤ 2.51%, which indicated system suitability.

Interpretation and Conclusion: The intended analyte is stable below 10ºC in all the performed stability experimentation & within the acceptance limits. It can be used for investigating drug concentration in routine quality control analysis in API & its pharmaceutical dosage forms.

Keywords :

Bortezomib; Bortezomib D3; method validation; HPLC-ESI-MS/MS; human plasma; multiple reaction monitoring.

Full Article - PDF    Page 1-17

DOI : 10.9734/BJPR/2016/30565

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